Pulmonary chemoreflexes elicited by intravenous
injection of lactic acid in anesthetized rats
LU-YUAN LEE, ROBERT F. MORTON, AND JAN M. LUNDBERG
Department of Physiology, University of Kentucky Medical Center, Lexington, Kentucky 40536-0084;
and Department of Pharmacology, Karolinska Institut, S-104 01 Stockholm, Sweden
pulmonary C fibers; vagal reflexes; apnea; hyperpnea; capsaicin; perineural capsaicin treatment
NONMYELINATED C-fiber afferent nerve endings can be
stimulated by acid solutions injected into the circulation of various tissues or organ systems, including the
heart, the abdominal viscera, and the limb muscles (24,
27, 28, 30). Lactic acid, a major product of anaerobic
tissue metabolism, seems to be particularly effective in
stimulating these sensory endings (25, 27, 29).
The concentration of lactic acid in body fluids can be
elevated substantially during various physiological and
pathophysiological conditions. For example, lactic acid
can be produced in large quantities by the skeletal
muscles during anaerobic exercise. The production of
lactic acid can be also elevated locally as the result of
tissue ischemia. Lungs are perfused by the total venous
return and are therefore fully exposed to the lactic acid
produced by the peripheral tissues. Furthermore, lungs
and airways are extensively innervated by nonmyelinated C-fiber afferents that play an important role in
regulating various airway functions (3). However, although an involvement of pulmonary C fibers has been
speculated in the tachypneic response to injection of
lactic acid in rabbits (5), the stimulatory effect of lactic
acid on these afferent endings and the consequent
cardiopulmonary reflex responses have not been fully
explored.
The objectives of this study were to 1) characterize
the reflexogenic cardiorespiratory effects of intravenous (iv) injection of lactic acid; we postulated that if
pulmonary C-fiber endings are activated by lactic acid,
bolus injection of this acid should elicit the pulmonary
chemoreflexes, as characterized by apnea, bradycardia,
and hypotension, in spontaneously breathing animals;
2) evaluate the role of vagal C-fiber afferents in eliciting these responses by blocking the conduction of these
afferents with perineural application of capsaicin (12)
to both cervical vagi; and 3) determine the stimulatory
effect of lactic acid on individual vagal pulmonary
C-fiber afferents by using the single-fiber electrophysiological recording technique.
METHODS
Young Sprague-Dawley rats (329 6 6 g, n 5 58) of either
sex were anesthetized with intraperitoneal injection of a-chloralose (100 mg/kg) and urethan (500 mg/kg), and supplemental doses of the same anesthetics were administered intravenously whenever necessary to maintain abolition of the pain
reflex evoked by pinching the skin of the hindlimbs. The
femoral artery and vein were cannulated for recording arterial blood pressure and for iv injections, respectively; the tip
of the venous catheter was positioned slightly below the entry
of the right atrium. Body temperature was maintained at
,36°C throughout the experiment with a temperature servocontroller and a heating pad placed under the animal. A short
tracheal cannula was inserted just below the larynx via a
tracheotomy, through which rats breathed spontaneously in
the supine position. Respiratory flow was measured with a
heated pneumotachograph and a differential pressure transducer (Validyne MP45) and was integrated (Grass 7P10) to
give tidal volume (VT ). Ventilatory signals were recorded on a
Grass polygraph (model 7) and also analyzed by an on-line
computer (CompuAdd model 433); respiratory frequency (f),
VT, and minute volume of ventilation (V̇I) were all analyzed
on a breath-by-breath basis. Results obtained from the computer analysis were routinely compared with those obtained
by hand calculation for accuracy.
Lactic acid (200 mg/ml; Sigma Chemical, St. Louis, MO)
was prepared in distilled water and capsaicin (400 µg/ml;
Sigma Chemical) in a vehicle of 10% Tween 80, 10% ethanol,
and 80% isotonic saline. Solutions of lactic acid and capsaicin
at the desired concentrations were then prepared daily by
dilution with distilled water and isotonic saline, respectively,
on the basis of the animal’s body weight. The volume of each
bolus injection was 0.2 ml; the lactic acid solution injected at
the dose of 0.2 mmol/kg was slightly hypertonic (osmolarity:
335–440 mosmol/l; Wescor osmometer, model 5100B). At least
20 min elapsed between two lactic acid injection challenges.
Before each challenge, the rat’s lungs were hyperinflated
0161-7567/96 $5.00 Copyright r 1996 the American Physiological Society
2349
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Lee, Lu-Yuan, Robert F. Morton, and Jan M. Lundberg. Pulmonary chemoreflexes elicited by intravenous injection of lactic acid in anesthetized rats. J. Appl. Physiol. 81(6):
2349–2357, 1996.—Experiments were carried out to characterize the cardiorespiratory reflex responses to intravenous
injection of lactic acid and to determine the involvement of
vagal bronchopulmonary C-fiber afferents in eliciting these
responses in anesthetized rats. Bolus injection of lactic acid
(0.2 mmol/kg iv) immediately elicited apnea, bradycardia,
and hypotension, which were then followed by a sustained
hyperpnea. The immediate apneic and bradycardiac responses to lactic acid were completely abolished by bilateral
vagotomy and were absent when the same dose of lactic acid
was injected into the left ventricle. The subsequent hyperpneic response was substantially attenuated by denervation
of carotid body chemoreceptors. After a perineural capsaicin
treatment of both vagus nerves to block the conduction of C
fibers, lactic acid no longer evoked the immediate apnea and
bradycardia, whereas the hyperpneic response became more
pronounced and sustained, presumably because of the removal of the inhibitory effect on breathing mediated by
pulmonary C-fiber activation. Single-unit electrophysiological recording showed that intravenous injection of lactic acid
consistently evoked an abrupt and intense burst of discharge
from the vagal C-fiber afferent endings in the lungs. In
conclusion, the cardiorespiratory depressor responses induced by lactic acid are predominantly elicited by activation
of vagal pulmonary C fibers.
2350
PULMONARY CHEMOREFLEXES ELICITED BY LACTIC ACID
Fig. 1. Experimental records illustrating effect of vagotomy on reflex responses to injection of lactic acid in an
anesthetized rat (305 g). A: control; lactic acid (0.2
mmol/kg iv) was injected into a femoral venous catheter
at 1st arrowhead and flushed from catheter into vein as a
bolus at 2nd arrowhead. Tip of catheter was positioned
just below entry of right atrium. B: response to injection
of same dose of lactic acid 30 min after bilateral cervical
vagotomy. VT, tidal volume (upward, inspiration); ABP,
arterial blood pressure.
compared the ventilatory responses to injections of the same
dose of lactic acid before and after sectioning both carotid
sinus nerves in six vagotomized rats. The carotid sinus nerve
was identified on each side of the neck with the aid of a
dissecting microscope; carotid body denervation was verified
by a complete abolition of the hyperpneic response to inhalation of 10 ml of 100% nitrogen.
In seven additional rats, we examined whether the responses to lactic acid injections showed any dose-related
pattern. The responses to three different doses of lactic acid
(0.1, 0.15, and 0.2 mmol/kg iv) were tested in random
sequence in each rat. Hypertonic saline (1.4% NaCl solution)
with the osmolarity (420 mosmol/l) closely matching that of
the highest concentration of lactic acid was injected as
control.
Series 2: effect of perineural capsaicin treatment of vagi on
responses to lactic aid injection. To assess the role of C-fiber
afferents, we used the method of perineural application of
capsaicin to both cervical vagus nerves to block the neural
conduction in these fibers; this method was modified from
that reported first by Jancso and Such (12) and used successfully in our previous study (14). Briefly, cotton strips soaked
in capsaicin solution (0.25 mg/ml) were wrapped around a
2- to 3-mm segment of the isolated cervical vagus nerves for
15–20 min and then removed. Our criterion for a successful
treatment was a complete abolition of the reflex responses to
capsaicin injection (1.0 µg/kg iv); capsaicin was chosen because of its specificity and potency in stimulating the nonmyelinated C-fiber afferent endings. The response to lactic acid
injection was tested twice before and then again within 30
min after completion of the treatment; in a previous study, we
showed that the blocking effect of this treatment lasted for
.60 min (14, 16). To determine whether the perineural
capsaicin treatment affected neural conduction in myelinated
afferent fibers, we also compared the reflex apneic responses
to lung inflation (tracheal pressure 6 cmH2O) during the
control phase and after the treatment in each rat. A total of 10
rats were studied in this series.
Previous investigators have shown that slow continuous iv
infusion of hydrochloric acid causes the endogenous release
from platelets of thromboxane A2, which, in turn, can stimu-
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(tracheal pressure .10 cmH2O) to establish a constant volume history (20).
Three series of experiments were carried out as described
below.
Series 1: ventilatory response to lactic acid injection. This
series was carried out in 10 rats to study the cardiorespiratory reflex responses elicited by iv injection of lactic acid (0.2
mmol/kg). The same dose of lactic acid was injected at least
twice during control to establish the reproducibility of the
response in each animal. The response to lactic acid was
tested again 30 min after bilateral cervical vagotomy to
assess the possible involvement of vagus nerves in eliciting
these responses.
To determine the magnitude of the change in blood pH
caused by lactic acid injection, arterial blood samples were
drawn from four rats both before and within 3–10 s after the
injection; the after-injection sampling time was chosen to
coincide with the apneic response and to correct for the
estimated circulation time between the lungs and the femoral
artery. Because of the minimal sample volume (0.09 ml)
required for the blood-gas analyzer (Instrumentation Laboratory model 1306), it took 5–7 s to collect the blood sample. The
pH of mixed venous blood was also measured in five additional rats that were artificially ventilated after midline
thoracotomies; blood samples were drawn from the right
ventricle both before and within 6 s after the injection of lactic
acid either via a catheter advanced from the right jugular
vein (n 5 1) or by a needle inserted directly into the right
ventricle (n 5 4).
To determine whether the reflex effects observed in this
study were elicited from stimulation of receptors located in
the lungs, the same dose of lactic acid was also injected 30
min later into a catheter that was inserted into the right
carotid artery and advanced retrogradely until its tip was
positioned in the left ventricle in five additional rats; the
position of the catheter was monitored by the pressure trace
and confirmed by postmortem examination.
After the injection of lactic acid, hyperventilation was
consistently elicited after the initial depressor responses and
it persisted even after bilateral vagotomy. To determine the
possible involvement of carotid body chemoreceptors, we
PULMONARY CHEMOREFLEXES ELICITED BY LACTIC ACID
increase, reaching a peak of 189.6 6 37.1 ml/min (P ,
0.01) at the third breath (Fig. 2); the baseline ventilatory parameters were calculated by averaging 10 consecutive breaths immediately before injection in each
rat. Both the immediate apneic and the subsequent
hyperpneic responses to the lactic acid injection were
highly reproducible in the same animals (Fig. 2). Although the apnea and bradycardia occurred only transiently, the hypotensive response usually lasted for
.10 s after the injection (e.g., Fig. 1). Arterial blood pH
decreased substantially from a baseline of 7.44 6 0.02
to 7.16 6 0.04 (n 5 4, P , 0.01; paired t-test) after the
injection of lactic acid, and it returned to the baseline
within 1 min in the two rats tested. In five additional
rats, mixed venous blood pH was measured in the blood
samples drawn directly from the right ventricle, and it
decreased from a baseline of 7.39 6 0.06 to 7.01 6 0.05
after the injection (n 5 5, P , 0.01; paired t-test).
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late pulmonary C fibers (13, 26). To test the possible involvement of this effect in the ventilatory response to lactic acid,
we compared the responses to lactic acid injections during the
control phase and 30 min after administration of indomethacin (15 mg/kg iv), a cyclooxygenase inhibitor, in six additional
rats. Indomethacin (Sigma Chemical) was first dissolved in
polyethylene glycol and then diluted at a 1:1 ratio in saline to
a final concentration of 15 mg/ml for slow infusion. We have
previously shown that one-third of this dose was sufficient to
prevent the bronchoconstrictive effect mediated by the endogenous release of thromboxane A2 in guinea pig lungs (11).
Series 3: effect of lactic acid injection on pulmonary C-fiber
afferents. To further examine whether lactic acid can exert a
stimulatory effect on vagal pulmonary C-fiber endings, we
recorded directly the afferent activity of vagal C fibers in the
lungs of nine rats using a ‘‘single-fiber’’ recording technique,
similar to that described previously (15, 16). Briefly, rats were
paralyzed with pancuronium bromide (Pavulon, 0.05 mg/kg
iv); its effect was allowed to wear off periodically so that the
depth of anesthesia could be monitored. After a midsternal
thoracotomy, the lungs were ventilated with a respirator, and
the expiratory line was placed 3 cm under water to prevent
the lungs from collapsing. The right or left vagus nerve was
isolated from the adjacent carotid artery and sectioned as far
rostrally as possible. The distal end of the cut vagus nerve
was immersed in a pool of mineral oil and placed on a small
dissecting platform. A thin nerve filament was then teased
away from the desheathed nerve trunk, and its afferent
activity was recorded with a monopolar platinum-iridium
hook electrode. The nerve activity was amplified (Grass
P511K), monitored on an audio monitor (Grass AM-8), and
displayed on an oscilloscope (Tektronix 502A). The thin
filament was further split until the afferent nerve activity
from a single unit was electrically isolated. The vagus nerve
was also sectioned or ligated just above the diaphragm to
eliminate the nerve activity arising from the lower visceral
organs.
Pulmonary C fibers usually have a sparse and irregular
discharge but can be activated by hyperinflation of the lungs.
Once the afferent activity of the single unit(s) had been
identified by lung inflation (to 3 or 4 VT ), capsaicin (1.0 µg/kg)
was injected as a bolus via the venous catheter; only fibers
that were activated within 2 s (usually ,1 s) after the
injection were included in this study. The response to bolus
injection of lactic acid (0.2 mmol/kg iv) was tested 15 min
later. Finally, the general location of each receptor was
identified by gentle palpation of the lungs with a small glass
rod after the experiment; those receptors having a location
that could not be identified in the lung structures were not
included.
Statistical analysis. Results were analyzed statistically by
either a one-way or a two-way repeated-measures analysis of
variance, unless otherwise mentioned. When a positive interaction was indicated by the analysis of variance test, the
responses were further compared by using a post hoc analysis
(Fisher’s least significant difference). P , 0.05 was considered
significant. All data are presented as means 6 SE.
2351
RESULTS
Series 1: ventilatory response to lactic acid injection.
Bolus injection of lactic acid (0.2 mmol/kg) immediately
elicited apnea, bradycardia, and hypotension, which
were followed by a more sustained hyperpnea (e.g., Fig.
1). On the first breath after the injection, V̇I decreased
from a baseline of 124.3 6 11.8 to 38.2 6 5.4 ml/min
(n 5 10, P , 0.001) and then quickly started to
Fig. 2. Effect of bilateral vagotomy on ventilatory responses to a
bolus injection of lactic acid (0.2 mmol/kg iv). Vertical dashed lines
depict time of injection. Two control tests separated by .20 min were
performed in each rat to determine reproducibility of response.
Injection was then repeated 30 min after bilateral cervical vagotomy.
Data represent means 6 SE of 10 rats.
2352
PULMONARY CHEMOREFLEXES ELICITED BY LACTIC ACID
Fig. 3. Experimental records illustrating a comparison between responses to lactic acid injected via
intravenous and left ventricular routes in an anesthetized rat (350 g). A: response to a bolus of lactic acid
(0.2 mmol/kg) injected via a femoral venous catheter.
B: same dose of lactic acid injected via a left ventricular catheter. See legend of Fig. 1 for further explanation.
Injection of the dose of 0.2 mmol/kg elicited both the
immediate apneic and the subsequent hyperpneic responses, similar to those found in other series (e.g., Fig.
2); these responses were markedly greater than those
induced by the lower doses of lactic acid in the same
rats. Control injections of hypertonic saline did not
elicit any detectable responses (Fig. 5).
Series 2: effect of perineural capsaicin treatment of
vagi on responses to lactic acid injection. Perineural
capsaicin treatment of both vagi did not cause any
significant change in the baseline VT, f, V̇I, arterial
blood pressure, or heart rate (n 5 10; e.g., Figs. 6 and
7). The reflex apnea induced by lung inflation was not
significantly different during the control phase (12.3 6
2.4 s, n 5 10) and after the treatment (9.7 6 0.7 s, P .
0.05; paired t-test). However, the cardiorespiratory
depressor responses elicited by capsaicin injection were
completely eliminated by the treatment (Figs. 6 and 7).
Similarly, in the same animals, the apnea and bradycardia elicited immediately by the injection of lactic acid
were also abolished after the treatment, whereas the
more sustained hypotensive response was attenuated
but not completely blocked (Figs. 6 and 7). In contrast,
the subsequent hyperpneic response to lactic acid injection was clearly enhanced, primarily because of a
marked increase in f (Figs. 6 and 7).
Pretreatment with indomethacin altered neither the
immediate apneic response nor the subsequent hyperpneic response to lactic acid injection in six additional
rats (Fig. 8). Cardiovascular depressor responses to
lactic acid were also not altered by the indomethacin
pretreatment.
Series 3: effect of lactic acid injection on pulmonary
C-fiber afferents. Vagal pulmonary C fibers discharged
only sparsely and irregularly during control; their
average baseline activity (over a 10-s interval) was
0.14 6 0.08 impulses/s (n 5 12) . All these C-fiber
afferents were activated by hyperinflation of the lungs
(e.g., Fig. 9), and their locations were identified in the
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After bilateral vagotomy, the immediate apnea and
bradycardia were completely abolished, but a small
reduction in VT was frequently found on the first breath
after the acid injection (e.g., Figs. 1 and 2). In contrast,
the hyperpneic response became more pronounced after vagotomy; both the intensity and the duration of the
hyperpnea markedly increased (Figs. 1 and 2).
When the same dose of lactic acid was injected
directly into the left ventricle via the carotid arterial
catheter in five additional rats with intact vagi, the
apnea and bradycardia that were elicited immediately
by the iv injection 30 min earlier were absent in four of
these animals (e.g., Fig. 3) and markedly reduced in the
remaining one. In contrast, the hyperpneic response
still persisted and occurred sooner in all animals,
occasionally accompanied by an augmented breath 2–5
breaths later (e.g., Fig. 3). In addition, left ventricular
injection of lactic acid also induced a systemic hypotension but to a lesser degree than did the iv injection, and
the accompanying bradycardia was absent (e.g., Fig. 3).
Sectioning both carotid sinus nerves completely abolished the hyperpneic response to inhalation of 10 ml
nitrogen in the six vagotomized rats. In the same
animals, denervation of carotid bodies also markedly
diminished the increase in V̇I induced by the injection
of lactic acid (0.2 mmol/kg iv); the peak increases in V̇I
(averaged over five consecutive breaths) were 265.2 6
43.8 and 174.3 6 31.2 ml/min (n 5 6, P , 0.01) before
and after the carotid body denervation, respectively
(Fig. 4).
The cardiorespiratory responses to lactic acid injections showed a dose-related pattern (Fig. 5). Injection of
lactic acid at a lower dose (0.1 mmol/kg iv) did not elicit
significant change in any of the ventilatory parameters,
although a bradypnea was found immediately after the
injection in two of the seven rats tested. A higher dose
(0.15 mmol/kg) evoked a mild but significant reduction
in f (P , 0.05) and V̇I (P , 0.05) on the first breath after
the injection but did not induce any delayed hyperpnea.
PULMONARY CHEMOREFLEXES ELICITED BY LACTIC ACID
2353
Fig. 4. Effect of bilateral carotid body denervation on ventilatory
responses to a bolus injection of lactic acid (0.2 mmol/kg iv) in
vagotomized rats. Vertical dashed lines depict time of injection. BV,
bilateral vagotomy; CBD, carotid body denervation. Data represent
means 6 SE of 6 rats.
lung structures. Each of these afferent fibers was
excited abruptly after a bolus injection of capsaicin (1.0
µg/kg iv). A bolus injection of lactic acid (0.2 mmol/kg iv)
evoked a short burst of discharge (e.g., Figs. 9 and 10)
within 1 s after the injection in 11 of these 12 receptors,
and the activity usually returned to the baseline level
within 5 s (Figs. 9 and 10). The peak fiber activity after
the injection was 6.84 6 1.12 impulses/s (averaged over
a 3-s interval), and the responses were reproducible in
the same fibers.
DISCUSSION
This study clearly describes a biphasic ventilatory
response to iv injection of a bolus of lactic acid in
anesthetized spontaneously breathing rats: an immediate apneic response followed by hyperpnea. The initial
apnea was accompanied by bradycardia and hypotension; these responses resembled the classic ‘‘pulmonary
chemoreflex,’’ which has been repeatedly demonstrated
by previous investigators with injections of various
chemical agents (e.g., capsaicin) or endogenous media-
Fig. 5. Ventilatory responses to iv injections of 3 different doses of
lactic acid (LA) tested in random sequence. An interval of at least 20
min elapsed between 2 tests. Responses to hypertonic saline (osmolarity 5 420 mosmol/l) injections were tested as controls. Data represent
means 6 SE of 7 rats.
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tors (e.g., bradykinin) into the pulmonary circulation
(3, 4). Because the chemical substances that elicit the
pulmonary chemoreflex invariably stimulate the vagal
pulmonary C-fiber afferents, it is believed that activation of these afferents is responsible for triggering the
cardiorespiratory depressor responses (3, 22). The results of this study lend additional support to this
notion, because the initial apnea and bradycardia that
followed the lactic acid injection were completely abolished by perineural capsaicin treatment of both cervical vagi, which has been shown to selectively block the
conduction of C-fiber afferents (12, 14). Indeed, our
study of the individual vagal pulmonary C-fiber afferents has further demonstrated the stimulatory effect of
lactic acid on these sensory endings.
The excitatory effect of lactic acid on nociceptors has
been described in various other organ systems. For
example, it has been reported that lactic acid is a very
effective stimulus of C-fiber afferent endings in the limb
skeletal muscles (24, 28) and in the gastrointestinal
tract (27). However, the action of lactic acid on the
C-fiber afferent endings in the lungs has not been
2354
PULMONARY CHEMOREFLEXES ELICITED BY LACTIC ACID
Fig. 7. Effect of perineural capsaicin treatment of both vagi on ventilatory responses to injections of capsaicin and
lactic acid. Left, responses to a bolus injection of capsaicin (1 µg/kg iv); right, responses to lactic acid (0.2 mmol/kg
iv). Vertical dashed lines depict time of injections. Data represent means 6 SE of 10 rats.
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Fig. 6. Experimental records illustrating effect of perineural capsaicin treatment of vagi on responses to injections
of capsaicin and lactic acid in an anesthetized rat (330 g). Left (A and B), control; right (C and D), after perineural
capsaicin treatment of both vagi. Top (A and C), responses to bolus injections of capsaicin (1 µg/kg iv); bottom (B and
D), lactic acid (0.2 mmol/kg iv). An interval of at least 20 min elapsed between 2 tests. See legend of Fig. 1 for further
explanation.
PULMONARY CHEMOREFLEXES ELICITED BY LACTIC ACID
previously characterized. Continuous perfusion of isolated guinea pig lungs with acidic buffer at a pH of 5.0
has been shown to evoke the release of various sensory
neuropeptides, such as tachykinins and calcitonin generelated peptides, presumably from C-fiber afferents
(18). Sustained application of acidic buffer of the same
low pH has also been demonstrated to stimulate C
fibers in guinea pig trachea in vitro (6). In the present
study, the change in blood pH caused by lactic acid
injection was considerably smaller and lasted for only a
very short duration, but the injection nevertheless
caused a distinct excitatory effect on C-fiber endings in
the lungs. Although C fibers in the abdominal viscera
and skeletal muscles seem to be particularly sensitive
to hydrogen ions carried by the lactic acid, as compared
with the responses to other acidic solutions (25, 27, 29),
whether lactic acid is more potent than other forms of
acid in stimulating pulmonary C fibers remains to be
determined.
After injection into the venous blood, lactic acid can
react with bicarbonate ions and produce CO2, which is
then eliminated from the lungs (19). This reaction has
been shown in our preliminary trials in anesthetized
rats; the end-tidal CO2 concentration increased immediately after injection of lactic acid (0.2 mmol/kg iv) from
,5 to 7–8% and lasted for three to four breaths (L.-Y.
Lee and R. F. Morton, unpublished observations). This
change may explain the marked difference in pH between mixed venous blood and systemic arterial blood
found in this study. We believe that these changes in
blood pH caused by the lactic acid injection led to a
transient reduction of pH in the pulmonary interstitial
fluid, which, in turn, activated these C-fiber sensory
endings in the lungs.
The transduction mechanism of the acid-induced
excitation of C-fiber endings is not fully understood. It
has been suggested that low-pH buffer may trigger the
release of an endogenous ligand for the ‘‘capsaicin
receptor’’ and thereby activate the C-fiber afferents and
evoke the release of tachykinins from these endings,
because these effects of acid could be completely blocked
in the isolated guinea pig trachea and lungs by capsazepine (6, 18), a specific antagonist to the capsaicin
receptor. The possible involvement of cyclooxygenase
metabolites in the acid-induced activation of C fibers
has also been reported in other visceral organs (7, 17).
Indeed, it has been shown that a slow, continuous
infusion of hydrochloric acid triggers platelet to release
thromboxane A2, which, in turn, can stimulate pulmonary C fibers (13, 26). However, we can rule out the
possibility that the stimulatory effect on pulmonary C
fibers by the bolus injection of lactic acid in the present
study is mediated by the action of endogenous cyclooxygenase metabolites because the pulmonary chemoreflex elicited by lactic acid was not affected by pretreatment with indomethacin (Fig. 8). Alternatively, it has
been shown that hydrogen ions can directly excite
capsaicin-sensitive C-fiber neurons of the rat dorsal
root ganglia by evoking a sustained inward current as
the result of an increase in monovalent cation (Na1, K1,
Cs1 ) conductance in the neuronal membrane (2, 23).
Whether this proton-activated current is responsible
for generating action potentials in the pulmonary Cfiber endings by lactic acid remains to be investigated.
After either vagotomy or perineural capsaicin treatment of both vagi, the same dose of lactic acid no longer
triggered the initial apneic response. However, neither
vagotomy nor perineural treatment prevented the subsequent hyperpnea. On the contrary, the hyperpneic
response to lactic acid was markedly enhanced and
prolonged, presumably because of the elimination of
the inhibitory effect caused by the pulmonary C-fiber
activation. Bolus injection of lactic acid (0.2 mmol/kg iv)
lowered pH in the arterial blood abruptly and transiently from 7.44 to 7.16 in this study; this acidotic load
presumably stimulated peripheral chemoreceptors and
thereby elicited a reflex hyperpneic response (1). This
hypothesis is supported by our observation that sectioning both carotid sinus nerves eliminated ,75% of the
hyperpnea induced by lactic acid injection in vagotomized rats. The remaining hyperpneic response is
probably mediated through the excitation of other
sensory receptors (e.g., aortic body chemoreceptors).
Furthermore, injection of lactic acid is known to lead to
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Fig. 8. Effect of indomethacin on ventilatory responses to injection of
lactic acid (0.2 mmol/kg iv). r, Responses 30 min after a pretreatment
with indomethacin (15 mg/kg iv). Vertical dashed lines depict time of
injections. Data represent means 6 SE of 6 rats.
2355
2356
PULMONARY CHEMOREFLEXES ELICITED BY LACTIC ACID
a transient increase of arterial CO2 tension (19), which
may stimulate the central chemoreceptors and induce
hyperventilation.
In vagotomized rats, injection of lactic acid still
triggered a small reduction in VT of the first breath
immediately after the injection (e.g., Figs. 1, 2, and 6).
This response is presumably elicited by activation of
nonvagal afferents, and the receptor type(s) and location(s) are not identified in this study. Because of the
rapid onset of the response, the receptors are probably
located in or near the pulmonary circulation. We suspect that sympathetic afferents arising from the lungs
and/or heart may be involved (3, 30).
Although stimulation of pulmonary C-fiber afferents
is known to induce the transient systemic hypotension
that accompanies the reflex bradycardia (3, 4, 16), the
more sustained hypotensive response to a bolus injection of lactic acid observed in this study probably
involved, to some extent, a direct vasodilatory effect of
hydrogen ions on the peripheral vascular smooth
muscles (9). This speculation is supported by our
observations that the hypotensive response was also
induced by the left ventricular injection of lactic acid
Fig. 10. Responses of vagal C-fiber afferents in lungs to injections of capsaicin and lactic acid. A: responses to a
bolus injection of capsaicin (1.0 µg/kg
iv). B: responses to lactic acid (0.2
mmol/kg iv). Data represent means 6
SE of 12 fibers.
(e.g., Fig. 3) and could not be completely abolished by
either bilateral vagotomy or perineural capsaicin treatment of both vagi (e.g., Figs. 1 and 6).
It is well recognized that pulmonary C-fiber afferent
endings are very sensitive to various inhaled irritants
and to certain blood-borne autocoids and that they,
therefore, play an important role in regulating the
respiratory defense functions in both physiological and
pathological conditions (3). Activation of these afferent
endings by chemical substances such as lactic acid is
known to elicit the pulmonary chemoreflex, as described above, accompanied by reflex bronchoconstriction and hypersecretion of mucus (3). In addition,
activation of these afferents is believed to be involved in
evoking the dyspneic sensation in certain pathophysiological conditions (e.g., pulmonary edema or lung
inflammation) (8, 21). It has been postulated that the
dyspneic sensation during strenuous exercise could
result from an excitation of pulmonary C-fiber afferents
caused by pulmonary congestion or a mild increase in
interstitial pressure in the lungs under those conditions (21, 22). However, on the basis of our results in
this study, it seems conceivable that lactic acid may
Downloaded from http://jap.physiology.org/ by 10.220.33.6 on June 18, 2017
Fig. 9. Experimental records illustrating stimulatory effect of lactic acid on a pulmonary C fiber arising from ending
in right lower lobe of an anesthetized paralyzed open-chest rat (335 g). A: response to lung inflation. B: response to a
bolus injection of capsaicin (1.0 µg/kg iv). C: response to a bolus injection of lactic acid (0.2 mmol/kg iv).
Hyperinflation of lungs was produced by occluding expiratory line of respirator for 3 consecutive cycles, and
inflation was prolonged by turning off the respirator. Fifteen minutes elapsed between B and C. AP, action
potentials; Pt, tracheal pressure. See legend of Fig. 1 for further explanation.
PULMONARY CHEMOREFLEXES ELICITED BY LACTIC ACID
also play a part in the genesis of the dyspneic sensation
during severe exercise. Lactic acid is an important
product of anaerobic metabolism, and the production of
lactic acid can increase drastically during heavy exercise; blood lactate concentrations in the range of 10–20
mmol/l have been reported during heavy exercise in
healthy human volunteers (10). Several other pathological conditions that could lead to excessive production of
lactic acid in the tissue include local tissue ischemia
and inflammation. Whether pulmonary C-fiber endings
are activated by lactic acid endogenously produced
under those conditions remains to be determined.
Received 18 April 1996; accepted in final form 22 July 1996.
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The authors thank Dr. Mary K. Rayens for statistical analysis of
the data and Margareta Stensdotter, Kevin Kwong, and Ju-Lun Hong
for technical assistance.
This study was supported by Grants HL-40369 and HL-52172
from the National Heart, Lung, and Blood Institute and by Grant
14X-6554 from the Swedish Medical Research Council. L.-Y. Lee was
a recipient of the Senior International Fellowship (TW 01734)
awarded by the National Institutes of Health Fogarty Center.
Address for reprint requests: L.-Y. Lee, Dept. of Physiology, Univ.
of Kentucky, Lexington, KY 40536-0084.
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