Journal of Gerontology: MEDICAL SCIENCES
1997. Vol. 52A, No. 2, M94-M96
Copyright 1997 by The Gerontological Society of America
The Influence of Age on Isometric
and Isotonic Rat Detrusor Contractions
Ping Kong Lieu, Alfa Sa'adu, E. Olu Orugun, and James G. Malone-Lee
Centre for Geriatric Medicine, University College London Medical School.
Background. Reductions in detrusor shortening velocity and detrusor contractility have been observed in association
with aging. The reasons for these changes are unclear.
Methods. We examined the isometric and isotonic responses of detrusor, taken from the bladder body, to adenosine 5'triphosphate (ATP), noradrenaline (NA), serotonin, and acetylcholine (Ach) in vitro, using bladders from young (30
days) and old (365 days) Sprague-Dawley rats, to determine whether there were any significant age-related differences.
Results. Isometric contractions with ATP at doses of 1O3 to 1CH molar concentrations produced greater contractile
forces in old rats when compared to young rats (p — .0136 at IO~2 mole of ATP). Isotonic contractions at similar
concentrations also produced significant differences between the young and the old rats, the latter being faster (p =
.0225). Isometric contraction with noradrenaline produced significant differences between young and old rats, the latter
being stronger. This became apparent at 10"4 molar concentration of noradrenaline (p = .0043). Isometric contractions
with serotonin also produced significantly greater contractions in the old rats when compared to young rats. The
differences became apparent at 10"* molar concentration of serotonin (p = .045). There were no age-related differences in
isotonic and isometric contractile responses to acetylcholine in the doses used in our experimental setup.
Conclusions. Age-related differences in isometric function were detected in response to ATP, NA, and serotonin.
Differences in isotonic function were only found in response to ATP and Ach.
M
ANY physiological changes in micturition associated
with aging have been described (1,2). Urinary incontinence is common, with an increased incidence in old age (3).
A common urodynamic finding in elderly patients with
lower urinary tract symptoms is detrusor instability (4).
Voiding problems in the elderly have also been described
(5). Detrusor shortening velocity and contractility are reduced, especially in elderly women (1). The reasons why the
elderly bladder behaves in this manner are not entirely clear.
It is possible that the changes reflect alterations in the
physiological properties of the detrusor muscle and its innervation, or its responses to neurotransmitters.
We examined, in vitro, both the isometric and isotonic
responses of the detrusor, taken from the bladder body, to
adenosine 5'-triphosphate (ATP), acetylcholine (Ach), serotonin, and noradrenaline (NA), using bladders from young
and old rats.
ATP depolarizes neuronal and smooth muscle membrane,
and the depolarizing effect may be mediated by directly
opening the nonselective cationic L-type channels which
allow calcium to pass into the cell. ATP does not mobilize
calcium from intracellular pools (6). Acetylcholine, on the
other hand, acts via the receptor gated channels, releasing
calcium from intracellular stores. Acetylcholine has little
effect on the membrane (6,7). Serotonin receptors have been
identified in peripheral neurons and, although the precise
pharmacological action in relation to the bladder is unknown,
it is thought to produce depolarization and modulate transmitter release (7). There are sympathetic (hypogastric) nerves
that supply the bladder body, and these vary from species to
species (7,8). Sympathetic nerves release neuropeptide Y
M94
(NPY) and noradrenaline (NA) (9). Noradrenaline, by acting
via alpha adrenergic receptors, is known to produce inositol
triphosphate as a secondary messenger which releases calcium
from sarcoplasmic reticulum. Inositol triphosphate is produced from phosphoinositol biphosphate, which is resident in
cell membrane and activated by phospholipase C. Phospholipase C is activated by a noradrenaline/receptor complex,
possibly through the G protein (10). The differences in behavior of these agonists may therefore give indirect evidence of
the mechanisms responsible for differences observed between
the age groups.
METHODS
Two groups of female Sprague-Dawley rats were used:
young (30 days) and old (365 days). The usual mean survival
time of these female rats, nonexercised, is 476 days (11).
They were reared in an animal house under sterile conditions. The mean weight for the young rats was 260 grams
and the mean weight for the old rats was 310 grams.
The rats were sacrificed in a carbon dioxide chamber and
their urinary bladders removed. Longitudinal strips of the
detrusor muscle from the posterior wall were cut and suspended in an organ bath containing Tyrodes solution kept at
a constant temperature of 37°C by a flow heater (Grant
Instrument, Cambridge, U.K.). A mixture of 95% O2 and
5% CO2 continuously gassed the mixture in the bath. The
dimensions of each full-thickness bladder strip were 10 mm
long by 2 mm wide. The lower end of each strip was fixed to
a steel tube carrying the gas mixture while the upper end was
suspended by a silk suture to a Grass force transducer
electronically connected to an Apple Macintosh computer
RAT DETRUSOR CONTRACTIONS
which processed the data using the Acqknowledge 2.1 software package. For isometric experiments the strips were tied
directly to the force displacement transducers. In isotonic
experiments, a spring designed to generate a force of 1 gram
per centimeter extension was tied to the upper end of each
muscle strip. The springs were then tied to the force displacement transducers. A resting tension of 1 gram was
applied to all the specimens, and a 30-minute period for
equilibration followed before the start of the experiments.
The velocity of shortening during isotonic contractions
was calculated by differentiating the change of detrusor
length in mm, with respect to time in seconds. Isometric
contractions were quantified by measuring the force generated in millinewtons (mN) (g x 9.81).
The effects of adenosine 5'-triphosphate (ATP), noradrenaline, serotonin, and acetylcholine (all obtained from
Sigma Chemical, St. Louis) on both isotonic and isometric
bladder contractility were studied.
Statistical comparisons were made using the MannWhitney test. The W statistic, 95% confidence interval (CI)
of the difference between means and the /?-value of the
difference were quoted. The 95% level of confidence was
used to reject the null hypothesis.
RESULTS
Isometric contractile responses of bladder body muscle
strips to ATP in old rats were greater than in young rats
(mean young rats .644 mN, mean old rats 1.12 mN; 95% CI
of the difference .1569 to .9418 mN;/? = .0136 at 10"2mole
of ATP (Figure 1). Isotonic contractions of the detrusor
muscle to ATP (at 10"3 to 10"2 molar concentrations of ATP)
were faster in older rats compared to younger rats (mean
young rats .055 mm/sec, mean old rats .183 mm/sec; W =
43; 95% CI of the difference .038 to .213 mm/sec; p =
.0225 at 10-2 molar concentration of ATP).
Isometric contractions with noradrenaline were greater in
the detrusor strips from older rats when compared to younger
rats (mean young rats .005 mN, mean old rats 3.57 mN; W
= 57; 95% CI of the difference 1.60 to 5.8 mN, p = .0043
at 10"4 molar concentrations of noradrenaline (Figure 2).
Isometric contractions of detrusor strips with serotonin
showed significant differences between the young and old
rats (mean young rats 1.04 mN, mean old rats 2.27 mN; W
= 26; 95% CI of the difference .019 to 2.52; p = .0453 at
10"4 molar concentration of serotonin) (Figure 3). However,
within the range of noradrenaline and serotonin concentrations from 10~* to 103 molar used, no isotonic responses were
elicited in both old and young rats in our experimental setup.
Although higher values for isotonic and isometric contractile responses of bladder muscle strips to acetylcholine were
observed in older rats when compared to young rats, these did
not reach statistical significance in the concentrations used
(10* to 10~2 molar concentrations of acetylcholine).
DISCUSSION
In our study, isometric contractile responses were observed with all the autonomic agonists tested. Significant
age-related isometric contractile differences were observed
with adenosine triphosphate (ATP), noradrenaline (NA),
and serotonin despite the initial length of the detrusor strips
being the same. At the initial load of one gram, isotonic contractile responses were only observed with ATP and acetylcholine (Ach). We demonstrated age-related changes in
isotonic contractions only to ATP.
The age-related isometric differences observed with ATP,
NA, and serotonin, when compared to Ach, probably reflect
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Figure 1. Isometric contraction with ATP. Comparison of the in vitro
dose response curves to ATP of detrusor muscle strips of young (n = 6) and
old (n = 6) rats. Bars represent standard deviations.
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Concentration of Noradrenaline (molar)
Figure 2. Isometric contraction with noradrenaline. Comparison of the in
vitro dose response curves to noradrenaline of detrusor muscle strips of
young (n =• 6) and old (n = 6) rats. Bars represent standard deviations.
LIEUETAL.
M96
groups observed with ATP, reflect the observed changes in
isometric contractions between the same age groups. Our
study provides indirect evidence of changes at the detrusor
membrane level between young and old rats. Conclusive
results would require direct physiological studies on the
detrusor membrane. If the findings are substantiated, it may
help in explaining some of the urodynamic changes observed
during voiding in later life. Research into the understanding
of the differences between age groups would need to be
concentrated on the detrusor membrane.
In conclusion, we have demonstrated significant agerelated differences in isometric contractile responses with
respect to ATP, noradrenaline, and serotonin in the detrusor
strips from rat bladder body. Age-related isotonic contractile
responses were demonstrated only to ATP. We suggest that
the differences observed reflect changes in mechanisms of
activation of contraction, and this may be at the level of the
detrusor membrane.
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ACKNOWLEDGMENTS
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Figure 3. Isometric contraction with serotonin. Comparison of the in
vitro dose response curves to serotonin of detrusor muscle strips of young (n
= 6) and old (« = 6) rats. Bars represent standard deviations.
differences in activation of contraction (6). ATP and serotonin effect their responses by depolarizing muscle membrane which results in increases in intracellular calcium (7).
Noradrenaline mediates its action via inositol triphosphate
which is formed by the noradrenaline/alpha-1 adrenoceptor
receptor complex stimulating the hydrolysis of the membrane lipid-phosphatidylihositol (4,5)-diphosphate. This reaction is carried out by a phospholipase C (10). It is therefore
possible to speculate that the age-related changes in isometric responses reflect, indirectly, changes in the detrusor
muscle membranes of young and old rats. This is supported
by the fact that acetylcholine, which has little effect on
muscle membrane but mediates its responses via a receptor
gated channel (6), was able to produce similar forces of
contraction in young as in old rats. This observation is also
indirect evidence that the contractile mechanisms including
intracellular calcium, actin X myosin interactions, and
intracellular calmodulin x calcium interactions are probably the same in the age groups studied.
Isotonic contractile responses were only elicited with
acetylcholine and ATP. This would suggest that ATP and
acetylcholine were the only agonists capable of generating
enough force to overcome the initial calibrated tension load
of 1 gram force in our experimental setup. We chose a 1
gram load because of the inertia experienced with smaller
loads. However, the 1 gram load may have been too high.
The differences in isotonic contraction between the age
This study was supported by the Sandoz Gerontological Research
Foundation.
We are grateful to Mr. Dave Sheppard for his kind assistance with the
breeding of the animals. We thank Dr. Irfan Wahedna for his comments on
the manuscript.
Dr. Lieu is now with the Department of Geriatric Medicine, Tan Tock
Seng Hospital, Singapore 308433.
Address correspondence to Dr. James G. Malone-Lee, Centre for Geriatric Medicine, University College London Medical School, St. Pancras
Hospital, St. Pancras Way, London NW1 OPE. E-mail: [email protected]
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5. Resnick NM, Yalla SV, Laurino E. The pathophysiology of urinary
incontinence among institutionalized elderly persons. N Engl J Med
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6. Illes P, Wolfgang N. Neuronal ATP receptors and their mechanism of
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11. Retzlaff E. Effect of daily exercise on life-span of albino rats. Geriatrics 1966;21:171-7.
Received May 25, 1994
Accepted May 17, 1996