PhD Thesis
Interstitial changes in trapezius muscle during
repetitive low-force work
- Studies with microdialysis in healthy subjects and patients with
work-related trapezius myalgia
Lars Rosendal
2004
National Institute of Occupational Health, Denmark
- Interstitial changes in trapezius muscle during repetitive low-force work -
PREFACE
This PhD project was initiated by senior researcher, PhD, Karen Søgaard at the
National Institute of Occupational Health (NIOH), Copenhagen, Denmark, as part of
the research strategy aiming at increasing our knowledge about the underlying
mechanisms behind work-related muscle pain due to repetitive low-force work. The
studies presented in this thesis have either been approved by the ethical committee of
Copenhagen and Frederiksberg, Denmark (KF 01-152/01), or the ethical committee of
Linköping University, Sweden (Dnr. 03-031).
Mentorship was provided by senior researcher, PhD, Jesper Kristiansen, NIOH, as well
as professor, Dr.med, Michael Kjær and senior researcher, PhD, Henning Langberg,
both from the Institute of Sports Medicine Copenhagen, Bispebjerg University
Hospital, Denmark.
The project was co-financed by the National Institute of Occupational Health and the
Danish Research Agency (641-01-0035).
Copenhagen, March 31st 2004
Lars Rosendal
Thesis submitted: April 1st 2004
Thesis defended: November 25th 2004
Evaluation committee:
Associate professor, Bente Stallknecht (head)
Department of Medical Physiology, The Panum Institute, Denmark
Professor, Urban Ungerstedt
Department of Physiology and Pharmacology, The Karolinska Institute, Sweden
Professor, Nina Vøllestad
Institute of Nursing and Health Sciences, University of Oslo, Norway
2
- Lars Rosendal -
CONTENT
PREFACE .............................................................................................................................2
CONTENT.............................................................................................................................3
LIST OF PAPERS.................................................................................................................5
SUMMARY (ENGLISH) ......................................................................................................7
RESUME (DANSK) ..............................................................................................................9
LIST OF ABBREVIATIONS..............................................................................................11
1. INTRODUCTION ...........................................................................................................13
1.1 PREVALENCE AND RISK FACTORS OF WORK-RELATED MUSCULOSKELETAL DISORDERS ...13
1.2 TRAPEZIUS MUSCLE .....................................................................................................14
1.3 PROPOSED PATHOMECHANISMS BEHIND MYALGIA FROM REPETITIVE, LOW-FORCE WORK 15
1.4 MUSCLE PAIN AND NOCICEPTION ..................................................................................19
1.5 MICRODIALYSIS ...........................................................................................................20
2. AIM..................................................................................................................................23
2.1 HYPOTHESIS ................................................................................................................23
3. METHODS ......................................................................................................................25
3.1 PARTICIPANTS .............................................................................................................25
3.2 PROTOCOLS .................................................................................................................25
3.3 OVERVIEW OF USED METHODS AND EXERCISE PROTOCOLS .............................................26
3.4 EXERCISE PROTOCOLS ..................................................................................................27
3.5 MICRODIALYSIS ...........................................................................................................28
3.6 ELECTROMYOGRAPHY .................................................................................................30
3.7 BIOCHEMICAL MEASUREMENTS ....................................................................................31
3.8 RPE, VAS, AND ALGOMETRY ......................................................................................32
3.9 CLINICAL EXAMINATION AND QUESTIONNAIRE ..............................................................33
3.10 STATISTICS ................................................................................................................33
4. RESULTS ........................................................................................................................35
4.1 RESPONSE TO THE INSERTION TRAUMA, HEALTHY SUBJECTS ..........................................35
4.2 RESPONSE TO RLW AND SUS, HEALTHY SUBJECTS .......................................................35
4.3 RESPONSE TO RLW, PATIENTS WITH WORK-RELATED TRAPEZIUS MYALGIA AND HEALTHY
CONTROLS .........................................................................................................................39
3
- Interstitial changes in trapezius muscle during repetitive low-force work -
5. DISCUSSION ..................................................................................................................43
5.1 MAIN FINDINGS ...........................................................................................................43
5.2 APPLICABILITY OF MICRODIALYSIS IN TRAPEZIUS MUSCLE .............................................43
5.3 MUSCLE RESPONSE TO INSERTION TRAUMA ...................................................................44
5.4 MUSCLE METABOLISM DURING RLW AND DURING INTENSE STATIC CONTRACTION ........45
5.5 CYTOKINE RESPONSE TO RLW .....................................................................................47
5.6 MUSCLE PAIN AND ALGESIC SUBSTANCES IN TRAPEZIUS MYALGIA .................................49
7. CONCLUSION................................................................................................................51
8. PERSPECTIVE ...............................................................................................................53
ACKNOWLEDGEMENTS.................................................................................................55
REFERENCE LIST ............................................................................................................57
PAPER I
PAPER II
PAPER III
4
- Lars Rosendal -
LIST OF PAPERS
The present thesis is based on three papers.
They will be referred to in the text by Roman numerals.
I.
Rosendal, L., Blangsted, A. K., Kristiansen, J., Søgaard, K., Langberg, H., Sjøgaard,
G., & Kjær, M. (2004). Interstitial muscle lactate, pyruvate, and potassium
dynamics in the trapezius muscle during repetitive low-force arm movements,
measured with microdialysis. Acta Physiol Scand In press.
II.
Rosendal, L., Søgaard, K., Kjær, M., Sjøgaard, G., Langberg, H., & Kristiansen, J.
(2004). Increase in interstitial interleukin-6 of human skeletal muscle with
repetitive low-force exercise. J Appl Physiol In Press,
doi: 10.1152/japplphysiol.00130.2004.
III.
Rosendal, L., Larsson, B., Kristiansen, J., Peolsson, M., Søgaard, K., Kjaer, M.,
Sörensen, J., & Gerdle, B. (2004). Increase in muscle nociceptive substances and
anaerobic metabolism in patients with trapezius myalgia: microdialysis in rest and
during exercise. Pain 112, 345-355
5
- Interstitial changes in trapezius muscle during repetitive low-force work -
6
- Lars Rosendal -
SUMMARY (ENGLISH)
Introduction
Chronic pain in the musculoskeletal system due to working life conditions is an
important socio-economic problem in the industrialized world. Muscle pain is frequent
in the upper trapezius muscle in occupational groups employed with highly repetitive
work tasks. Even though the problem is widely recognized, the underlying mechanisms
behind the development of work-related muscle pain are not well understood. Several
pathomechanisms have been proposed and although they differ, they have one
important feature in common, which is that muscle nociceptors are activated or
sensitized by metabolites and algesics released into the interstitial space in response to
repetitive low-force work.
The aim of the present thesis was to determine the interstitial trapezius muscle
responses to repetitive low-force work in healthy subjects and in patients with workrelated trapezius myalgia – with respect to metabolism, inflammatory mediators and
potential algesic substances.
Methods
The microdialysis technique was used to study the interstitial muscle:
- Cytokine and muscle damage response to the insertion trauma in healthy
participants.
- Metabolic and cytokine responses to repetitive low-force work and to intense
static shoulder flexion in healthy participants.
- Metabolic, blood flow and algesic response to repetitive low-force work in
patients with work-related trapezius myalgia and in healthy controls.
Main findings and conclusions
The present thesis demonstrated that the microdialysis technique enabled measurement
of low and high molecular weight substances related to metabolism, cytokine response,
and algesics in trapezius muscle in response to repetitive low-force work.
It was demonstrated that metabolites accumulate in the trapezius muscle during 20 min
of repetitive low-force work. An increase in muscle lactate was found indicating that
7
- Interstitial changes in trapezius muscle during repetitive low-force work -
anaerobic metabolism is accelerated, even though the muscle activity level was below
10 % of max.
The cytokine interleukin-6, which has been speculated to have important metabolic and
anti-inflammatory properties, increased substantially during repetitive low-force work
and the increase could only to a minor degree be explained by the insertion trauma per se.
Finally, and perhaps most important of all, work-related trapezius myalgia was shown
to be associated with increased anaerobic metabolism as well as increased levels of
potential algesic substances (serotonin, glutamate) locally in the painful muscle tissue
– indicating that peripheral nociceptive processes could be activated. These changes in
metabolites and algesics were not associated with a reduced blood flow response in the
patients with trapezius myalgia.
8
- Lars Rosendal -
RESUME (DANSK)
Introduktion
Kroniske muskelsmerter, opstået i forbindelse med erhvervsarbejde, er et stort problem
og udgør en betydelig økonomisk byrde hvad angår sygdomsfravær, sundhedsudgifter
og førtidspensioner. En væsentlig del af disse muskelsmerter forekommer i skuldernakke muskulaturen (trapezius) efter længerevarende statisk eller ensidig gentaget
arbejde (EGA). Til trods for en generel anerkendelse af problemet er årsagen til
udvikling af arbejdsrelaterede muskelsmerter fortsat uvis. Der findes flere hypoteser
om hvordan muskelbesvær i forbindelse med EGA udvikles, og et fællestræk ved
hypoteserne er at der som følge af EGA sker en frigivelse af metabolitter og
smertemedierende substanser til den interstitielle væskefase hvor smertereceptorer
aktiveres eller sensibiliseres.
Formålet med denne Ph.d.-afhandling er at bestemme det lokale (interstitielle)
trapezius muskelrespons på EGA i raske individer og i patienter med arbejdsbetinget
skulder-nakke besvær – specielt med hensyn til metabolisme, pro og anti-inflammatoriske
mediatorer og potentielle smertemedierende substanser.
Metoder
For at undersøge det lokale muskelrespons blev mikrodialysekatetre indlagt i trapeziusmusklen for at studere:
- det inflammatoriske respons på traumet i forbindelse med kateterindlæggelsen,
hvilket blev undersøgt i raske individer.
- det metaboliske og inflammatoriske respons på EGA og på intens statisk
arbejde, hvilket blev undersøgt i raske individer.
- metabolismen, blodgennemstrømningen og koncentrationen af potentielle
smerte-medierende substanser, målt i hvile og under EGA i patienter med
arbejdsbetinget skulder-nakke besvær og i raske kontroller.
Resultater og konklusioner
Studierne viste at mikrodialyse teknikken kan anvendes på trapezius musklen i hvile og
under forskellige arbejdstyper. Specielt kan metoden med fordel bruges til at studere
ændringer i intramuskulære substanser ved lette belastninger, som ikke afspejles i
blodbanen.
9
- Interstitial changes in trapezius muscle during repetitive low-force work -
Det blev fundet at metabolismen i trapeziusmusklen i væsentlig grad påvirkes af 20
min EGA (muskelbelastning ca. 10% af max). Til trods for den lave muskelbelastning
påvistes laktatkoncentrationen således at stige under EGA, hvilket indikerer en øget
anaerob energiomsætning. Desuden blev en særdeles stor stigning i interleukin-6 påvist
som følge af EGA. Denne stigning kunne kun i mindre grad tilskrives selve traumet.
De vigtigste fund er dog at den anaerobe energiomsætning er væsentlig forøget, både i
hvile og under EGA, i patienter med arbejdsbetinget skulder-nakke besvær, hvilket
ikke kunne tilskrives en lavere blodgennemstrømning. Desuden var koncentrationen af
potentielt smertemedierende substanser væsentligt forøget i patientgruppen, hvilket
indikerer at muskelsmerten til dels kan skyldes en lokal, perifer smertereceptoraktivering.
10
- Lars Rosendal -
LIST OF ABBREVIATIONS
5-HT
EMG
IL-1
IL-6
ISF
K+
MU
MVC
PPT
RLW
RR
RPE
SUS
VAS
WTM
: Serotonin (5-hydroxytryptamine)
: Electromyography
: Interleukin-1 beta
: Interleukin-6
: Interstitial fluid
: Potassium
: Motor unit
: Maximum voluntary contraction
: Pressure pain threshold
: Repetitive low-force work
: Relative recovery
: Rating of perceived exertion
: Sustained static shoulder flexion
: Visual analogue scale
: Work-related trapezius myalgia
11
- Interstitial changes in trapezius muscle during repetitive low-force work -
12
- Lars Rosendal -
1. INTRODUCTION
Chronic pain syndromes within the musculoskeletal system due to working life
conditions are an important socio-economic problem in the industrialized world.
A total of 39 % of the reported work-related diseases in Denmark are of
musculoskeletal origin (Westgaard & Winkel, 1997) and, the number of work-related
musculoskeletal disorders associated with “repeated trauma” in the USA numbered
240.000 cases in 2000 (Bureau of Labor Statistics, 2001). Thus the number of people
affected is enormous, causing a negative impact on society. This impact includes a
reduction in workforce and income as well as increased costs for medical treatment,
sick leave and early retirement, and the cost in the Nordic countries and Holland is
estimated to be between 0.5 and 2% of the Gross National Product (Johansson et al.,
2003b). Work-related musculoskeletal disorders are particularly frequent in the
trapezius muscle, primarily in occupational groups employed with highly repetitive
work tasks. Even though the problem is widely recognized, the underlying mechanisms
behind the development of work-related muscle pain are not well understood. Both
peripheral as well as central mechanisms have been proposed (Edwards, 1988;
Henriksson, 1988;Hägg, 1991;Sjøgaard & Søgaard, 1998;Gissel, 2000;Johansson et
al., 2003a;Barr & Barbe, 2004).
1.1 Prevalence and risk factors of work-related musculoskeletal disorders
Work-related musculoskeletal disorder is defined as a multi-factorial concept that
includes work exposure as a significant contributor to the development (WHO, 1985).
Myalgia refers to pain that originates from muscle tissue, and chronic myalgia is the
single largest category of work-related illnesses in USA, Japan and the Nordic
countries (Sjøgaard et al., 1995;Bernard, 1997). Of these disorders, myalgia in the
shoulder-neck region is predominant, and the most frequent site for myalgia is the
upper (descending) part of the trapezius muscle (Johansson et al., 2003b).
Work-related trapezius myalgia (WTM) often worsen during the course of the working
day (Hagberg, 1996), and the person suffering from WTM is often initially free of pain
during leisure time but the pain gradually increases to being present at all time.
Epidemiological studies have shown that self-reported shoulder-neck myalgia for more
than 30 days in the last year, is prevalent in many occupations that involve repetitive
job tasks (Jensen et al., 1998;Fredriksson et al., 2000;Jensen, 2003). Furthermore, a
recent systematic review reports evidence that highly repetitive work and forceful arm
or hand movements cause neck and shoulder disorders, and strong evidence that work
13
- Interstitial changes in trapezius muscle during repetitive low-force work -
activities involving prolonged static loads on the neck and shoulder muscles increase
neck and shoulder disorders (Bernard, 1997). Epidemiological research supports that
both physical and psychosocial factors related to work could play a role in the
development of these disorders. However, the evidence for an etiologic role is thought
to be stronger for the former than for the latter (Johansson et al., 2003b).
1.2 Trapezius muscle
The anatomic structures in the shoulder girdle interact in a complex manner.
The movement of the shoulder occurs in a coordinated action of the gleno-humeral
joint and scapula and many muscles interact. The trapezius muscle has several
important actions in the shoulder muscle function, especially as a stabilizer of the
scapula in hand and arm movements, and as a stabilizer of the neck for adequate visual
precision. The trapezius muscle is a large, flat, triangle-shaped muscle that extends
from the back of the scull, down to the lower thoracic region and from the spine to the
acromion. The trapezius muscle is divided into three parts, exerting their separate
actions – the upper (descending) part, the middle (horizontal) part and the lower
(ascending) part (Figure 1). The upper part was the object for the present investigation
and it has the following main functions; a) raise the shoulder against gravity, b) lateral
flexion of the neck, c) head rotation, and d) neck hyper extension (when contracted
bilaterally).
Figure 1 Schematic drawing of the trapezius muscle.
Reprinted with permission from (Lindman, 1992).
14
- Lars Rosendal -
Surface electromyography (EMG) amplitude from the upper trapezius is widely used
as a measure of the shoulder-neck load during occupational work. The EMG activity
level of the upper trapezius muscle during occupational work of repetitive character
(repetitive low-force work, RLW) has been reviewed (Jensen et al., 1999) and
demonstrated to be approximately 15 % EMG max during assembly work and sewing
machine operations (Christensen, 1986;Jensen et al., 1993b), approx. 7 % EMGmax in
cleaners (Nordander et al., 2000), and 4-6 % EMGmax in office workers and computer
operators (Nordander et al., 2000;Blangsted et al., 2003).
1.3 Proposed pathomechanisms behind myalgia from repetitive, low-force
work
A substantial amount of money has been put into research but despite that, the
advances in methods for the prevention, treatment and rehabilitation of chronic workrelated myalgia is lacking. Recently, it was proposed that this is due to a lack of
understanding of the basic pathophysiological mechanisms behind chronic muscle pain
and that such knowledge is a prerequisite for efficient prevention, treatment and
rehabilitation (Johansson et al., 2003b).
It has been proposed that nociceptors may be triggered through the release of different
metabolic products related to muscle contraction (MacLean et al., 1998;Sjøgaard et al.,
2000) and this section summarizes some of the proposed pathomechanisms.
1.3.1 The Cinderella Hypothesis
One characteristic feature of low-force repetitive work is that it implies low force
requirements as described above. Inhomogeneous activation of a muscle exerting low
forces is a characteristic that may be considered a risk factor for the development of
work-related muscle pain (Sjøgaard et al., 2000). The functional unit in muscles is a
motor unit (MU), consisting of one motorneuron and the muscle fibers it innervates
(between a few and thousands). It is well-known that muscular tissue is recruited in a
hierarchical manner according to the size principle, starting with the MU with the
lowest threshold (Henneman & Olson, 1965;Zajac & Faden, 1985). This has led to the
development of the Cinderella hypothesis by Hägg; stating that during prolonged lowlevel contraction, a few MU will become fatigued or exhausted and thereby be
relatively overloaded despite that the muscle as a whole is working at a low energy
demand (Hägg, 1991). Indeed, so-called Cinderella fibers have been identified - a
stereotype recruitment pattern is evident during static as well as dynamic contractions
(Søgaard, 1995;Sjøgaard & Søgaard, 1998). The continuous activity of a subset of
15
- Interstitial changes in trapezius muscle during repetitive low-force work -
muscle fibers will involve a high local energy turnover and may result in a localized
increase in the intramuscular pressure around the fibers, thus impeding blood flow to
the parts of the muscle that need the oxygen most. This would turn metabolism towards
a more anaerobic state resulting in increased lactate levels and perhaps decreased
pyruvate levels. In support of this hypothesis, biopsy studies on subjects with workrelated myalgia have indicated various structural changes and mitochondrial
disturbances indicating disturbed metabolism. In subjects with trapezius myalgia, an
increased frequency of type-I fibers as well as increased size has been demonstrated
(Kadi et al., 1998), indicating a load-induced hypertrophy (Larsson et al., 1988).
Furthermore, the existence of type-I muscle fibers with a ragged appearance and
increased subsarcolemmal mitochondrial damage (ragged red fibers) and a reduced
capillarization per fiber cross-sectional area has been demonstrated (Larsson et al.,
1988;Kadi et al., 1998;Larsson et al., 2000b), which could be indicative of a disturbed
energy turnover (Kadi et al., 1998;Hägg, 2000;Larsson et al., 2004). These
morphological changes have been suggested to be associated with a reduced blood
flow and thereby ischemia in the muscle during work (Travell JG et al.,
1942;Johansson et al., 2003b). In support of this, the blood flow in the upper trapezius
has been shown to be reduced in patients with myalgia (Larsson et al., 1999). Thus
there seems to be some chronic changes in muscle characteristics, however, whether
occupational RLW is associated with in-vivo increased anaerobic metabolism locally
in the muscle tissue has not been investigated. Neither has it been investigated whether
a reduced muscle blood flow in trapezius myalgia is associated with increased
metabolite accumulation locally in muscle tissue during RLW, indicating a disturbed
energy turnover.
The morphological changes in muscle tissue from patients with work-related
musculoskeletal disorders have also been suggested to be due to inflammatory events
locally in the tissue due to microruptures from repeated motions (Barr & Barbe, 2004).
Cytokines are classical markers of inflammatory activity. The cytokines interleukin-1
beta (IL-1) and interleukin-6 (IL-6) have previously been considered proinflammatory and associated with disease (Waage & Steinshamn, 1993), and have also
been suggested to associated with muscle damage in response to exercise (Bruunsgaard
et al., 1997). Recent data, however, suggest that in response to exercise, IL-6 is
produced locally in muscle tissue (Ostrowski et al., 1998;Steensberg et al., 2000) and
connective tissue (Langberg et al., 2002b), without the presence of muscle damage
(Brenner et al., 1999). Furthermore, IL-6 may primarily posses anti-inflammatory
characteristics and may regulate metabolic activity during exercise (Pedersen et al.,
16
- Lars Rosendal -
2001b;Pedersen et al., 2003;Starkie et al., 2003). The role of these cytokines in relation
to inflammatory activity, cellular damage, and metabolism have, however, not been
investigated in response to RLW.
1.3.2 The Vicious circle and Pain adaptation models
The vicious circle model is another complementary theory (Johansson & Sojka, 1991),
which has recently been revised (Johansson et al., 2003a). It proposes that
accumulation of metabolites locally in muscle may disrupt normal muscle control and
coordination. The model suggests that in situations when chemosensitive group III-IV
muscle afferents located in the interstitial space are activated (various substances are
proposed below)  the activity in muscle spindles are increased via reflexes 
thereby increasing the activity in the -motoneurons  causing a less efficient motor
control  resulting in an increase in metabolites and inflammatory substances  again
acting on group III-IV muscle afferents and thus creating a positive feedback loop – a
vicious circle. In this way, prolonged muscle activation leading to fatigue causes an
accumulation of local muscle metabolites and algesics thereby activating nociceptors,
and the pain activates a self-perpetuating vicious circle keeping the pain present. Of
notion is, however, that this model is based on findings from animal studies only.
A contrasting model, the pain adaptation model has been proposed (Lund et al., 1991).
This model proposes a tendency to avoid or change painful movement patterns.
Locally, the pain adaptation model predicts that muscle pain affects muscle activation
by inhibition of agonists and excitation of antagonist muscles, thus lowering the
activity of the working muscles (agonists) (Lund et al., 1991). It is proposed that this
pain modulation model may initiate a protective mechanism by muscle “relaxation”,
which is in contrast to the vicious circle model that suggests pain perpetuation as a
consequence. It has however been shown that during low-force contractions,
experimentally induced pain does not affect muscle activation, suggesting that pain
during this type of contraction does not elicit a pain-adaptive response (Birch et al.,
2000).
1.3.3 The Calcium Hypothesis
It has been suggested that calcium plays a crucial role in the initialization and
progression of biochemical events that lead to muscle myalgia (Edwards et al.,
1984;Jackson et al., 1985;Sjøgaard et al., 2000;Gissel, 2000;Westerblad et al., 2000).
Transportation of calcium ions between extra and intracellular compartments plays a
key role in muscle contraction. Moreover, intracellular free calcium is a key regulator
17
- Interstitial changes in trapezius muscle during repetitive low-force work -
of many cell processes including oxidative status and metabolism, and primary
activator of many enzymes that are important for maintaining the structural integrity of
the cells. Changes in the level of intracellular free calcium ions may therefore have
adverse consequences for the cells.
The basis of the “calcium-hypothesis” is the suggestion that RLW can cause an
increase in resting levels of intracellular free calcium. The increase in resting levels is
caused by two effects: Firstly, RLW is suggested to exceed the buffer capacity of the
sarcoplasmic reticulum, thus leading to reduced capacity for reabsorption of released
calcium. Secondly, increased permeability of the cell membrane caused by constantly
repeated excitation cycles lead to increased influx of extracellular calcium and
therefore also contributes to a further increased intracellular calcium level (Gissel &
Clausen, 2001). The long-term consequence for the cells could be a release of
metabolites and algesic substances into the interstitium as well as extensive structural
damage eventually leading to the death of the cell. Leakage of the intracellular enzyme
lactate dehydrogenase (LDH), is a measure of a loss of membrane structure and is
suggested to be caused by calcium induced membrane protein degradation (Gissel,
2000;Gissel & Clausen, 2003). The “calcium-hypothesis” of RLW-induced myalgia
describes how a normal physiological mechanism (the calcium in and outflow cycles
during muscle fiber contraction) under certain conditions (RLW) has the potential to
initiate a series of events that can lead to a chronic condition with characteristics
similar to the clinical features associated with chronic work-related myalgia. Some
support for this hypothesis has been reported; after prolonged low frequency
stimulation, an increased intracellular Ca ++ content in stimulated muscle tissue was
reported concomitant with an increased LDH leakage (Gissel, 2000;Gissel & Clausen,
2001;Gissel & Clausen, 2003).
To summarize, muscle pain from RLW is associated with morphological muscle
changes, perhaps indicating a disturbed energy turnover. At a motor control level, these
changes are speculated to be due to a constant activation of the same MU, increasing
the local metabolic load. On a cellular level a constant activation of the same muscle
cells is speculated to lead to increased intracellular calcium levels that may initiate
events leading to cellular damage. It is important to realize that the pathomechanisms
described above are not mutually exclusive, but may rather be complementary
interacting and may act simultaneously or at different steps and time points during
disease progression of chronic work-related myalgia. However, one important feature
of all models is that the consequence is a release of metabolites and algesics into the
18
- Lars Rosendal -
interstitial space sensitizing or activating the muscle nociceptors, leading to the
sensation of pain in the muscle.
1.4 Muscle pain and nociception
Pain is defined as: “an unpleasant sensory and emotional experience associated with
actual or potential tissue damage, or described in terms of such damage”
(International Association of the Study of Pain) and the experience of pain is therefore
always subjective. In contrast, nociception is related to activity in peripheral or central
receptors (nociceptors). Peripheral nociceptors can be activated by noxious stimuli of
mechanical, thermal and chemical nature or by stimuli that approaches a noxious level
(Mense, 1993;Millan, 1999). Muscle nociceptors are functionally specialized free
nerve endings, located in the interstitial space – type III (A, myelinated) and IV (C,
unmyelinated) afferent fibers. The typical locations of free nerve endings in skeletal
muscle are the wall of arterioles and the connective tissue. Nociceptors can be
activated or sensitized by several chemical substances, e.g. bradykinin, lactic acid,
glutamate, potassium (K+) and serotonin (5-HT) (Mense, 1993;Millan, 1999;GravenNielsen & Mense, 2001).
Bradykinin is suggested to be released in response to trauma and inflammation (Blais,
Jr. et al., 1999), is associated with inflammatory pain (Dray & Perkins, 1993;
Hargreaves et al., 1993) and has been shown to induce pain and to sensitize
nociceptors when injected into muscle tissue (Jensen et al., 1990;Babenko et al.,
1999;Mørk et al., 2003). Bradykinin has a potent vasodilator effect (O'Kane et al.,
1994), and has been shown to be released in response to muscle contraction (Langberg
et al., 2002a). Bradykinin has been measured with the microdialysis technique in rats
(Boix et al., 2002) and in humans in connection to surgery (Hargreaves et al., 1993)
and exercise (Langberg et al., 2002a).
Lactate and K+ are released from muscle tissue in response to contraction. Lactate is
the product of anaerobic energy production from glycolysis (MacLean et al.,
1999;Linossier et al., 2000;von Duvillard, 2001) and lactate is suggested to be
associated to muscle fatigue and pain (Kniffki et al., 1978;Fitts, 1994;Axelson et al.,
2002) although controversy exists on its physiological role in this connection (Rotto &
Kaufman, 1988;Nielsen et al., 2001). K+ is lost to the interstitium during muscle
contraction via voltage-dependent K+ channels activated during action potentials. An
accumulation of interstitial K+ is speculated to have several physiological effects
including regulation of blood flow, development of fatigue, and sensation of muscle
pain (Jensen et al., 1993a;Sejersted & Sjøgaard, 2000). The local muscle K+ response
19
- Interstitial changes in trapezius muscle during repetitive low-force work -
to intense exercise has previously been measured with microdialysis (Green et al.,
2000;Juel et al., 2000).
Glutamate is an excitatory neurotransmitter released from peripheral afferent fibers in
response to nociceptive activation and inflammation (Millan, 1999). Glutamate is a
potent pain modulator in the human central nervous system acting through the
ionotrophic glutamate receptor N-methyl-D-aspartate (NMDA) and acting additive
with the nociceptive role of other substances such as substance P (Alfredson et al.,
2001b). Furthermore, glutamate injected into muscle tissue increases pain intensity and
lowers mechanical pain thresholds (Cairns et al., 2003;Svensson et al., 2003).
Glutamate has been measured in peripheral tendon tissue with the microdialysis
technique (Alfredson et al., 1999;Alfredson et al., 2000) and the presence of peripheral
NMDA receptors has been demonstrated in animal (Coggeshall & Carlton, 1998) and
man (Alfredson et al., 2001a;Alfredson et al., 2001b).
Serotonin (5-HT) has been shown to induce pain and hyperalgesia when injected into
muscle tissue (Babenko et al., 1999;Mørk et al., 2003) and has recently been measured
in painful muscle tissues by microdialysis (Ernberg et al., 1999). 5-HT can be released
from activated platelets, endothelial cells, and mast cells (Jensen et al., 1990;Millan,
1999;Levine & Reichling, 1999;Ernberg et al., 1999;Mørk et al., 2003). Furthermore,
peripheral sensory nerves have been shown to contain 5-HT and may therefore also be
a source for 5-HT (Abramovici et al., 1991), however, the source for muscle 5-HT is
not yet fully elucidated.
In summary, the proposed algesics are all likely candidates to effect muscle
sensitization and pain in relation to work-related muscle disorders. In response to
RLW, using small muscle groups, the absolute changes in metabolites as well as
algesics in muscle tissue may be to small to be detected in systemic blood samples,
although they may be of considerable magnitude locally in the muscle tissue.
Furthermore, the target organs for initiating muscle pain (nociceptors) are located in
the interstitial space, therefore microdialysis was chosen as a technique to study
interstitial muscle response to RLW.
1.5 Microdialysis
Microdialysis is a technique based upon a concept of perfusion of hollow dialysis
tubes, which allow for determination of interstitial concentration of various substances
in vivo in animals and humans (Delgado et al., 1972;Ungerstedt & Hallstrom,
1987;Arner et al., 1988). Briefly, the principle in microdialysis is that a microdialysis
20
- Lars Rosendal -
catheter is inserted in the tissue of interest. The catheter is being perfused with a
solution (termed perfusate) resembling the interstitial fluid (ISF). Substances diffuse
over the membrane and into the perfusate in a concentration-dependent manner, and
samples (dialysate) can be collected for later analysis (Figure 2).
Originally, microdialysis was developed for in vivo determination of biochemical
processes in the extracellular compartments of the brain (Bito et al., 1966;Ungerstedt
& Pycock, 1974;Ungerstedt, 1991). In 1974, Ungerstedt created a microdialysis
catheter that resembled an artificial blood capillary by perfusing a hollow fiber
containing a dialysis membrane obtained from an artificial kidney (Ungerstedt &
Pycock, 1974). This was the beginning of the development of the microdialysis
catheters that we use today.
Figure 2 Schematic description of the principle in microdialysis. A custom-made microdialysis catheter
is inserted into the target tissue and perfused with a physiological solution mimicking the interstitial fluid.
The catheter enters and exits the tissue at different points as shown. Black dots represent molecules.
Reprinted from with permission (Stallknecht, 2003).
Microdialysis has been applied to skin (Birklein et al., 2000), adipose tissue (Arner et
al., 1988;Stallknecht et al., 2001), tendon (Alfredson et al., 1999), peritendon
(Langberg et al., 2001), and bone (Thorsen et al., 1996). Muscle tissue has also been
studied with microdialysis, especially with respect to metabolism (Lönnroth,
1997;Henriksson, 1999;MacLean et al., 1999;Lundberg et al., 2002) and recently
microdialysis was applied to the trapezius muscle to study tension-type headache
(Ashina et al., 2002;Ashina et al., 2003). However, this PhD project is the first attempt
to study repetitive low-force work and work-related myalgia by means of
microdialysis. In spite of the fact that microdialysis today is a quite extensively used
21
- Interstitial changes in trapezius muscle during repetitive low-force work -
technique in research as well as in clinical work such as during surgery, only very few
studies have reported the effect of the insertion trauma on the substances measured.
1.5.1 Catheters
Most published studies applying the microdialysis technique are using commercially
available catheters (CMA-microdialysis, Stockholm, Sweden) with a pore size of 20
kDa. The advantage of this type of catheter is that only one incision is needed (see
Figure 3). The disadvantage may be that it is difficult to keep the catheter in a fixed
position in a working muscle. The catheters used in the present thesis are custom made
(I, II, III), based on experience from my work at the Sports Medicine Research Unit
(Bispebjerg Hospital, Copenhagen, Denmark) (Rosendal, 2000;Langberg et al., 2001).
In contrast to the commercial available catheters, the custom made enters and exits the
muscle tissue at different points (Figure 2). Hence the position stability in a working
muscle may be somewhat better. The use of custom made catheters also allowed for
manufacturing of different catheters with small as well as large molecular weight cutoffs (5 kDa and 3000 kDa). Thus the use of catheters with different cut-offs allowed
determination of substances with both low (e.g. metabolites) and high molecular
weight (e.g. cytokines) in the present PhD work. Of note is that CMA microdialysis
has recently (2003) manufactured a catheter with high molecular weight cut-off (100
kDa, CMA71).
Figure 3 Commercially available
catheters, CMA60 (CMA Microdialysis, Stockholm, Sweden).
The catheter is inserted into the
tissue of interest by a cannula and
has only one insertion point. The
diffusion between interstitial fluid
and membrane occurs at the tip of
the catheter as shown.
Reprinted with permission from
CMA microdialysis.
22
- Lars Rosendal -
2. AIM
The aim of the present thesis was to determine the local trapezius muscle responses to
repetitive low-force work in healthy subjects and in patients with work-related
trapezius myalgia with respect interstitial changes in metabolism, inflammatory
mediators and potential algesic substances.
The specific aims of the present thesis were to:
-
Investigate whether microdialysis is feasible in the trapezius muscle, and to
study to what extent the insertion trauma affects the local muscle cytokine
response.
-
To study whether local muscle metabolite accumulation, electrolyte flux, and
cytokines were affected by standardized repetitive low-force work.
-
Study whether differences exist in local muscle metabolite accumulation,
blood flow, and algesic substances between patients with trapezius myalgia
and healthy controls during rest and standardized repetitive low-force work.
2.1 Hypothesis
The working hypothesis was that:
-
Repetitive low-force work does affect local muscle metabolism but that this
may not be reflected systemically.
-
Painful muscle tissue, caused by repetitive low-force work, will reflect an
increased metabolite accumulation due to a reduced blood flow.
-
The local concentrations of algesic substances will be increased in painful
muscle tissue with the potential of sensitizing local nociceptors that will
become hyper-responsive to mechanical exposure.
23
- Interstitial changes in trapezius muscle during repetitive low-force work -
24
- Lars Rosendal -
3. METHODS
3.1 Participants
The participants in study A+B were healthy males without any known muscle
disorders whereas the participants in study C were female patients with trapezius
myalgia and healthy female controls. Three participants took part in both studies A+B.
For a more detailed description of the patient group, see Paper III. The characteristics
of the participants are summarized in Table 1.
Table 1 Participant data. Data are mean (range).
Study
Sex.
Health status.
Number of
participants.
Age, years.
Height, cm.
Weight, kg.
A
B
C
Male
Healthy
6
Male
Healthy
6
Trapezius myalgia
19
Healthy
20
32 (26-40)
181 (176-184)
81 (75-89)
30 (28-33)
180 (173-188)
81 (68-89)
41 (21-61)
165 (153-175)
65 (52-87)
36 (26-56)
166 (153-178)
63 (48-82)
Female
3.2 Protocols
3.2.1 Pilot study
A pilot study on three healthy participants (two males, one female) was conducted to
evaluate whether microdialysis was feasible in the trapezius muscle. The result
revealed that a) only negligible pain was associated with insertion of the microdialysis
catheters in the trapezius muscle and b) that the microdialysis catheters were all
functional after static, intermittent static and dynamic arm exercises of 10 min
duration.
3.2.2 Study A
In order to study to what extent the local muscle cytokine response was affected by the
insertion trauma, a microdialysis catheter (3000 kDa) was inserted into the trapezius
muscle on the dominant side of six healthy participants. The participants rested sitting
upright for two hours. For further details, see Paper II.
25
- Interstitial changes in trapezius muscle during repetitive low-force work -
3.2.3 Study B
In order to evaluate whether local and systemic metabolism and cytokines were
affected by a standardized repetitive low-force work task, two custom-made
microdialysis catheters (cut-off: 5kDa and 3000 kDa) were inserted into the dominant
trapezius muscle of six participants. After a 60 min resting period (to allow tissue
recovery from the insertion trauma), the participants performed a 20 min standardized
repetitive arm movement task, mimicking an occupational job situation (RLW, see
description later). This was followed by a 60 min recovery period (Recovery 1). In
order to create a situation in which an increase in anaerobic muscle metabolism would
occur in both interstitium and blood, the participants performed a 10 min sustained
static shoulder flexion at 20% MVC (SUS, see description later), followed by the
second 60 min recovery period (Recovery 2). For further details, see Paper I.
3.2.4 Study C
In the last study, the aim was to investigate if there exist local muscular differences in
anaerobic metabolism, algesic substances, and blood flow between patients with workrelated trapezius myalgia and healthy controls during rest and when exposed to
repetitive low-force work. The groups studied consisted of 20 healthy controls and 19
patients with chronic WTM who had been referred to a multidisciplinary pain center,
diagnosed, and clinically examined (see below). Two custom-made microdialysis
catheters (cut-off: 5kDa and 3000 kDa) were inserted into the dominant (most painful)
trapezius muscle where after the participants rested for 140 min, performed a 20 min
standardized repetitive arm movement task, mimicking an occupational job situation
(RLW) followed by a 2-hour recovery period. For further details, see Paper III.
3.3 Overview of used methods and exercise protocols
The methods used in the studies are summarized in table 2.
26
- Lars Rosendal Table 2 Methods and exercise protocols used in studies.
Study
A
Exercise protocol:
Repetitive low-force contraction
Intense static contraction
Microdialysis
Blood samples
MVC
EMG
RPE
VAS
PPT
Pain drawing
Clinical examination
Blood flow determination
Ultrasound B-scan (muscle thickness)
X
B
C
X
X
X
X
X
X
X
X
X
X
X
X
X
X
X
X
3.4 Exercise protocols
3.4.1 Repetitive low-force work (RLW)
To mimic a standardized occupational work situation, a repetitive low-force exercise
protocol was developed. The exercise protocol consisted of moving short wooden
sticks back and forth between standardized positions 30 cm apart on a giant pegboard
at a frequency of 1 Hz. The participants performed the RLW in a seated position with
the pegboard placed 30 cm in front of them, measured from the elbow with the upper
arm hanging vertically and the elbow in a 90 flexion (Figure 4). The RLW was
performed for 20 min in study B+C.
3.4.2 Static shoulder flexion
The sustained static shoulder flexion protocol (SUS) was developed to create a
situation in which a significant change in metabolite accumulation should be expected
locally in the trapezius muscle. SUS served as a positive control situation for the ability
of the microdialysis technique to detect changes in metabolism during exercise. The
aimed load during SUS was 20% MVC and it was performed as a 90 shoulder flexion
in the sagittal plane. The load was placed just proximal to the elbow joint at the center
of gravity of the arm and 20% MVC was including the weight of the arm (5% of body
weight) (Jensen et al., 1993b). To calculate the load, MVC was performed (see later).
27
- Interstitial changes in trapezius muscle during repetitive low-force work -
Figure 4 Repetitive low-force work. The participants performed 20 min of repetitive low-force work by
moving short wooden sticks back and forth between standardized positions 30 cm apart on a giant
pegboard at a frequency of 1 Hz. The exercise was intended to mimic an occupational job situation with
repeated monotonous work at a low-force contraction level.
3.5 Microdialysis
A thin microdialysis catheter is placed in the muscle tissue, and the catheter is
continuously perfused with a solution resembling the ISF. Water-soluble substances in
the ISF diffuse across the membrane and enter the perfusate in a concentrationdependent manner. The concentration of a substance in the dialysate reflects the
concentration in the tissue, making it possible to detect values of and changes in
interstitial concentrations of specific metabolites and mediators. The exchange of
substances being sampled is limited by the molecular weight and characteristics of the
substances, pore size of the dialysis membrane (cut-off), membrane length, perfusion
flow, and blood flow.
In the ideal system, equilibrium between the concentrations of a substance in the
dialysate and the concentrations in the ISF would be reached. However, since diffusion
is dependent on tissue-properties like interstitial pressure and temperature, properties
of the dialysis membrane and perfusion rate, correction for the rate of exchange is
needed in order to calculate the interstitial concentration of a substance accurately
(Lönnroth, 1997).
28
- Lars Rosendal -
3.5.1 Relative recovery and interstitial calculations
In order to determine the interstitial concentration, the in-vivo relative recovery (RR)
needs to be determined. Several different techniques have been used (Jacobson et al.,
1985;Lönnroth et al., 1987;Bolinder et al., 1992) however the internal reference
technique (Scheller & Kolb, 1991), was used in the present thesis. The advantages of
the internal reference technique are that each catheter is calibrated in situ, that
interstitial changes are time-wise rapidly detected when conditions change e.g. in
response to exercise, and that relatively large flow rates can be used, enabling a larger
sample volume for biochemical analysis.
With the internal reference technique, the microdialysis catheters are calibrated in situ
by perfusing the catheters with a perfusate containing an indicator substance that
resembles the substance of interest, but which can be distinguished from the substance
of interest during analysis of the dialysate (Figure 5). The indicator substance can
either be the substance of interest labeled with radioactivity or a radioactive labeled
substance with the same diffusion characteristics as the substance of interest. By using
different labeled substances such as (3H, 14C), it is potentially possible to monitor the
diffusion of multiple molecules in the same catheters simultaneously. It is assumed that
the RR from the interstitial fluid to perfusate of unlabelled metabolite equals relative
loss from the perfusate to interstitial fluid of labeled metabolite (Scheller & Kolb,
1991;Lönnroth & Strindberg, 1995). RR is calculated for each microdialysis catheter as
RR = (cpmp – cpmd)/ cpmp, where cpmp was counts per min in the perfusate and cpmd
in the dialysate. The interstitial concentrations (Ci) are calculated as Ci = (Cd – Cp)/RR
+ Cp, where Cd was dialysate concentration and Cp perfusate concentration.
*
Perfusate
Dialysate
*
*
Figure 5 The principle of microdialysis. The microdialysis probe is being perfused with a solution
resembling the interstitial fluid (perfusate). Substances diffuse over the membrane and into the perfusate in
a concentration-dependent manner and samples (dialysate) can be collected for later analysis. The internal
reference technique is used for determination of in situ recovery of compounds in the microdialysis
catheter. A radioactive compound (*) with similar characteristics as the molecule of interest is added to the
perfusate. The relative loss can be calculated based on the difference between the radioactivity left in the
dialysate and that of the perfusate. Reprinted with permission from (Langberg, 1999).
29
- Interstitial changes in trapezius muscle during repetitive low-force work -
3.5.2 Blood flow
Nutritive trapezius muscle blood flow was estimated by the microdialysis ethanol
technique (Hickner et al., 1994) using 3H2O instead of ethanol (Stallknecht et al.,
1999). A small amount of 3H2O was added to the perfusate, and the blood flow is
calculated as the ratio of 3H2O in the dialysate and the perfusate (the outflow-to-inflow
ratio). The ratio varies inversely with the local blood flow in the tissue (Hickner et al.,
1994;Stallknecht et al., 1999).
3.5.3 Construction of microdialysis catheters
Low-cut catheter
For sampling of low weight molecules, costume-made microdialysis catheters were
constructed. These catheters were made from single plasmaphoresis hollow fibers (0.4
mm in diameter, molecular mass cut-off 5 kDa; Alwall, GFE 11, Gambro Dialysatoren,
Hechingen, Germany), glued to a gas-tight nylon inlet and outlet tubing (Portex
Autoclavable Nylon Tubing, Portex Limited, Smiths Industries, Kent, England), and
with a suture thread (Johnson & Johnson, Brussels, Belgium) glued to the membrane to
improve the mechanical stability of the fiber. The length of the microdialysis
membrane was 30 mm.
High-cut catheter
For determination of molecules with high molar weight, another type of microdialysis
catheter was constructed from a single plasmaphoresis hollow fiber (0.4 mm in
diameter, molecular mass cut-off 3000 kDa; Asahi Medicals, Japan), glued to a gastight nylon inlet and outlet tubing (Portex Autoclavable Nylon Tubing, Portex Limited,
Smiths Industries, Kent, England) and with an inner wire (100 m stainless steel wire)
to improve the mechanical stability of the fiber. The length of the dialysis membrane,
available for diffusion, was 30 mm.
Both types of microdialysis catheters were packed in covers and sterilized
(ETO-sterilization, Maersk Medicals, Denmark).
3.6 Electromyography
To evaluate the work-load during RLW and SUS (study B, Paper I), electromyography
(EMG) was recorded from the trapezius muscle by bipolar surface electrodes. The
center of each pair of electrodes was placed directly above the microdialysis membrane
and 2 cm lateral to the midpoint between the seventh cervical vertebrae and the lateral
end of acromion (Jensen et al., 1993c). The inter-electrode distance was 20 mm.
30
- Lars Rosendal -
The EMG signal was amplified, band-pass filtered and sampled on dataloggers with a
sampling frequency of 1024 Hz. The signals were transferred to a computer where they
were visually quality checked, high-pass filtered and full-wave rectified. The analysis
of the EMG signal during the two exercise periods was performed in the time and in
the frequency domain by calculating the root-mean-square (EMGRMS) value and the
mean power frequency (EMGMPF).
3.6.1 Maximal voluntary contraction and force
A strain-gauge force transducer was used to measure the force exerted by the
participants during the MVC. The MVC was a bilateral 90o shoulder flexion with
resistance placed just proximal to the elbow joint at the center of gravity of the arms
(Jensen et al., 1993b). The MVC was performed 3 times with 2 min between each
attempt.
3.7 Biochemical measurements
The biochemical measurements were all conducted with validated, commercial assays
and methods. An overview of the biochemical measurements applied in each of the
studies is given in Table 3. Most of the analytical methods are described in detail in
the respective papers. However, a few analytical methods that are not reported in any
of the papers are described below.
3.7.1 Interleukin-1 beta (IL-1)
Interstitial muscle IL-1 was measured with a Quantikine® assay from R&D systems
(Minneapolis, MN, USA). In order to meet the minimum sample volume requirements
of the assay, dialysate samples were diluted 20-fold with calibrator diluent. The lowest
calibration standard was used as the level of detection for the cytokine in the dialysate,
which was 10 pg/ml when adjusting for the dilution factor.
3.7.2 Lactate dehydrogenase (LDH)
Interstitial muscle LDH activity was measured by a Cytotoxicity Detection Kit (LDH)
from Roche Molecular Biochemicals (Roche Diagnostics, Manheim, Germany), which
has a level of detection of 21.5 mU/ml.
31
- Interstitial changes in trapezius muscle during repetitive low-force work -
3.7.3 Bradykinin (BKN)
The analysis for interstitial muscle BKN was performed with a radioimmunoassay kit,
RIK 7151 from Peninsula laboratories Inc (CA, USA). The level of detection when
adjusted for dilution factor was 25 pg/ml.
Table 3 Biochemical measurements from interstitial fluid (ISF) and blood samples.
Study
A
Lactate
ISF
Plasma
Pyruvate
Potassium (ISF, Plasma)
Interleukin-6
ISF
Plasma
Interleukin-1 (ISF)
Lactate dehydrogenase
ISF
Plasma
Creatine kinase, (Plasma)
Glutamate, (ISF)
Serotonin (5-HT), (ISF)
Bradykinin, (ISF)
X
X
B
C
X
X
X
X
X
X
X
X
X
X
X
X
X
X
X
3.8 RPE, VAS, and algometry
Rating of perceived exertion (RPE) in the shoulder region was performed during RLW
and SUS in study B (Paper I). RPE was rated according to a 10-graded Borg scale
(from 0 to 9) (Borg, 1982) where 0 corresponds to “no perceived exertion” and 9
corresponds to “maximum perceived exertion”.
Pain intensity was assessed in study C by a 100 mm visual analogue scale (VAS)
ranging from 0 (no pain) to 100 mm (worst possible pain) (Collins et al., 1997)
(Paper III).
Muscular tenderness, measured as pressure pain threshold (PPT), was determined in
study C in connection with the clinical examination (see below). PPT was determined
with algometry as it has previously been done (Persson et al., 2000). PPT was
32
- Lars Rosendal -
determined on both sides, at 3 points over each trapezius muscle (lateral, middle and
medial), and reference measurements were made on the tibialis anterior muscle on both
legs. All PPT measurements were conducted 3 times intercepted by approximately
1 min.
3.9 Clinical examination and questionnaire
In study C, the Nordic Ministry Council Questionnaire was used to survey pain in the
last 12 month (Kuorinka et al., 1987) and a structured interview that also concerned the
course of pain and work-tasks before onset of pain was conducted. The myalgia was
considered to be work-related when it was reported to have started in connection to
highly repetitive or static work tasks and was continuously getting worse during the
work day. The participants were examined by a standardized clinical examination
(Ohlsson et al., 1994) to ensure that they complied with the inclusion criteria (the
inclusion and exclusion criteria are described in detail in Paper III).
3.10 Statistics
For detail on the applied statistical tests, see the statistical paragraphs of the papers.
The level of significance was set at P<0.05. In general, the residuals were always
checked for normal distribution and variance homogeneity. Parametric statistics were
preferred if data were normally distributed.
33
- Interstitial changes in trapezius muscle during repetitive low-force work -
34
- Lars Rosendal -
4. RESULTS
The Roman numerals in parenthesis (I, II, III) refer to the paper in which more detailed
information can be found.
4.1 Response to the insertion trauma, healthy subjects
4.1.1 Cytokine response (II)
The muscle interstitial IL-1 concentration was below the level of detection at all time
points in the two participants who were measured.
In the first sample (5 min), IL-6 was below the level of detection in 5 of 6 participants.
During the initial hour of rest, the interstitial IL-6 concentration gradually increased to
359  171 pg/ml after 65 min (P<0.001). After 105 min of rest, the interstitial muscle
IL-6 concentration had increased to 489  202 pg/ml but, this was not statistically
different from the previous level at 65 min (Figure 6 B).
4.1.2 Lactate dehydrogenase (LDH)
A release of LDH into the interstitial space indicates cell membrane leakage.
The interstitial muscle LDH response to microdialysis catheter insertion is shown in
(Figure 6 A). The LDH level that was initially 2127 mU/ml, decreased by 55 % within
the first 30 min (P<0.05) and was negligible after 105 min. At 105 min, the interstitial
LDH level was significantly reduced below the level at previous time points and was
not further reduced (data not shown).
4.2 Response to RLW and SUS, healthy subjects
Rating of perceived exertion (I)
Ratings of perceived exertion increased during RLW from 0  0 at the start of the
exercise to 3.2  0.5 during the 20th min. During the sustained static shoulder flexion
(SUS), RPE increased from 0  0 to 8.5  0.3 at the end of the 10th min.
4.2.1 Electromyography and work-load (I)
During the 20 min RLW, the average muscle activity level (work-load) in the active
trapezius muscle was 8-9 % EMGmax, and 2-3 % EMGmax in the contralateral trapezius
muscle (control). EMG root mean square (RMS) increased in both the active and the
control trapezius muscle and mean power frequency (MPF) decreased in the active
trapezius muscle, all P<0.001 (Figure 7 A+C).
35
- Interstitial changes in trapezius muscle during repetitive low-force work -
During the 10 min SUS, the average muscle activity level in the active trapezius
muscle was initially (first min) 15  3 % EMGmax and increased to 33  9 % during the
last min (P<0.03). In the control trapezius muscle, the activity level increased
significantly from 2  1 % EMGmax to 9  4 %. Furthermore, EMGRMS increased in
both the active and the control trapezius muscle and EMGMPF decreased in the active
trapezius muscle, all P<0.001 (Figure 7 B+D).
Figure 6 Interstitial muscle IL-6 and LDH in response to an insertion trauma.
Interstitial muscle concentrations of lactate dehydrogenase (LDH, panel A) and interleukin-6 (IL-6, panel B)
in response to the insertion of a microdialysis catheter in the upper trapezius muscle of healthy male
participants (n=6). Interstitial muscle IL-6 and LDH measurements represent the concentration over a 10
min sampling period. Data are given as mean ± SEM.
* denotes significant difference from corresponding initial level (5 min) (P<0.05),
# denotes significant difference from initial level and from levels at 25 min and 65 min (P<0.05).
4.2.2 Metabolism (I)
In response to RLW, interstitial muscle lactate increased by 38% (P<0.001), remained
increased during the initial 10 min recovery (P<0.003) and returned to baseline levels
after 20 min recovery (Figure 8 A). In contrast, plasma lactate was not affected by
RLW (Figure 8 A). Interstitial muscle pyruvate levels were constant during RLW, but
increased transiently 20 min after RLW (Figure 8 B).
36
- Lars Rosendal -
During SUS, interstitial muscle lactate increased by 43% (P<0.003) and further
increased by 96% during the initial 10 min recovery (P<0.001) (Figure 8 A).
Thereafter, the lactate level gradually declined and after 60 min, it was no longer
different from the baseline level (P=0.84) (Figure 8 A). Plasma lactate also increased
in response to SUS (P<0.001) and was back to baseline level after 30 min recovery
(Figure 8 A). Interstitial muscle pyruvate was constant during SUS, but increased
significantly immediately after SUS and remained elevated for 30 min (all P<0.001)
before it returned to baseline level (Figure 8 B).
Figure 7 Electromyography. Recordings of the root mean square (EMGRMS) (A, B) and the mean power
frequency (EMGMPF) (C, D) from the active trapezius muscle (●, filled circle) and the contra-lateral
control trapezius muscle (○, open circle). EMG was recorded during 20 min repetitive low-force work
(RLW) (A, C) and during 10 min of 90  sustained static shoulder flexion (SUS) at 20% MVC (B, D).
Sampling of blood from the contralateral control arm during RLW introduced movement artifacts in the
EMG recordings, and the corresponding EMGRMS values have therefore been omitted. The shortest time
holding the original workload during SUS is shown with a vertical dotted line. Data are given as mean 
SEM. * Significant time effect (Page test).
37
- Interstitial changes in trapezius muscle during repetitive low-force work -
Figure 8 Lactate, pyruvate,
interleukin-6, and potassium dynamics
in the upper trapezius muscle.
Interstitial muscle (bar) and plasma
(line) lactate (panel A), interstitial
muscle pyruvate (panel B), interstitial
muscle interleukin-6 (IL-6) (panel C),
plasma IL-6 (panel D), and muscle
dialysate potassium (K+) (panel E)
during: baseline rest (BL 1, BL 2), 20
min repetitive low-force work (RLW),
10 min 90  sustained static shoulder
flexion at 20% MVC (SUS), and
recovery after exercises (Recovery 1+2).
The gray bar indicates measurements
performed during exercise. Interstitial
muscle measurements represent the
concentration over a 10 min sampling
period and are shown as bars, whereas
plasma levels are shown as lines.
Measurements are performed in healthy
males (n = 6) and data are given as
mean ± SEM.
* denotes significant difference from
the corresponding baseline value (BL1
or BL2) (P< 0.05).
38
- Lars Rosendal -
4.2.3 Cytokine response (II)
As in study A, muscle dialysate IL-1 was below the level of detection (10 pg ml-1) at
all time points (rest, RLW and recovery) in all 6 participants. The interstitial IL-6
concentration was initially (5 min) also below the level of detection, however, a
gradual increase in IL-6 was found during the resting period following the insertion of
microdialysis catheters (similarly as in study A). A small and insignificant increase in
IL-6 was noted after 30 min rest but IL-6 continued to increase, being significantly
increased after 55 min of rest (P<0.001). In response to RLW, interstitial IL-6
increased to 1246  461 pg/ml - significantly increased compared to the pre-exercise
level of 289  128 pg/ml (Figure 8 C). The interstitial IL-6 concentration continued to
increased for the 30 min recovery period following RLW, ending at a level of 2133 
477 pg/ml. Plasma IL-6 did numerically increase during the experiment, however, this
was not statistically significant (Figure 8 D).
4.2.4 Potassium (I)
Dialysate K+ concentrations increased significantly during RLW and tended to remain
elevated during the initial 10 min recovery period (P=0.08) where after it decreased to
resting levels (Figure 8 E). In contrast, plasma K+ concentrations remained unaffected
by the low-force work protocol – between 3.8 and 4.0 mmol/l.
During SUS, the dialysate K+ dynamics was similar as during RLW, with respect to
both concentrations and changes (Figure 8 E). Also during SUS, plasma K+ concentrations
remained at levels not different from resting levels.
4.3 Response to RLW, patients with work-related trapezius myalgia and
healthy controls
There were no differences between patients with work-related trapezius myalgia
(WTM) and healthy controls with respect to muscle thickness over the midpoint of the
trapezius muscle (12.0  0.5 vs. 13.0  0.6 mm) and superjacent tissue (9.5  0.5 vs.
8.6  0.5 mm) determined with ultrasound or with respect to indications of muscle
damage, determined from plasma CK (95  9 vs. 109  14 U/l) and plasma LDH (348
 13 vs. 347  24 U/l). Reference intervals for women are CK; <150 U/l and LDH;
<480 U/l respectively.
39
- Interstitial changes in trapezius muscle during repetitive low-force work -
4.3.1 Pain intensity and pressure pain threshold (III)
The average PPT was lower in patients with WTM than in controls at all sites (lateral
part, middle and medial part) in both the right and the left trapezius muscle (Table 4).
No significant group differences in PPT were noted at the control sites on the right and
left tibialis anterior muscles (Table 4).
VAS was higher in patients with WTM than in controls at all time points during rest,
RLW and recovery (P<0.001) (Figure 9 A). In response to RLW, VAS immediately
increased in patients with WTM, peaking at the end of exercise at 70 ± 5 mm (all,
P<0.001) followed by a slow decrease during recovery (Figure 9 A). VAS was
significantly increased for 1 h post-exercise. In controls, there was a small increase in
VAS during RLW, which became significant only at the end of exercise, peaking at 7 ±
2 mm, immediately returning to levels not significantly different from baseline (Figure
9 A).
4.3.2 Blood flow (III)
There were no differences in trapezius muscle blood flow between the two groups
during baseline rest and RLW (Figure 9 B). In both groups, RLW was associated with
an increased blood flow. During the initial recovery period, blood flow had decreased
to baseline level in controls whereas it remained significantly increased in patients with
WTM (Figure 9 B).
Table 4 Pressure pain threshold (PPT)
Groups
Location
Right trapezius, lateral part
Right trapezius, middle part
Right trapezius, medial part
Left trapezius, lateral part
Left trapezius, middle part
Left trapezius, medial part
Left tibialis anterior
Right tibialis anterior
Healthy
Controls
(n=19)
288±16
269±17
327±25
365±25
280±15
272±19
280±19
278±19
Work-related
Trapezius Myalgia
(n=20)
152±17
143±18
143±17
161±16
124±11
160±13
241±25
265±28
Statistics
p-values
<0.001 #
<0.001 #
<0.001 #
<0.001 #
<0.001 #
<0.001 #
0.232 ns
0.705 ns
Data are mean ± SEM and values are in kilopascal (kPa). The value of each location is based on the mean
value of three recordings. # denotes significant difference between groups, independent samples t-test.
40
- Lars Rosendal -
4.3.3 Metabolism (III)
Interstitial lactate and pyruvate concentrations were at all time points (baseline, RLW,
and recovery) higher in patients with WTM than in controls (all P<0.001) (Figure 9 C, D).
In response to RLW, lactate and pyruvate increased in both groups, but only
significantly in the patients with WTM (Figure 9 C, D).
4.3.4 Algesic substances (III)
Dialysate bradykinin concentrations were below the level of detection at all time points
(baseline, RLW, and recovery).
In contrast, interstitial muscle 5-HT was 6-fold higher in patients with WTM than in
controls (22.9  6.7 vs. 3.8  1.3 nmol/l) during baseline rest (P<0.01). The interstitial
concentration of 5-HT was positively correlated to resting VAS (r = 0.55, P<0.001) but
not to PPT.
The interstitial glutamate levels were higher in patients with WTM than in controls at
all time points (P<0.05) (Figure 9 E). In patients with WTM, baseline glutamate levels
were negatively correlated to PPT (r = -0.47, P<0.001). Glutamate levels increased
significantly in both groups during RLW and in patients with WTM, the glutamate
levels were positively correlated to VAS (r = 0.45, P<0.01). Following RLW, glutamate
immediately decreased to baseline levels in both groups, and in the patients with
WTM, interstitial glutamate was decreased below the baseline level after 2 h recovery
(Figure 9 E).
41
- Interstitial changes in trapezius muscle during repetitive low-force work -
Figure 9 Muscle pain, blood flow, lactate,
pyruvate, and glutamate levels in the upper
trapezius muscle of patients with workrelated trapezius myalgia and healthy
controls.
Pain ratings (VAS) (panel A), muscle blood
flow (panel B), interstitial lactate (panel C),
interstitial pyruvate (panel D), and interstitial
glutamate (panel E) levels in the upper
trapezius muscle of female patients with
chronic trapezius myalgia (, n = 19) and
healthy female controls (, n = 20).
Measurements are performed during baseline
rest (BL, 140-160 min), during 20 min
repetitive low-force work (RLW), and
during recovery from work (RECOVERY,
180-300 min).
Blood flow (outflow to inflow ratio of 3H2O),
lactate, pyruvate and glutamate were
measured in 20 min sampling periods.
In panel B, note that a reduction in outflow
to inflow ratio indicates an increase in blood
flow. All data are mean ± SEM.
# denotes significant overall difference in
mean values between groups.
* denotes significant difference from the prework baseline value.
42
- Lars Rosendal -
5. DISCUSSION
5.1 Main findings
The main findings in the present thesis are a) metabolism and algesics can be studied
in trapezius muscle with microdialysis, b) that repetitive low-force work is associated
with an increase in local anaerobic metabolism and a substantial increase in interstitial
IL-6, c) that these changes could not be detected in systemic blood samples, and finally
that d) work-related trapezius myalgia is associated with increased anaerobic
metabolism as well as increased levels of potential algesic substances locally in the
painful muscle tissue – indicating that even after long lasting pain experience, peripheral
nociceptive processes may be activated.
5.2 Applicability of microdialysis in trapezius muscle
The conducted studies have shown that microdialysis is a feasible technique for
studying local metabolism and electrolyte flux in trapezius muscle tissue during rest
and during static as well as dynamic work protocols. Insertion of microdialysis
catheters into the trapezius muscle was possible in both males and females as well as in
healthy subjects and in patients with trapezius myalgia, and was not associated with
significant increases in VAS (healthy) or only a temporary increase (patients with
WTR) (Paper III). Once the custom made catheters had been positioned in the trapezius
muscle, they remained functional throughout the study periods during rest as well as
during exercise in 100 out of 102 cases ~ 98 %.
5.2.1 Microdialysis vs. blood samples
The marked changes in metabolites in the muscle interstitium during RLW were not
associated with parallel changes in plasma levels. This may be due to the relatively
small muscle mass involved in the repetitive low-force arm work, reuptake by inactive
tissues and muscle compartments, or metabolization by other tissues (such as the liver).
Whether the lack of plasma changes is due to one or more of the possibilities described
above is not possible to determine from the present studies. However, it shows that the
microdialysis technique can provide physiological knowledge not attainable with
systemic blood sampling. Of note is, however, that venous effluent blood sampling was
not performed from the trapezius muscle and that this particular technique may provide
more sensitive measurements than systemic venous blood measurements. Still, venous
effluent blood sampling is very difficult from trapezius muscle. Furthermore,
microdialysis enable measurements performed directly in painful tissue, and this may
43
- Interstitial changes in trapezius muscle during repetitive low-force work -
be a very important feature when addressing differences is local concentrations of
algesics between painful and non-painful tissue, as it was done in study C.
5.3 Muscle response to insertion trauma
Like all invasive measurements, one drawback is the trauma applied to the tissue that is
studied. In microdialysis, the insertion of microdialysis catheters into a muscle causes
an insertion trauma. In order to allow the tissue to recover from the insertion trauma, a
resting period always follows the insertion before baseline measurements are obtained.
During the initial studies (Paper I), a resting period of 60 min following insertion was
used, and during that period, interstitial IL-6 increased slightly, being significantly
increased after 55 min. A control experiment was therefore conducted (Paper II). This
experiment confirmed that IL-6 slowly increases during the initial hour, where after the
interstitial IL-6 concentration was relative stable in the 2nd hour (Figure 6 B). Thus it
seems that interstitial IL-6 increases in response to trauma with a delay of more than 30
min. The delay in IL-6 could suggest that this is due to transcriptional activation of IL6 locally in the muscle tissue, as it has been shown in response to exercise (Keller et
al., 2001). In response to an insertion trauma, the first dialysate sample normally has
an increased concentration of substances released from the cells, followed by a rapid
decrease in the concentrations of most substances (Ungerstedt, 1991). In line with this,
increased concentrations of lactate and K+ were initially seen in study B before it
stabilized (Figure 8 A, E). A disturbed interstitial milieu can last from a few minutes
to several hours, depending on biological differences between tissues (Ungerstedt,
1991). Baseline levels have been shown to be reached within 10-20 min for e.g.
neurotransmitters in brain (Meyerson et al., 1990), within 60 min for lactate in skeletal
muscle (Rosdahl et al., 1998),(Henriksson, 1999) and within 120 min for inflammatory
mediators in peritendinous tissue (Langberg et al., 1999). To study to what extent the
muscle cells are damaged during the insertion, interstitial LDH (intracellular enzyme)
was measured. The result showed that LDH decreased rapidly within the first 20 min,
but it also showed that the interstitial LDH concentration was not negligible until 105
min after the insertion (Figure 6 A). This finding does not suggest that the cells are
“leaking LDH” for almost 2 hours – more likely, cellular repair occur relatively fast
(McNeil & Steinhardt, 1997). However, it demonstrates that the interstitial milieu may
be disturbed for up to 2 hours, and suggest that “true” baseline values may not be
obtainable in that period. Therefore in study C, the recovery period following the
insertion of the microdialysis catheters was extended to 2 hours before baseline
measurements were obtained.
44
- Lars Rosendal -
5.4 Muscle metabolism during RLW and during intense static contraction
Marked local changes in interstitial metabolites in the upper trapezius muscle were
found in response to 20 min repetitive low-force work. The increase in lactate showed
that anaerobic metabolism was increased in the trapezius muscle during RLW, despite
that the activity level was only 8-9 % EMGmax (Paper I). This finding could be due to
an inhomogeneous muscle activation in the trapezius muscle, thereby imposing a
heavy load on some muscle parts, which may indicate that the Cinderella hypothesis
(Hägg, 1991) holds true for trapezius muscle activation during RLW. A continuous
activation of the same motor units has previously been shown in muscle during static
and dynamic contractions (Søgaard, 1995). Another possibility is that the increase in
lactate could be caused by an insufficient local muscle blood flow and thereby
ischemia in the vicinity of the active muscle fibers as it has been proposed (Travell JG
et al., 1942;Larsson et al., 1990). Blood flow was not measured in study B and it is
thus not possible to directly answer this question. However, the results from study C do
not corroborate this suggestion because it was observed that local muscle
microcirculation was in fact increased, in trapezius muscle during RLW in healthy
participants (Figure 9 B, Paper III).
An increase in interstitial metabolites is speculated to be one of the corner stones in the
pathophysiology behind pain development in work-related muscle disorders (Edwards,
1988;Henriksson, 1988;Hägg, 1991;Sjøgaard & Søgaard, 1998;Hägg, 2000;Sjøgaard et
al., 2000;Johansson et al., 2003b). Recently however, Vøllestad and Røe expressed
doubt as to whether anaerobic metabolism is at all increased in response to RLW
(Johansson et al., 2003b). They based their doubt on a lack of in-vivo evidence in the
literature and on the lack of increased lactate levels in muscle biopsies during an
intermittent work protocol in quadriceps femoris (Vøllestad et al., 1988). Instead, they
proposed that force oscillations during RLW will increase the shear stress of muscle
fibers moving in relation to each other within the muscle, which could elicit
mechanical nociceptor activation (Johansson et al., 2003b). Although sheer stress may
very well contribute to the pain mechanism during RLW – especially when muscle
levels of algesics with sensitizing effects are increased (see section on algesics), it has
previously been shown that anaerobic metabolism indeed can increase in response to
low intensity work. An increase in muscle lactate was demonstrated in the early phase
of low-force static work at 5% MVC (Sjøgaard, 1988), and recently a continuously
increasing interstitial lactate level was demonstrated in trapezius muscle in response to
static contraction at 10% MVC (Ashina et al., 2002). An increase in interstitial lactate
in response to RLW was also found in study C, although the increase was only
45
- Interstitial changes in trapezius muscle during repetitive low-force work -
significant in the group with muscle pain. Highly surprising, the interstitial lactate
levels were significantly increased by approx. 30 % at baseline rest as well as during
RLW and recovery in the patients with trapezius myalgia (Figure 9 C). The
physiological role of such an increase is not known, but it may indicate that the local
muscle activity level is increased in WTM, even at rest. Supporting this idea, it was
shown that trapezius myalgia was associated with a diminished ability to relax painful
muscle tissue during resting periods between contractions (Larsson et al., 2000a),
which could potentially pose a greater metabolic load on trapezius muscle.
Furthermore, increased resting EMG concomitantly with a lowered blood flow has also
been reported from patients with WTM (Larsson et al., 1999), which could also
increase the lactate levels. A reduced blood flow has repeatedly been suggested to be
one of the most likely mechanism behind WTM (Larsson et al., 1988;Larsson et al.,
1990;Larsson et al., 1999). However, local muscle blood flow was not reduced in the
patients with WTM compared with healthy controls and the increased lactate levels
cannot be ascribed hereto alone. On the other hand, some morphological evidence
suggest that type I muscle fiber size is increased in subjects with WTM (Kadi et al.,
1998) and that capillarization per fiber cross sectional area is reduced (Kadi et al.,
1998;Larsson et al., 2004) which, could reduce the oxygen diffusion potential at the
same absolute blood flow. Furthermore, an increased appearance of ragged red muscle
fibers have been reported in subjects with WTM (Larsson et al., 1988;Kadi et al.,
1998;Larsson et al., 2000b) which is suggested to indicate an energy crisis (Kadi et al.,
1998;Hägg, 2000;Larsson et al., 2004). Based on the data in the present thesis, it could
be suggested that the biopsy data are now supported by in vivo evidence of altered
metabolism locally in trapezius muscle of patients with WTM, but that these changes
are not due to reductions in blood flow. Of note is however, that it is not possible
quantify the size of the blood flow with the microdialysis technique.
In response to RLW, a post-exercise increase in interstitial pyruvate was noted (Figure 8 B).
An increase in pyruvate following exercise has previously been reported (Axelson et
al., 2002) although the literature is conflicting and increased pyruvate levels have also
been reported during exercise (Linossier et al., 2000;Boschmann et al., 2002). This
increase in pyruvate following an exercise-induced increase in lactate could reflect the
oxidative status of the local muscle area in which microdialysis was performed. Thus, a
release of lactate from muscle cells during exercise, when the oxygen availability is
limited compared to the demand, can be followed by a cellular uptake and conversion
into pyruvate by lactate dehydrogenase, which may be used for aerobic metabolism
during the recovery period when oxygen delivery is more abundant.
46
- Lars Rosendal -
The increased lactate level during and following the static shoulder flexion was
expected, and SUS served as a positive control situation to test the ability of the
microdialysis technique to detect changes in muscle metabolism during exercise. The
increase in lactate was a confirmation of the marked metabolite accumulation
occurring in muscle tissue during intense exercise, which has previously been shown
(MacLean et al., 1999;Linossier et al., 2000;Boschmann et al., 2002).
During the repetitive low-force work as well as the intense static contractions, an
increase in muscle K+ concentration was noted (Figure 8 E). An increase in muscle K+
during exercise, which has repeatedly been shown (Green et al., 1999;Green et al.,
2000;Juel et al., 2000;Lott et al., 2001) is the result of a cellular leakage through K+
channels during excitation. It is interesting, however, that interstitial K+ concentrations
increased to similar levels during the two work protocols despite a large difference in
external work-load. This may further indicate that an inhomogeneous muscle activation
is evident during RLW pointing towards a Cinderella fiber recruitment pattern during
RLW.
5.5 Cytokine response to RLW
The increased metabolic activity during RLW was associated with a substantial
increase in interstitial levels of IL-6 but not IL-1. The cytokines IL-1 and IL-6 have
classically been considered pro-inflammatory cytokines because of the high measured
plasma levels in inflammatory diseases and severe infections (Waage & Steinshamn,
1993), however, recent data suggest that IL-6 may primarily posses metabolic and antiinflammatory characteristics (Pedersen et al., 2001b;Pedersen et al., 2003;Starkie et
al., 2003). When plasma IL-6 was initially shown to increase in response to intense
exercise (Haahr et al., 1991;Nehlsen-Cannarella et al., 1997;Pedersen et al., 1998) it
was suggested to be due to muscle damage from intense eccentric contractions
(Bruunsgaard et al., 1997). In contrast to this idea, increased plasma levels were also
found during non-damaging exercise (Brenner et al., 1999) and it has been shown that
the local production in the exercising muscles can account for the increased plasma
levels (Ostrowski et al., 1998;Steensberg et al., 2000).
The increase in interstitial IL-6 during RLW (Figure 8 C) occurred concomitant with
the increase in local muscle metabolism, which led me to suggest that a contractioninduced increase in metabolism could trigger a production of IL-6. That IL-6 has the
capability to influence metabolism has previously been suggested by Pedersen and coworkers (Pedersen et al., 2001a). They suggested that IL-6 released from muscle to
blood acts in a hormone-like fashion exerting metabolic control. In support of this idea,
47
- Interstitial changes in trapezius muscle during repetitive low-force work -
it has been shown that IL-6 has the ability to induce lipolysis in adipose tissue and to
some extent to increase the hepatic glucose release and thereby increase the total
substrate availability (Stouthard et al., 1995;Tsigos et al., 1997;Kanemaki et al.,
1998;Pedersen et al., 2003). Furthermore, IL-6 has been shown to be released faster
and in larger amounts when the muscle glycogen content is low, supporting its
connection to metabolic status (Keller et al., 2001;MacDonald et al., 2003). The
suggestion that IL-6 acts in a hormone-like fashion is based on the assumption that
plasma IL-6 increases during exercise, as found during intense exercise. However,
plasma IL-6 did not increase significantly in response to RLW (Figure 8 D) despite
that the increase in interstitial IL-6 was of such magnitude (20 fold higher than peak
plasma values during intense exercise) that it potentially should result in an increase in
plasma IL-6. The local effect of IL-6 within the muscle tissue is unknown at present,
but it has been suggested that IL-6 may potentially function as a local metabolic
regulator as well (Pedersen et al., 2001b), which would be consistent with the
simultaneous increases in metabolites found in the present thesis.
Another function of IL-6 may be to exert an anti-inflammatory effect. It has been
shown that IL-6 exerts a negative feedback on tumor necrosis factor-, and it is
suggested that this suppressing effect on low-grade inflammation could be one
mechanism by which exercise has positive health effects (Pedersen & Bruunsgaard,
2003;Starkie et al., 2003). Although highly speculative, the substantial increase in
interstitial IL-6 during low-force contractions could potentially be a link to the positive
health effects of low to moderate intensity exercise on classical diseases such as insulin
resistance, type 2 diabetes, and cardiovascular disease (Blair & Brodney, 1999) that
have been suggested to be caused by low-grade inflammation. Unfortunately it was not
possible to address possible inter-relationship between the anti-inflammatory effect of
IL-6 and the pro-inflammatory effect of IL-1 during low-force work, as it was not
possible to determine the interstitial concentration of IL-1 at any time point. IL-6 has
also been suggested to be released in response to muscle damage (Bruunsgaard et al.,
1997), and it seems to be confirmed by the increase in IL-6 found in response to the
insertion trauma (Figure 6 B), however, this effect was delayed by more than 30 min.
It has previously been shown that a trauma results in increased IL-6 messenger RNA
(Kami & Senba, 1998). It could therefore be speculated that the IL-6 increase during
RLW could mainly be due to an effect of the insertion trauma, but it was shown that
the increase caused by the insertion trauma only accounted for 20-30 % of the exercise
induced increase (Paper II). Of course, it cannot be excluded that the exercise per se
created additional cellular trauma due to friction between the microdialysis catheter
48
- Lars Rosendal -
and the surrounding muscle cells. However, when the slow increase in IL-6 after the
insertion trauma is compared to the steep increase during exercise, it is not likely that
the possible additional trauma during exercise would contribute significantly to the IL6 concentration during exercise. Therefore, it’s concluded that local muscle IL-6 does
respond to trauma by a slow increase but that the major increase seen during exercise is
contributable to the exercise per se.
5.6 Muscle pain and algesic substances in trapezius myalgia
Study C showed that trapezius muscle pain was increased and pressure pain thresholds
reduced in the patients with work-related trapezius myalgia compared with healthy
controls, and that these changes were significantly correlated to interstitial levels of 5HT and glutamate respectively. Furthermore, it was shown that PPT in control sites
(tibialis anterior, Table 4) did not differ between the groups indicating that the muscle
pain was not entirely due to central sensitization. Chronic pain has otherwise been
suggested to result in extensive sensitization of the central nervous system attributable
to changes in function of the neural pathway themselves, rather than peripheral
nociceptive pain (Mense, 1993;Sheather-Reid & Cohen, 1998), however the increased
levels of potential algesic substances in trapezius muscle suggest that peripheral
nociception may persist in chronic work-related trapezius muscle pain.
In the literature, bradykinin and 5-HT have been suggested to be likely candidates for
nociceptor activation in clinical muscle pain (Mense, 1991;Mense, 1993;Millan,
1999;Graven-Nielsen & Mense, 2001) and local injections of these candidates have
been shown to induce muscle pain and hyperalgesia to mechanical stimuli (Jensen et
al., 1990;Babenko et al., 1999;Mørk et al., 2003). Unfortunately, it was not possible to
determine the interstitial concentrations of bradykinin in the present thesis although it
has previously been done in calf muscle with microdialysis (Langberg et al., 2002a). In
contrast, it was possible to determine the interstitial 5-HT level, and this level was
approx. 6-fold higher in patients with WTM than in controls. Interestingly, the
interstitial 5-HT concentration, found in patients with WTM was of the same
magnitude (20 nmol/l) as used in one injection study demonstrating a nociceptive
effect of 5-HT and bradykinin (Babenko et al., 1999), which indicates that the obtained
interstitial level has a physiological relevance in relation to muscle pain. 5-HT alone is
speculated to primarily exert a sensitizing effect, because its effect is greatly increased
in combination with other algesic substances. Therefore it is very interesting that the
interstitial levels of lactate and glutamate, that are also suggested to have nociceptoractivating characteristics, were also significantly increased in patients with WTM.
49
- Interstitial changes in trapezius muscle during repetitive low-force work -
Although the lactate level was increased in WTM, it is unlikely that lactate per se is
important for pain mediation since it only activates muscle nociceptors at
supraphysiological concentrations (Mense, 1993). The role of glutamate in relation to
muscle pain has not been fully elucidated, but it has recently been shown that injection
of glutamate into muscle tissue results in increased pain intensity and lowered PPT
(Cairns et al., 2003;Svensson et al., 2003). It is noteworthy that these studies used
concentrations much higher than observed in muscle tissue in patients with WTM in
study C. However, a significant negative correlation with PPT was demonstrated in
patients with WTM, indicating that glutamate is relevant in relation to muscle pain in
trapezius myalgia. Not all pain syndromes are associated with increased glutamate
levels – identical trapezius muscle levels were found in patients with tension-type
headache and healthy controls (Ashina et al., 2003). It is likely however, that the
etiology of pain in trapezius muscle is not comparable in WTM and chronic tensiontype headache. In support of this, chronic tension-type headache has been suggested to
primarily be due to central sensitization from prolonged input from e.g. tender points
(Bendtsen & Ashina, 2000). On the other hand, increased peripheral glutamate levels
have been found in other clinical pain disorders (Alfredson et al., 1999;Alfredson et
al., 2000;Alfredson et al., 2001b) and it has been demonstrated that centrally,
glutamate acts through NMDA receptors, which have recently been demonstrated to be
present in peripheral tissue also (Alfredson et al., 2001a;Alfredson et al., 2001b).
It should be kept in mind that chronic pain is multifactorial in concept, and that
psychosocial factors may also significantly contribute to pain mediation and
perception, however, it is very interesting that the present thesis demonstrates that
work-related trapezius myalgia is associated with increased interstitial muscle levels of
potential algesic substances, indicating the possibility of peripherally activated
nociceptive processes in muscle tissue.
50
- Lars Rosendal -
7. CONCLUSION
In conclusion, the present thesis demonstrated that the microdialysis technique enabled
measurement of low and high molecular weight substances related to metabolism,
cytokine response, and algesics in trapezius muscle in response to repetitive low-force
work.
It was demonstrated that the trapezius muscle accumulates metabolites during 20 min
of repetitive low-force work. Even though the muscle activity level was below 10 % of
max, an increase in lactate was found, indicating that anaerobic metabolism is
accelerated.
The cytokine interleukin-6, which has been speculated to have important metabolic and
anti-inflammatory properties, increased substantially during repetitive low-force work
and the increase could only to a minor degree be explained by the insertion trauma per
se.
Finally, and perhaps most important of all, work-related trapezius myalgia was shown
to be associated with increased anaerobic metabolism as well as increased levels of
potential algesic substances locally in the painful muscle tissue – indicating that
peripheral nociceptive processes may be activated. These changes were not associated
with a reduced blood flow response in the patients with trapezius myalgia.
51
- Interstitial changes in trapezius muscle during repetitive low-force work -
52
- Lars Rosendal -
8. PERSPECTIVE
Some novel findings have emerged from the present thesis, however, as it is in science
– solving one question immediately leads to five new questions asked – therefore some
ideas on how to proceed from here are given…
It would be interesting to create an experimental model system in which temporary
changes, similar to those found in patients with work-related trapezius myalgia, could
be induced and studied. This may be done by investigating whether exercise-induced
muscle damage (DOMS), resulting in localized muscle pain would result in local
increases in metabolism and/or potential algesic substances. The purpose of such a
study is to investigate if DOMS can provide an experimental pain model in which
changes, pathomechanisms, and preventive treatments could be studied.
The findings of increased levels of potentially algesic substances in painful muscle
tissue of patients with trapezius myalgia give rise to a line of new studies to be
conducted, e.g.:
 It could be investigated whether painful muscle tissue from occupational work
also shows signs of increased levels of other potential algesic substances such
as substance P, potassium, prostaglandin’s, or calcitonin gene related peptide
(CGRP).
 Furthermore, to study the causal relationship between increased levels of
algesics and muscle pain, it could be studied whether local blocking of the
algesic substances, by using the microdialysis technique, has an affect on pain
perception.
Finally, the potential of the microdialysis technique may be explored further. For
instance it is desirable to be able to conduct microdialysis in muscle tissue during “real
occupational work” with a duration of several hours or maybe even days. This would
require that the technique is modified in order to adopt it to portable pumps etc. In
collaboration with my Swedish colleges (Björn Gerdle and Britt Larsson), an initial
attempt to perform microdialysis in freely moving humans by using portable pumps
has been performed, and the results are promising.
53
- Interstitial changes in trapezius muscle during repetitive low-force work -
54
- Lars Rosendal -
ACKNOWLEDGEMENTS
I would like to express my sincere gratitude to everyone who have helped me take my
first (staggering) steps into the world of science – I feel convinced that without your
support I would not have gotten half the way, and I am certain that it would not have
been as joyful.
I am particular grateful to:
-
-
-
PhD, Jesper Kristiansen, who has patiently served as my mentor and
“biochemistry guide”. Thank you for always believing in me, supporting me,
and for always being willing to go the extra mile with me.
Professor, Dr. med, Michael Kjær, for getting me started as well as for
competent mentorship and efficient help writing the papers.
PhD, Karen Søgaard for initiating this PhD project and for providing me with
competent mentorship, even though you were not officially given any credit.
Moreover, thanks for all your smiles, laughs, enthusiasm, and physiological
knowledge.
PhD, Henning Langberg, for providing mentorship and smiles when it was
most needed and for initially teaching me about microdialysis.
Also I would like to extent my warm thanks to:
-
-
-
-
Professor, Björn Gerdle and PhD, Britt Larsson for your excellent
collaboration, support and catching enthusiasm. Also, thanks for welcoming
me in your laboratory. I hope that our future collaboration will be as fruitful as
it has begun.
Professor, Dr.med.Sc, Gisela Sjøgaard, for inspiring physiological discussions
and valuable manuscript comments.
Colleagues at the National Institute of Occupational Health and the Sports
Medicine Research Unit, especially the PhD groups – you showed me that a
life with science is not only competitive and tough, but also warm and caring –
thanks for all the smiles and laughter we shared.
Anne Abildtrup for your great humor and for providing skilled technical
assistance with the biochemical analyses, and Bodil Holst for linguistic help as
well as for posing as a model on the cover.
Last, but definitely not least, I want to thank all the participants for making this
thesis possible – I am very grateful for your contributions.
55
- Interstitial changes in trapezius muscle during repetitive low-force work -
Finally I would like to express my warm thanks to the most colorful and supportive of
all persons - my wife Rie-Emilia. If it was not for you, your faith in me, and for all
your support when things did not turn out the way I wanted, these three years would
have been much harder than they have actually been…
56
- Lars Rosendal -
REFERENCE LIST
Abramovici, A., Daizade, I., Yosipovitch, Z., Gibson, S. J., & Polak, J. M. (1991). The distribution of peptidecontaining nerves in the synovia of the cat knee joint. Histol.Histopathol. 6, 469-476.
Alfredson, H., Forsgren, S., Thorsen, K., Fahlstrom, M., Johansson, H., & Lorentzon, R. (2001a). Glutamate NMDAR1
receptors localised to nerves in human Achilles tendons. Implications for treatment? Knee.Surg.Sports
Traumatol.Arthrosc. 9, 123-126.
Alfredson, H., Forsgren, S., Thorsen, K., & Lorentzon, R. (2001b). In vivo microdialysis and immunohistochemical
analyses of tendon tissue demonstrated high amounts of free glutamate and glutamate NMDAR1 receptors, but
no signs of inflammation, in Jumper's knee. J Orthop.Res. 19, 881-886.
Alfredson, H., Ljung, B. O., Thorsen, K., & Lorentzon, R. (2000). In vivo investigation of ECRB tendons with
microdialysis technique--no signs of inflammation but high amounts of glutamate in tennis elbow. Acta Orthop
Scand 71, 475-479.
Alfredson, H., Thorsen, K., & Lorentzon, R. (1999). In situ microdialysis in tendon tissue: high levels of glutamate, but
not prostaglandin E2 in chronic Achilles tendon pain. Knee.Surg.Sports Traumatol.Arthrosc. 7, 378-381.
Arner, P., Bolinder, J., Eliasson, A., Lundin, A., & Ungerstedt, U. (1988). Microdialysis of adipose tissue and blood for
in vivo lipolysis studies. Am.J.Physiol. 255, E737-E742.
Ashina, M., Stallknecht, B., Bendtsen, L., Pedersen, J. F., Galbo, H., Dalgaard, P., & Olesen, J. (2002). In vivo evidence
of altered skeletal muscle blood flow in chronic tension-type headache. Brain 125, 320-326.
Ashina, M., Stallknecht, B., Bendtsen, L., Pedersen, J. F., Schifter, S., Galbo, H., & Olesen, J. (2003). Tender points are
not sites of ongoing inflammation -in vivo evidence in patients with chronic tension-type headache. Cephalalgia
23, 109-116.
Axelson, H. W., Melberg, A., Ronquist, G., & Askmark, H. (2002). Microdialysis and electromyography of
experimental muscle fatigue in healthy volunteers and patients with mitochondrial myopathy. Muscle.Nerve. 26,
520-526.
Babenko, V., Graven-Nielsen, T., Svensson, P., Drewes, A. M., Jensen, T. S., & Arendt-Nielsen, L. (1999).
Experimental human muscle pain and muscular hyperalgesia induced by combinations of serotonin and
bradykinin. Pain 82, 1-8.
Barr, A. E. & Barbe, M. F. (2004). Inflammation reduces physiological tissue tolerance in the development of work related musculoskeletal disorders. J Electromyogr.Kinesiol. 14, 77-85.
Bendtsen, L. & Ashina, M. (2000). Sensitization of myofascial pain pathways in tension-type headache. In The
headaches, eds. Olesen, J., Tfelt-hansen, P., & Welch, K. M. A., pp. 573-577. Lippincott, Williams & Wilkins,
Philadelphia.
Bernard, B. (1997). Musculoskeletal disorders and workplace factors: A critical review of epidemiologic evidence for
work-related musculoskeletal disorders of the neck, upper extremity, and low back, 2 ed., pp. I-C-59. U.S.
Department of Health and Human Services, NIOSH, Cincinnati, USA.
Birch, L., Christensen, H., Arendt-Nielsen, L., Graven-Nielsen, T., & Sogaard, K. (2000). The influence of
experimental muscle pain on motor unit activity during low-level contraction. Eur.J Appl.Physiol 83, 200-206.
57
- Interstitial changes in trapezius muscle during repetitive low-force work Birklein, F., Weber, M., & Neundorfer, B. (2000). Increased skin lactate in complex regional pain syndrome: evidence
for tissue hypoxia? Neurology 55, 1213-1215.
Bito, L., Davson, H., Levin, E., Murray, M., & Snider, N. (1966). The concentrations of free amino acids and other
electrolytes in cerebrospinal fluid, in vivo dialysate of brain, and blood plasma of the dog. J.Neurochem. 13,
1057-1067.
Blair, S. N. & Brodney, S. (1999). Effects of physical inactivity and obesity on morbidity and mortality: current
evidence and research issues. Med Sci Sports Exerc 31, S646-S662.
Blais, C., Jr., Adam, A., Massicotte, D., & Peronnet, F. (1999). Increase in blood bradykinin concentration after
eccentric weight-training exercise in men. J.Appl.Physiol 87, 1197-1201.
Blangsted, A. K., Hansen, K., & Jensen, C. (2003). Muscle activity during computer-based office work in relation to
self-reported job demands and gender. Eur.J.Appl.Physiol 89, 352-358.
Boix, F., Rosenborg, L., Hilgenfeldt, U., & Knardahl, S. (2002). Contraction-related factors affect the concentration of a
kallidin-like peptide in rat muscle tissue. J.Physiol 544, 127-136.
Bolinder, J., Ungerstedt, U., & Arner, P. (1992). Microdialysis measurement of the absolute glucose concentration in
subcutaneous adipose tissue allowing glucose monitoring in diabetic patients. Diabetologia 35, 1177-1180.
Borg, G. A. (1982). Psychophysical bases of perceived exertion. Med.Sci.Sports Exerc. 14, 377-381.
Boschmann, M., Rosenbaum, M., Leibel, R. L., & Segal, K. R. (2002). Metabolic and hemodynamic responses to
exercise in subcutaneous adipose tissue and skeletal muscle. Int J Sports Med 23, 537-543.
Brenner, I. K., Natale, V. M., Vasiliou, P., Moldoveanu, A. I., Shek, P. N., & Shephard, R. J. (1999). Impact of three
different types of exercise on components of the inflammatory response. Eur.J.Appl.Physiol Occup.Physiol 80,
452-460.
Bruunsgaard, H., Galbo, H., Halkjaer-Kristensen, J., Johansen, T. L., MacLean, D. A., & Pedersen, B. K. (1997).
Exercise-induced increase in serum interleukin-6 in humans is related to muscle damage. Journal of Physiology
499, 833-841.
Bureau of Labor Statistics. Nonfatal illnesses cases by selected categories, private industry, 2000. 2001. US
Department of Labour, Bureau of Labour Statistics, Washington, DC.
Cairns, B. E., Svensson, P., Wang, K., Hupfeld, S., Graven-Nielsen, T., Sessle, B. J., Berde, C. B., & Arendt-Nielsen,
L. (2003). Activation of peripheral NMDA receptors contributes to human pain and rat afferent discharges
evoked by injection of glutamate into the masseter muscle. J.Neurophysiol. 90, 2098-2105.
Christensen, H. (1986). Muscle activity and fatigue in the shoulder muscles of assembly-plant employees. Scand.J.Work
Environ.Health 12, 582-587.
Coggeshall, R. E. & Carlton, S. M. (1998). Ultrastructural analysis of NMDA, AMPA, and kainate receptors on
unmyelinated and myelinated axons in the periphery. J Comp Neurol. 391 , 78-86.
Collins, S. L., Moore, R. A., & McQuay, H. J. (1997). The visual analogue pain intensity scale: what is moderate pain
in millimetres? Pain 72, 95-97.
Delgado, J. M., DeFeudis, F. V., Roth, R. H., Ryugo, D. K., & Mitruka, B. M. (1972). Dialytrode for long term
intracerebral perfusion in awake monkeys. Arch.Int.Pharmacodyn.Ther. 198, 9-21.
Dray, A. & Perkins, M. (1993). Bradykinin and inflammatory pain. Trends Neurosci. 16, 99-104.
Edwards, R. H. (1988). Hypotheses of peripheral and central mechanisms underlying occupational muscle pain and
injury. Eur.J.Appl.Physiol 57, 275-281.
58
- Lars Rosendal Edwards, R. H., Newham, D. J., Jones, D. A., & Chapman, S. J. (1984). Role of mechanical damage in pathogenesis of
proximal myopathy in man. Lancet 1, 548-552.
Ernberg, M., Hedenberg-Magnusson, B., Alstergren, P., & Kopp, S. (1999). The level of serotonin in the superficial
masseter muscle in relation to local pain and allodynia. Life Sci 65, 313-325.
Fitts, R. H. (1994). Cellular mechanisms of muscle fatigue. Physiol Rev. 74, 49-94.
Fredriksson, K., Alfredsson, L., Thorbjornsson, C. B., Punnett, L., Toomingas, A., Torgen, M., & Kilbom, A. (2000).
Risk factors for neck and shoulder disorders: A nested case-control study covering a 24-year period.
Am.J.Ind.Med. 38, 516-528.
Gissel, H. (2000). Ca2+ accumulation and cell damage in skeletal muscle during low frequency stimulation. European
Journal of Applied Physiology and Occupational Physiology 83, 175-180.
Gissel, H. & Clausen, T. (2001). Excitation-induced Ca2+ influx and skeletal muscle cell damage. Acta Physiol Scand.
171, 327-334.
Gissel, H. & Clausen, T. (2003). Ca2+ uptake and cellular integrity in rat EDL muscle exposed to electrostimulation,
electroporation, or A23187. Am.J.Physiol Regul.Integr.Comp Physiol 285, R132-R142.
Graven-Nielsen, T. & Mense, S. (2001). The peripheral apparatus of muscle pain: evidence from animal and human
studies. Clin J Pain 17, 2-10.
Green, S., Bulow, J., & Saltin, B. (1999). Microdialysis and the measurement of muscle interstitial K+ during rest and
exercise in humans. J.Appl.Physiol. 87, 460-464.
Green, S., Langberg, H., Skovgaard, D., Bulow, J., & Kjaer, M. (2000). Interstitial and arterial-venous [K+] in human
calf muscle during dynamic exercise: effect of ischaemia and relation to muscle pain. J Physiol. 529 Pt 3, 849861.
Haahr, P. M., Pedersen, B. K., Fomsgaard, A., Tvede, N., Diamant, M., Klarlund, K., Halkjaer-Kristensen, J., &
Bendtzen, K. (1991). Effect of physical exercise on in vitro production of interleukin 1, interleukin 6, tumour
necrosis factor-alpha, interleukin 2 and interferon-gamma. Int.J Sports Med 12, 223-227.
Hagberg, M. (1996). ABC of work related disorders. Neck and arm disorders. BMJ 313, 419-422.
Hägg, G. M. (1991). Static work loads and occupational myalgia - a new explanation model. In Electromyographical
Kinesiology, eds. Anderson, P. A., Hobart, D. J., & Danoff, J. V., pp. 141-144. Elsevier Science Publishers B.V.,
Amsterdam.
Hägg, G. M. (2000). Human muscle fibre abnormalities related to occupational load. European Journal of Applied
Physiology and Occupational Physiology 83, 159-165.
Hargreaves, K. M., Roszkowski, M. T., & Swift, J. Q. (1993). Bradykinin and inflammatory pain. Agents Actions Suppl
41, 65-73.
Henneman, E. & Olson, C. B. (1965). Relations between structure and function in the design of skeletal muscles.
Journal of Neurophysiology 28, 581-598.
Henriksson, J. (1999). Microdialysis of skeletal muscle at rest. Proc.Nutr.Soc. 58, 919-923.
Henriksson, K. G. (1988). Muscle pain in neuromuscular disorders and primary fibromyalgia. European Journal of
Applied Physiology and Occupational Physiology 57, 348-352.
Hickner, R. C., Bone, D., Ungerstedt, U., Jorfeldt, L., & Henriksson, J. (1994). Muscle blood flow during intermittent
exercise: comparison of the microdialysis ethanol technique and 133Xe clearance. Clin.Sci.(Colch.) 86, 15-25.
59
- Interstitial changes in trapezius muscle during repetitive low-force work Jackson, M. J., Jones, D. A., & Edwards, R. H. (1985). Vitamin E and muscle diseases. J Inherit.Metab Dis. 8 Suppl 1,
84-87.
Jacobson, I., Sandberg, M., & Hamberger, A. (1985). Mass transfer in brain dialysis devices--a new method for the
estimation of extracellular amino acids concentration. J.Neurosci.Methods 15, 263-268.
Jensen, B. R., Fallentin, N., Byström, S., & Sjøgaard, G. (1993a). Plasma potassium concentration and doppler blood
flow during and following submaximal handgrip contractions. Acta Physiologica Scandinavica 147, 203-211.
Jensen, B. R., Schibye, B., Søgaard, K., Simonsen, E. B., & Sjøgaard, G. (1993b). Shoulder muscle load and muscle
fatigue among industrial sewing-machine operators. Eur J Appl Physiol Occup Physiol 67, 467-475.
Jensen, C. (2003). Development of neck and hand-wrist symptoms in relation to duration of computer use at work.
Scand J Work Environ.Health 29, 197-205.
Jensen, C., Borg, V., Finsen, L., Hansen, K., Juul-Kristensen, B., & Christensen, H. (1998). Job demands, muscle
activity and musculoskeletal symptoms in relation to work with the computer mouse. Scand J Work
Environ.Health 24, 418-424.
Jensen, C., Laursen, B., & Sjøgaard, G. (1999). Shoulder and neck. In Biomechanics in Ergonomics, ed. Kumar, S., pp.
201-220. Taylor & Francis, London Philadelphia.
Jensen, C., Vasseljen, O., & Westgaard, R. H. (1993c). The influence of electrode position on bipolar surface
electromyogram recordings of the upper trapezius muscle. European Journal of Applied Physiology and
Occupational Physiology 67, 266-273.
Jensen, K., Tuxen, C., Pedersen-Bjergaard, U., Jansen, I., Edvinsson, L., & Olesen, J. (1990). Pain and tenderness in
human temporal muscle induced by bradykinin and 5-hydroxytryptamine. Peptides 11, 1127-1132.
Johansson, H., Arendt-Nielsen, L., Bergenheim, M., Blair, S., van Dieen, J., Djupsjöbacka, M., Fallentin, N., Gold, J.
E., Hägg, G., Kalezic, N., Larsson, S.-E., Ljubisavljevic, M., Lyskov, E., Mano, T., Magnusson, M., Passatore,
M., Pedrosa-Domellöf, F., Punnett, L., Roatta, S., Thornell, L.-E., Windhorst, U., & Zukowska, Z. (2003a).
Epilogue: An integrated model for chronic work-related myalgia "Brussels Model". In Chronic work-related
myalgia - Neuromuscular mechanisms behind work-related chronic muscle pain syndromes, eds. Johansson, H.,
Windhorst, U., Djupsjöbacka, M., & Passatore, M., pp. 291-300. Centre for Musculoskeletal Research,
University of Gävle, Umeå, Sweden.
Johansson, H. & Sojka, P. (1991). Pathophysiological mechanisms involved in genesis and spread of muscular tension
in occupational muscle pain and in chronic musculoskeletal pain syndromes: a hypothesis. Med Hypotheses. 35,
196-203.
Johansson, H., Windhorst, U., Djupsjöbacka, M., & Passatore, M. (2003b). Chronic work-related myalgia, pp. 1-310.
Centre for Musculoskeletal Research, University of Gävle, Umeå, Sweden.
Juel, C., Pilegaard, H., Nielsen, J. J., & Bangsbo, J. (2000). Interstitial K(+) in human skeletal muscle during and after
dynamic graded exercise determined by microdialysis. Am.J Physiol.Regul.Integr.Comp.Physiol. 278, R400R406.
Kadi, F., Waling, K., Ahlgren, C., Sundelin, G., Holmner, S., Butler-Browne, G. S., & Thornell, L. E. (1998).
Pathological mechanisms implicated in localized female trapezius myalgia. Pain 78, 191-196.
Kami, K. & Senba, E. (1998). Localization of leukemia inhibitory factor and interleukin-6 messenger ribonucleic acids
in regenerating rat skeletal muscle. Muscle Nerve 21, 819-822.
60
- Lars Rosendal Kanemaki, T., Kitade, H., Kaibori, M., Sakitani, K., Hiramatsu, Y., Kamiyama, Y., Ito, S., & Okumura, T. (1998).
Interleukin 1beta and interleukin 6, but not tumor necrosis factor alpha, inhibit insulin-stimulated glycogen
synthesis in rat hepatocytes. Hepatology 27, 1296-1303.
Keller, C., Steensberg, A., Pilegaard, H., Osada, T., Saltin, B., Pedersen, B. K., & Neufer, P. D. (2001). Transcriptional
activation of the IL-6 gene in human contracting skeletal muscle: influence of muscle glycogen content. FASEB J
15, 2748-2750.
Kniffki, K. D., Mense, S., & Schmidt, R. F. (1978). Responses of group IV afferent units from skeletal muscle to
stretch, contraction and chemical stimulation. Exp.Brain Res. 31, 511-522.
Kuorinka, I., Jonsson, B., Kilbom, Å., Vinterberg, H., Biering-Sørensen, F., Andersson, G., & Jørgensen, K. (1987).
Standardised Nordic questionnaires for the analysis of musculoskeletal symptoms. Applied Ergonomics 18, 233237.
Langberg, H. The human Achilles tendon. Circulatory and metabolic changes with exercise. 1999. University of
Copenhagen. (PhD Thesis)
Langberg, H., Bjorn, C., Boushel, R., Hellsten, Y., & Kjaer, M. (2002a). Exercise-induced increase in interstitial
bradykinin and adenosine concentrations in skeletal muscle and peritendinous tissue in humans. J Physiol 542,
977-983.
Langberg, H., Olesen, J. L., Gemmer, C., & Kjaer, M. (2002b). Substantial elevation of interleukin-6 concentration in
peritendinous tissue, in contrast to muscle, following prolonged exercise in humans. J Physiol 542, 985-990.
Langberg, H., Rosendal, L., & Kjaer, M. (2001). Training-induced changes in peritendinous type I collagen turnover
determined by microdialysis in humans. J Physiol 534, 297-302.
Langberg, H., Skovgaard, D., Karamouzis, M., Bulow, J., & Kjaer, M. (1999). Metabolism and inflammatory mediators
in the peritendinous space measured by microdialysis during intermittent isometric exercise in humans.
J.Physiol.(Lond.) 515, 919-927.
Larsson, B., björk, J., Kadi, F., Lindman, R., & Gerdle, B. Capillary supply and moth eaten fibres in cleaners with and
without work-related trapezius myalgia and in healthy controls. Clin J Pain . 2004. In Press
Larsson, B., Bjork, J., Elert, J., & Gerdle, B. (2000a). Mechanical performance and electromyography during repeated
maximal isokinetic shoulder forward flexions in female cleaners with and without myalgia of the trapezius
muscle and in healthy controls. European Journal of Applied Physiology and Occupational Physiology 83, 257267.
Larsson, B., Bjork, J., Henriksson, K. G., Gerdle, B., & Lindman, R. (2000b). The prevalences of cytochrome c oxidase
negative and superpositive fibres and ragged-red fibres in the trapezius muscle of female cleaners with and
without myalgia and of female healthy controls. Pain 84, 379-387.
Larsson, R., Oberg, P. A., & Larsson, S. E. (1999). Changes of trapezius muscle blood flow and electromyography in
chronic neck pain due to trapezius myalgia. Pain 79, 45-50.
Larsson, S. E., Bengtsson, A., Bodegard, L., Henriksson, K. G., & Larsson, J. (1988). Muscle changes in work-related
chronic myalgia. Acta Orthop.Scand 59, 552-556.
Larsson, S. E., Bodegard, L., Henriksson, K. G., & Oberg, P. A. (1990). Chronic trapezius myalgia. Morphology and
blood flow studied in 17 patients. Acta Orthop.Scand 61, 394-398.
Levine, J. D. & Reichling, D. B. (1999). Peripheral mechanisms of inflammatory pain. In Textbook of pain, eds. Wall,
P. D. & Melzak, R., pp. 59-84. Churchill Livingstone, Edinburg.
61
- Interstitial changes in trapezius muscle during repetitive low-force work Lindman, R. (1992). Chronic trapezius myalgia - a morphological study. Arbete och Hälsa 34, 1-46.
Linossier, M. T., Dormois, D., Arsac, L., Denis, C., Gay, J. P., Geyssant, A., & Lacour, J. R. (2000). Effect of
hyperoxia on aerobic and anaerobic performances and muscle metabolism during maximal cycling exercise.
Acta Physiologica Scandinavica 168, 403-411.
Lönnroth, P. (1997). Microdialysis in adipose tissue and skeletal muscle. Horm.Metab.Res. 29, 344-346.
Lönnroth, P., Jansson, P. A., & Smith, U. (1987). A microdialysis method allowing characterization of intercellular
water space in humans. Am.J.Physiol. 253, E228-E231.
Lönnroth, P. & Strindberg, L. (1995). Validation of the 'internal reference technique' for calibrating microdialysis
catheters in situ. Acta Physiol.Scand. 153, 375-380.
Lott, M. E., Hogeman, C. S., Vickery, L., Kunselman, A. R., Sinoway, L. I., & MacLean, D. A. (2001). Effects of
dynamic exercise on mean blood velocity and muscle interstitial metabolite responses in humans. Am.J
Physiol.Heart.Circ.Physiol. 281, H1734-H1741.
Lund, J. P., Donga, R., Widmer, C. G., & Stohler, C. S. (1991). The pain-adaptation model: a discussion of the
relationship between chronic musculoskeletal pain and motor activity. Canadian Journal of Physiology and
Pharmacology 69, 683-694.
Lundberg, G., Olofsson, P., Ungerstedt, U., Jansson, E., & Sundberg, J. (2002). Lactate concentrations in human
skeletal muscle biopsy, microdialysate and venous blood during dynamic exercise under blood flow restriction.
Pflugers.Arch 443, 458-465.
MacDonald, C., Wojtaszewski, J. F., Pedersen, B. K., Kiens, B., & Richter, E. A. (2003). Interleukin-6 release from
human skeletal muscle during exercise: relation to AMPK activity. J.Appl.Physiol 95, 2273-2277.
MacLean, D. A., Bangsbo, J., & Saltin, B. (1999). Muscle interstitial glucose and lactate levels during dynamic exercise
in humans determined by microdialysis. J Appl Physiol 87, 1483-1490.
MacLean, D. A., LaNoue, K. F., Gray, K. S., & Sinoway, L. I. (1998). Effects of hindlimb contraction on pressor and
muscle interstitial metabolite responses in the cat. J Appl Physiol. 85, 1583-1592.
McNeil, P. L. & Steinhardt, R. A. (1997). Loss, restoration, and maintenance of plasma membrane integrity. J Cell Biol.
137, 1-4.
Mense, S. (1991). Considerations concerning the neurobiological basis of muscle pain. Canadian Journal of Physiology
and Pharmacology 69, 610-616.
Mense, S. (1993). Nociception from skeletal muscle in relation to clinical muscle pain. Pain 54, 241-289.
Meyerson, B. A., Linderoth, B., Karlsson, H., & Ungerstedt, U. (1990). Microdialysis in the human brain: extracellular
measurements in the thalamus of parkinsonian patients. Life Sci 46, 301-308.
Millan, M. J. (1999). The induction of pain: an integrative review. Prog.Neurobiol. 57, 1-164.
Mørk, H., Ashina, M., Bendtsen, L., Olesen, J., & Jensen, R. (2003). Experimental muscle pain and tenderness
following infusion of endogenous substances in humans. Eur.J.Pain 7, 145-153.
Nehlsen-Cannarella, S. L., Fagoaga, O. R., Nieman, D. C., Henson, D. A., Butterworth, D. E., Schmitt, R. L., Bailey, E.
M., Warren, B. J., Utter, A., & Davis, J. M. (1997). Carbohydrate and the cytokine response to 2.5 h of running. J
Appl Physiol 82, 1662-1667.
Nielsen, O. B., de Paoli, F., & Overgaard, K. (2001). Protective effects of lactic acid on force production in rat skeletal
muscle. J Physiol. 536, 161-166.
62
- Lars Rosendal Nordander, C., Hansson, G. A., Rylander, L., Asterland, P., Bystrom, J. U., Ohlsson, K., Balogh, I., & Skerfving, S.
(2000). Muscular rest and gap frequency as EMG measures of physical exposure: the impact of work tasks and
individual related factors. Ergonomics 43, 1904-1919.
O'Kane, K. P., Webb, D. J., Collier, J. G., & Vallance, P. J. (1994). Local L-NG-monomethyl-arginine attenuates the
vasodilator action of bradykinin in the human forearm. Br.J Clin Pharmacol. 38, 311-315.
Ohlsson, K., Attewell, R. G., Johnsson, B., Ahlm, A., & Skerfving, S. (1994). An assessment of neck and upper
extremity disorders by questionnaire and clinical examination. Ergonomics 37, 891-897.
Ostrowski, K., Rohde, T., Zacho, M., Asp, S., & Pedersen, B. K. (1998). Evidence that interleukin-6 is produced in
human skeletal muscle during prolonged running. J Physiol 508, 949-953.
Pedersen, B. K. & Bruunsgaard, H. (2003). Possible beneficial role of exercise in modulating low-grade inflammation
in the elderly. Scand.J.Med.Sci.Sports 13, 56-62.
Pedersen, B. K., Ostrowski, K., Rohde, T., & Bruunsgaard, H. (1998). The cytokine response to strenuous exercise.
Can.J.Physiol Pharmacol. 76, 505-511.
Pedersen, B. K., Steensberg, A., Keller, P., Keller, C., Fischer, C., Hiscock, N., van Hall, G., Plomgaard, P., &
Febbraio, M. A. (2003). Muscle-derived interleukin-6: lipolytic, anti-inflammatory and immune regulatory
effects. Pflugers Arch. 446, 9-16.
Pedersen, B. K., Steensberg, A., & Schjerling, P. (2001a). Exercise and interleukin-6. Curr.Opin.Hematol. 8, 137-141.
Pedersen, B. K., Steensberg, A., & Schjerling, P. (2001b). Muscle-derived interleukin-6: possible biological effects. J
Physiol 536, 329-337.
Persson, A. L., Hansson, G. A., Kalliomaki, A., Moritz, U., & Sjolund, B. H. (2000). Pressure pain thresholds and
electromyographically defined muscular fatigue induced by a muscular endurance test in normal women. Clin J
Pain 16, 155-163.
Rosdahl, H., Hamrin, K., Ungerstedt, U., & Henriksson, J. (1998). Metabolite levels in human skeletal muscle and
adipose tissue studied with microdialysis at low perfusion flow. Am J Physiol 274, E936-E945.
Rosendal, L. Military training - the effects of 12 weeks of military basic training on collagen type I metabolism and
adenosine, physical performance and incidence of injury in groups of soldiers with different physical fitness
level. 2000. University of Copenhagen. (Master Thesis)
Rotto, D. M. & Kaufman, M. P. (1988). Effect of metabolic products of muscular contraction on discharge of group III
and IV afferents. J Appl.Physiol 64, 2306-2313.
Scheller, D. & Kolb, J. (1991). The internal reference technique in microdialysis: a practical approach to monitoring
dialysis efficiency and to calculating tissue concentration from dialysate samples. J.Neurosci.Methods 40, 31-38.
Sejersted, O. M. & Sjøgaard, G. (2000). Dynamics and consequences of potassium shifts in skeletal muscle and heart
during exercise. Physiol.Rev 80, 1411-1481.
Sheather-Reid, R. B. & Cohen, M. L. (1998). Psychophysical evidence for a neuropathic component of chronic neck
pain. Pain. 75, 341-347.
Sjøgaard, G. (1988). Muscle energy metabolism and electrolyte shifts during low-level prolonged static contraction in
man. Acta Physiologica Scandinavica 134, 181-187.
Sjøgaard, G., Lundberg, U., & Kadefors, R. (2000). The role of muscle activity and mental load in the development of
pain and degenerative processes at the muscle cell level during computer work. European Journal of Applied
Physiology and Occupational Physiology 83, 99-105.
63
- Interstitial changes in trapezius muscle during repetitive low-force work Sjøgaard, G., Sejersted, O. M., Winkel, J., Smolander, J., Jørgensen, K., & Westgaard, R. (1995). Exposure assessment
and mechanisms of pathogenesis in work-related musculoskeletal disorders: Significant aspects in the
documentation of risk factors. In Work and health. Scientific basis of progress in the working environment., eds.
Svane, O. & Johansen, C., pp. 75-87. European Commission, Directorate-General V, Luxembourg.
Sjøgaard, G. & Søgaard, K. (1998). Muscle injury in repetitive motion disorders. Clin Orthop 21-31.
Søgaard, K. (1995). Motor unit recruitment pattern during low-level static and dynamic contractions. Muscle Nerve 18,
292-300.
Stallknecht, B. Influence of physical training on adipose tissue metabolism - with special focus on effects of insulin and
epinephrine. 1-84. 2003. Department of Medical Physiology. (Doctoral Dissertation)
Stallknecht, B., Donsmark, M., Enevoldsen, L. H., Fluckey, J. D., & Galbo, H. (1999). Estimation of rat muscle blood
flow by microdialysis probes perfused with ethanol, [14C]ethanol, and 3H2O. J Appl Physiol 86, 1054-1061.
Stallknecht, B., Lorentsen, J., Enevoldsen, L. H., Bulow, J., Biering-Sorensen, F., Galbo, H., & Kjaer, M. (2001). Role
of the sympathoadrenergic system in adipose tissue metabolism during exercise in humans. J Physiol 536, 283294.
Starkie, R., Ostrowski, S. R., Jauffred, S., Febbraio, M., & Pedersen, B. K. (2003). Exercise and IL-6 infusion inhibit
endotoxin-induced TNF-alpha production in humans. FASEB J. 17, 884-886.
Steensberg, A., van Hall, G., Osada, T., Sacchetti, M., Saltin, B., & Klarlund, P. B. (2000). Production of interleukin-6
in contracting human skeletal muscles can account for the exercise-induced increase in plasma interleukin-6. J
Physiol 529 Pt 1, 237-242.
Stouthard, J. M., Romijn, J. A., Van der, P. T., Endert, E., Klein, S., Bakker, P. J., Veenhof, C. H., & Sauerwein, H. P.
(1995). Endocrinologic and metabolic effects of interleukin-6 in humans. Am.J.Physiol 268, E813-E819.
Svensson, P., Cairns, B. E., Wang, K., Hu, J. W., Graven-Nielsen, T., Arendt-Nielsen, L., & Sessle, B. J. (2003).
Glutamate-evoked pain and mechanical allodynia in the human masseter muscle. Pain 101, 221-227.
Thorsen, K., Kristoffersson, A. O., Lerner, U. H., & Lorentzon, R. P. (1996). In situ microdialysis in bone tissue.
Stimulation of prostaglandin E2 release by weight-bearing mechanical loading. J Clin Invest 98, 2446-2449.
Travell JG, Rinzler S, & Herman M (1942). Pain and disability of the shoulder and arm. JAMA 417-422.
Tsigos, C., Papanicolaou, D. A., Kyrou, I., Defensor, R., Mitsiadis, C. S., & Chrousos, G. P. (1997). Dose-dependent
effects of recombinant human interleukin-6 on glucose regulation. J.Clin.Endocrinol.Metab 82, 4167-4170.
Ungerstedt, U. (1991). Microdialysis - principles and applications for studies in animals and man. J.Intern.Med. 230,
365-373.
Ungerstedt, U. & Hallstrom, A. (1987). In vivo microdialysis - a new approach to the analysis of neurotransmitters in
the brain. Life Sci. 41, 861-864.
Ungerstedt, U. & Pycock, C. (1974). Functional correlates of dopamine neurotransmission.
Bull.Schweiz.Akad.Med.Wiss. 30, 44-55.
Vøllestad, N. K., Sejersted, O. M., Bahr, R., Woods, J. J., & Bigland-Ritchie, B. (1988). Motor drive and metabolic
responses during repeated submaximal contractions in humans. J Appl.Physiol 64, 1421-1427.
von Duvillard, S. P. (2001). Exercise lactate levels: simulation and reality of aerobic and anaerobic metabolism. Eur J
Appl Physiol. 86, 3-5.
Waage, A. & Steinshamn, S. (1993). Cytokine mediators of septic infections in the normal and granulocytopenic host.
Eur.J.Haematol. 50, 243-249.
64
- Lars Rosendal Westerblad, H., Bruton, J. D., Allen, D. G., & Lannergren, J. (2000). Functional significance of Ca2+ in long-lasting
fatigue of skeletal muscle. European Journal of Applied Physiology and Occupational Physiology 83, 166-174.
Westgaard, R. H. & Winkel, J. (1997). Ergonomic intervention research for improved musculoskeletal health: A critical
review. International Journal of Industrial Ergonomics 20, 463-500.
Zajac, F. E. & Faden, J. S. (1985). Relationship among recruitment order, axonal conduction velocity, and muscle-unit
properties of type-identified motor units in cat plantaris muscle. J.Neurophysiol. 53, 1303-1322.
65
I
Rosendal, L., Blangsted, A. K., Kristiansen, J.,
Søgaard, K., Langberg, H., Sjøgaard, G., &
Kjær, M. (2004). Interstitial muscle lactate,
pyruvate, and potassium dynamics in the
trapezius muscle during repetitive low-force
arm movements, measured with
microdialysis. Acta Physiol Scand In press.
Uncorrected proof.
Printed with permission.
II
Rosendal, L., Søgaard, K., Kjær, M., Sjøgaard,
G., Langberg, H., & Kristiansen, J. (2004).
Increase in interstitial interleukin-6 of
human skeletal muscle with repetitive lowforce exercise. J Appl Physiol In Press,
doi: 10.1152/japplphysiol.00130.2004.
Uncorrected proof.
Printed with permission.
III
Rosendal, L., Larsson, B., Kristiansen, J.,
Peolsson, M., Søgaard, K., Kjaer, M.,
Sörensen, J., & Gerdle, B. (2004). Increase
in muscle nociceptive substances and
anaerobic metabolism in patients with
trapezius myalgia: microdialysis in rest and
during exercise. Pain 112, 345-355
Uncorrected proof.
Printed with permission.