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Central integration of the autonomic cardiorespiratory response to nasopharyngeal stimulation in the rabbit
Brain Research, 87 (1975) 171-179
171
© ElsevierScientificPublishing Company, Amsterdam - Printed in The Netherlands
CENTRAL INTEGRATION OF THE AUTONOMIC CARDIORESPIRATORY RESPONSE TO NASOPHARYNGEAL STIMULATION IN THE RABBIT
SAXON W. WHITE School of Medicine, The Flinders University of South Australia, Bedford Park, S.A. 5042 (Australia)
Environmental disturbances provoke in the intact animal a variety of autonomic responses each of which may be shown to be a function of the input profile and its central integration. In the rabbit, the nasal inhalation of small amounts of irritants such as smoke or ammonia vapour is followed by apnoea in expiration, a rise in arterial pressure, vagal bradycardia and widespread sympathetic adrenergic vasoconstriction resulting in a marked restriction of blood flow in over 90 ~ of the soft tissue mass4,9,16. The trigeminal nerve initiates the apnoea, and the arterial baroreceptors and sudden loss of lung inflation both contribute to the cardiac slowing 1°,15. The olfactory nerves do not play a significant role in the cardiovascular disturbance, and the effects are not a function of chemicals absorbed from the respiratory tract or of stimulation of optic nerves or receptors of the face and lower respiratory tract 1°,t4,15. The current studies were undertaken in unanaesthetized rabbits to examine the role of different brain regions in the strong autonomic activation evoked by nasopharyngeal stimulation. In addition, the separate components of the input profile were studied in relation to the heart rate and mesenteric vasoconstrictor responses, which were used as indices of autonomic excitation mechanisms. The effects of nasal inhalation of cigarette smoke were first examined in spontaneously breathing pontine (decerebrate), thalamic and sham-operated rabbits in which the carotid sinus and aortic nerves were intact, and in similar preparations when they were sectioned. Then in the same animals under conditions of controlled ventilation the separate effects of smoke stimulation without apnoea, and of apnoea without smoke, were studied. The results suggest that both the trigeminal and lung inflation components of the input profile cause activation of heart rate and vasoconstrictor neurones through mechanisms that operate at both bulbospinal and suprabulbar levels, and that convergence of these inputs with the arterial baroreceptor input serves to facilitate the autonomic responses. The experiments were carried out using New Zealand white rabbits weighing between 2.4 and 3.5 kg. Blood flow in the superior mesenteric artery was measured using a Doppler ultrasonic transducer, which was implanted at a preliminary laparotomy under halothane anaesthesia 10--14 days before an experiment. When connected to the Doppler flowmeter (Karl Pierson, Pierson Lab., 23032 Via Cereza Street, Mission Viejo, Calif. 92675) transducers chronically implanted in this way provide an
172 output (blood velocity) which is a linear function of blood flow rate 14,16. On the day of the experiment under halothane anaesthesia, the ear artery was cannulated for systemic arterial pressure measurement (Statham P23 Dc strain-gauge). A frontoparietal craniotomy was then performed and sham-operated rabbits, thalamic rabbits (in which cerebral hemispheres and basal ganglia were removed) and pontine rabbits (infracollicular decerebration) were prepared as described previously 7. There were 4 animals in each of these groups, and in 4 additional animals of each group the carotid sinus and aortic nerves (buffer nerves) were cut bilaterally in the neck immediately
Fig. 1. Method of deliveringcigarette smoke to nasal region of unanaesthetized rabbit.
173 •
$
II
$
II
•
AV mm
Hg
~
MF
~
~'F
56~
¢m/se¢
28 0
HR /rain
'L
o
4eo~ 240
_ _
- -
-
-
O I I'~
I 2 mlns
Fig. 2. Rabbit 2.5 kg, unanaesthetized. A continuous record is shown to illustrate both the stability of the circulation and the reproducibility of the response to successive cigarette smoke stimuli (at
arrows) delivered as in Fig. 1. (From ref. 15, with permission.) after the neurosurgical procedure, exactly as described previously 5. Sham-operated and thalamic animals recovered normal movement and posture within 0.5-1 h from the end of anaesthesia. Pontine rabbits lay on their side until 30 min before the start of the experiment; their body temperature was maintained by an electric blanket. Details of the postoperative management and posturing of these animals for experiments were as previously described 7. Nasopharyngeal stimulation was produced by blowing cigarette smoke gently through a narrow PVC tube to a position immediately in front of the animal's nose (Fig. 1), from where it was actively inhaled. The cardiorespiratory response in a given animal was highly reproducible (Fig. 2), and the response of a given animal was calculated as the mean of 3 stimuli given at 3-5 rain intervals. For each test, measurements of arterial pressure, heart rate, mesenteric blood flow and mesenteric 'conductance' (mesenteric flow/arterial pressure) were made just before smoke stimulation, and 7-12 sec later when the response was at its height. The duration of apnoea was also measured using a light AC-coupled carbonin-plastic strain-gauge strapped about the rabbit's thorax. The response to smoke stimulation was first observed during spontaneous ventilation and the animal was then given a short acting anaesthetic (propanidid, Epontol, Bayer) and intubated with a No. 14 Foregger paediatric endotracheal tube lubricated with Xylocaine jelly. It was then connected to a respiratory pump and ventilated at the average minute ventilation (1 1/min) and rate (60/min) of normal rabbits. While still under anaesthesia, muscular relaxation was induced by decamethonium iodide (1 mg/kg i.v., followed by 0.5 mg/kg i.v. every 20 min). The animals recovered from anaesthesia while ventilation was controlled, and periodic blood gas determinations were made and compared with the values for that animal when spontaneously breathing. Any differences were minimized by appropriate adjustments to the respiratory pump. No operative interventions were carried out while ventilation was controlled and decamethonium continued. Fifteen minutes after beginning controlled ventilation the circulatory effects of apnoea, in the absence of smoke stimulation, were studied by disconnecting the respiratory pump for 7-10 sec to simulate the apnoea of the normal response. Five minutes later the circulatory effects of blowing a small amount of smoke into the nose were studied while ventilation was maintained at 1 1/min. The alternate stimuli of apnoea alone and smoke alone were repeated twice more at 5 rain intervals.
174 The role of central mechanisms was assessed by comparing the evoked mean
changes in a given variable in the different preparations and the responses were expressed as per cent change using each animal as its own control. This method was used because in the present study the differences in initial resting values between preparations were generally ~mall and the conclusions reached by using either absolute or percentage changes were similar 17. Resting status of the different neural preparations. Because of the unusual nature of the preparations used in the present study, it is pertinent to review the status of the resting respiratory and circulatory variables before dealing with differences between responses. In rabbits with intact carotid sinus and aortic nerves resting respiration, arterial pressure, heart rate and mesenteric blood velocity differed little between shamoperated, thalamic and pontine animals (Table I). In rabbits with section of the buffer nerves, arterial pressure was higher than in corresponding groups in which the nerves were intact, whilst mesenteric blood velocity was lower. For this reason most of the quantitative comparisons of 'conductance' changes were made within each of the two subgroups of neural preparations with different buffer nerve status. Controlled ventilation had minimal effects on resting circulatory variables in any of the different preparations 17. Suprabulbar and bulbospinal regulation of respiration. In rabbits with intact buffer nerves, apnoea time was similar in sham-operated and pontine rabbits, but was shorter in thalamic animals (P = 0.05, Table II). In rabbits with sectioned buffer nerves, the duration of apnoea was longer than in the corresponding groups in which the nerves were intact, however, as in animals with intact buffer nerves the apnoea time of sham-operated and pontine animals was similar, but in thalamic animals it was shorter (P = 0.02). The longer apnoea of animals with an intact brain but with sectioned buffer nerves has been noted previously 10, and has been attributed to the absence of arterial chemoreceptor drive secondary to apnoea as asphyxial blood gas changes developed. The shorter duration of apnoea in thalamic animals compared TABLE I RESTING CARDIOVASCULARMEASUREMENTSIN SPONTANEOUSLYBREATHING NEURAL PREPARATIONS WITH AND WITHOUT INTACT CAROTID SINUS AND AORTIC NERVES Values a m averages a n d s t a n d a r d error o f m e a n for 4 animals. H R , heart rate, b e a t s / m i n ; AP, arterial pressure, m m H g ; MBV, mesenteric blood velocity, cm/sec.
Sham Thalamic Pontine
lntact carotid sinus and aortic nerves
Section carotid sinus and aortic nerves
HR
AP
MBV
HR
AP
MBV
264 (4.4) 266
94 (2.2) 93
23 (3.0) 21
292 (2.0) 245
104 (8.4) 106
16 (0.9) 14
(18.2)
(5.8)
(2.5)
(5.0)
(7.5)
(2.5)
220 (31.5)
82 (2.7)
22 (0.9)
293 (7.5)
94 (4.2)
13 (1.3)
175 TABLE II DURATION OF APNOEA EOR DIFFERENT NEURAL PREPARATIONS (GROUPS OF 4 ANIMALS) Values are averages 4- S.E.M.
Sham Thalamic Pontine
Intact carotid sinus and aortic nerves
Section carotid sinus and aortic nerves
7.8 ± 0.63 2.8 d: 1.60 9.3 d: 2.59
17.3 ± 1.44 8.5 ± 2.86 15.0 ± 1.50
with the other two neural preparations suggests that normally cerebral structures facilitate the apnoea initiated by trigeminal stimulation. The apnoea time of pontine animals on the other hand was the same as in sham-operated animals, suggesting in turn that diencephalic structures exert inhibitory effects on the reflex suppression of respiration evoked at bulbospinal levels, an effect normally masked by cerebral facilitation. It should be noted that the characteristic differences between the 3 neural preparations are present irrespective of the buffer nerve status, suggesting that chemoreceptor effects on respiration are predominantly exerted at bulbospinal levels. This finding is in agreement with conclusions drawn from other experiments in which respiratory drive was increased rather than inhibited7. Suprabulbar and bulbospinal regulation of heart rate. Trigeminal afferents could evoke their circulatory effects directly through projections to the pontomedullary reticular formation1, and/or indirectly due to changes associated with the sudden inhibition of respiration in expiration, i.e., the sudden reduction in the input via vagal afferents from lung inflation receptors2,3,1% In addition, with the onset of apnoea and bradycardia, there is a marked increase in atrial transmural pressure 13, and previous studies have shown that arterial baroreceptors contribute to the reduction in heart rate due, most probably, to the rise in arterial pressure. Changes in chemoreceptor activity contribute to the termination of the apnoea but appear to play little part in the bradycardia of the single bolus smoke stimulus10. In the current study, bradycardia was evoked by apnoea without smoke stimulation, and also by smoke stimulation without apnoea. The effects of apnoea alone on heart rate appeared to depend largely on the integrity of the buffer nerves, since after their section there was only a small residual bradycardia (P < 0.05, Fig. 3). This in turn was dependent on intact cerebral hemispheres (P < 0.05). It may be suggested, therefore, that the reflex heart rate effects of sudden loss of lung inflation are partly dependent on arterial barorecepter interactions with lung inflation afferents, and partly on arterial baroreceptor independent mechanisms. After buffer nerve section, the afferent drive induced by apnoea alone arises from lung inflation receptors, cardiac receptors, or both, and is transmitted through vagal afferents2,3,11,1L Other studies have indicated that the arterial baroreceptor-heart rate reflex is dependent on supra~pontine regions 6,8, and the present study suggests that the reflex response to
176 ISECTION C.S.+A.N.J
IINTACT C.S.+AN.I MEAN ARTERIAL PRESSURE %
t
Sh
Th
Po
•
II
m
I00
HEART RATE %
%
801 60 40
oO; oE
I '°-°
20
1
Fig. 3. Haemodynamic effects of simulated apnoea in expiration in the absence of cigarette smoke stimulation in groups of 4 sham-operated rabbits (Sh), thalamic rabbits (Th) and pontine rabbits (Po). Left 3 panels, preparations with intact carotid sinus and aortic nerves; fight 3 panels, preparations in which the carotid sinus and aortic nerves have been cut. The symbol on the left = 2 S.E. of mean difference between control and response values for any one of the 3 responses, calculated by analysis of variance. The mean absolute resting data for these rabbits and those in Figs. 4 and 5 are shown in Table I. a p n o e a alone made by vagal afferents on the one hand, and the carotid sinus and aortic nerves on the other are both dependent on suprapontine regions. The bradycardia induced by nasal smoke stimulation without apnoea in rabbits with section of the buffer nerves probably occurs t h r o u g h trigeminal afferents (Fig. 4). In these animals the reflex effects in part depend on intact cerebral hemispheres, since cardiac slowing was greatest in sham-operated animals (P < 0.05). W h e n the buffer nerves were intact, smoke stimulation without apnoea caused greater bradycardia (P < 0.05) which was similar in magnitude in all 3 neurological preparations. The trigeminal input thus appears to have direct effects on heart rate neurones which are dependent on mechanisms involving the cerebral hemispheres. In addition, there appears to be an interaction effect between trigeminal and arterial baroreceptor afferents at bulbospinal levels. iINTACT C.S.+A.N] Sh
ARTERIAL
PRESSURE HEART RATE %
'40
Th
!1
Po
[SECT,ON CS÷AN] Sh
Th
EQ
I l l
80I 6O 40
FLOW
60 F
CONDUCTANCE I % 2O Fig. 4. Effects of nasal stimulation with smoke during controlled ventilation. Notations as in Fig. 3.
177 I INTACT C.S.*A.N. I Sh MEAN ARTERIAL PRESSURE %
140 I00 L
RCTE
f
~
Th
milll
[ SECTION C.S.+A.N, [
Po
Sh ~ ,[
Th
Po
i..m
60f
FLOW CONDUCTANCE % 20
Fig. 5. Effectsof smoke stimulation in spontaneouslybreathing animals. The respiratory (apnoea time) data from these experiments is shown in Table II. Notations as in Fig. 3.
In view of the above findings, in the spontaneously breathing animal with section of the buffer nerves one would expect bradycardia of the smoke reflex to be present but less than normal in all neural preparations. Moreover, in view of the importance of the cerebral hemispheres for reflexes from both the trigeminal and lung inflation inputs, the cardiac slowing might be expected to be greater in the animals with intact brains. These expectations were confirmed experimentally and are shown in Fig. 5. In the intact animal, the bradycardia due to smoke inhalation thus appears to be a complex response depending on at least 3 peripheral inputs: (1) the direct effects of trigeminal stimulation, (2) the effects of apnoea predominantly through various vagal afferents and (3) the arterial baroreceptors. Suprabulbar and bulbospinal regulation of the mesenteric circulation. Central nervous regulation of the mesenteric circulation through adrenergic vasoconstrictor fibres is similar in certain respects to the regulation of heart rate. Thus in animals in which the buffer nerves are cut, the effects of apnoea without smoke stimulation and smoke stimulation without apnoea differ in magnitude between sham-operated, thalamic and pontine animals (Figs. 3 and 4). Mesenteric vasoconstriction evoked by apnoea alone is dependent on cerebral mechanisms, since only in sham-operated animals is there any fall in 'conductance' (P < 0.05, Fig. 3). Likewise, smoke stimulation alone, although reducing 'conductance' in all 3 neural preparations, does so to the greatest extent in sham-operated animals (Fig. 4). These data, as with the regulation of heart rate, suggest a role for the cerebral hemispheres in the regulation of sympathetic vasoconstrictor fibre activity through both the trigeminal and lung inflation inputs which are independent of the arterial baroreceptor input. Clearly, there is also a direct control through the trigeminal input, which appears to act at a bulbospinal level, but there is no equivalent control through the lung inflation input (el Figs. 3 and 4, right panels, pontine animals). When the buffer nerves are intact the reduction in conductance through either input is considerably enhanced. This possibly reflects a greater degree of reflex vasoconstriction; however, this interpretation is uncertain in view of the differences in resting conductance between animals with and
178 without section of the buffer nerves. Moreover, the rise in arterial pressure might be expected to stimulate arterial baroreceptor reflexes to minimize the mesenteric vasoconstriction rather than accentuate it. It is possible therefore that the arterial chemoreceptor input is responsible for the enhanced vasoconstriction, even though its role in the concomitant reflex bradycardia is negligible 10. An alternative explanation is that projections from trigeminal and vagal afferents converge on vasoconstrictor neurones also receiving arterial baroreceptor afferents, producing 'resetting' of barereflex threshold and gain s . In this way, constrictor effects would be enhanced despite the rise in arterial pressure. Of particular interest in thalamic animals with intact buffer nerves is the vasoconstriction evoked by apnoea alone, which is not present in these preparations when the buffer nerves are cut, or in pontine animals (Fig. 3). These data suggest that suprapontine interactions between lung inflation and arterial baroreceptor afferents play an important role in the vasoconstriction evoked by apnoea alone. Conclusions. It is concluded that when nasopharyngeal stimulation occurs in the intact rabbit there is suprabulbar as well as bulbospinal regulation of respiration and the circulation. The return of respiration after reflex suppression of breathing by trigeminal stimulation is partly a function of arterial chemoreceptor activity acting on bulbospinal neurones. Trigeminal, lung inflation and arterial baroreceptor afferents all appear to play a role in exciting heart rate and sympathetic vasoconstrictor pathways, and arterial baroreceptor-lung inflation and arterial baroreceptor-trigeminal interactions appear to mutually enhance autonomic activity at different levels of the central nervous system. An alternative explanation is that brain sections interrupt facilitatory pathways descending to autonomic neurones in bulbospinal regions. In both cases the results demonstrate the role of suprapontine mechanisms in the autonomic response to a complex input profile elicited by a simple environmental stimulus. These studies were carried out in collaboration with Dr. R. J. McRitchie and Prof. P. I. Korner. Mr. D. Lauff gave valuable technical assistance. The study was supported by research grants from the Life Insurance Medical Research Fund of Australia and New Zealand, the National Heart Foundation of Australia, the Australian Tobacco Research Foundation, the National Health and Medical Research Council of Australia and the Postgraduate Medical Foundation of the University of Sydney.
1 BRODAL,A., Neurological Anatomy, Oxford Univ. Press, London, 1969, 415 pp. 2 DALY, M. DE B., AND HAZZLEDINE, J. L., The effects of artificially induced hyperventilation on the
primary cardiac reflex response to stimulation of the carotid bodies in the dog, J. PhysioL (Lend.), 168 (1963) 872-889. 3 DALY, M. DE B., AND ROBINSON, B. H., An analysis of the reflex systemic vasodilator repsonse elicited by lung inflation in the dog, J. Physiol. (Lend.), 195 (1967) 387~106. 4 FORSTER,R. P., ANDNYBOER,J., Effect of induced apnoea on cardiovascular renal functions in the rabbit, Amer. J. Physiol., 183 (1955) 149-154. 5 KORNER, P. I., Effect of section of the carotid sinus and aortic nerves on cardiac output of the rabbit, J. Physiol. (Lend.), 180 (1965) 266-278.
179 6 KORNER,P. I., SHAW, J., WEST, M. J., AND OLIVER, J. R., Central nervous system control of baroreceptor reflexes in the rabbit, Circulat. Res., 31 (1972) 637-652. 7 KORNER,P. I., UTHER, J. B., AND WHITE, S. W., Central nervous integration of the circulatory and respiratory response to arterial hypoxemia in the rabbit, Circulat. Res., 24 (1969) 757-776. 8 KORNER, P. I., WEST, M. J., AND SHAW, J., Central nervous resetting of baroreceptor reflexes, Aust. J. exp. Biol. Med. ScL, 51 (1973) 53-64. 9 KRETSCHMER,F., Ll~oerReflexe vonder Nasenschleimhaut auf Atmung und Kreislauf, Sber. Akad. Wiss. (Wien), 62 (1870) 147. 10 McRITCHIE, R. J., ANO WHITE, S. W., Role of trigeminal, olfactory, carotid sinus and aortic nerves in the respiratory and circulatory response to nasal inhalation of cigarette smoke and other irritants in the rabbit, Aust. J. exp. BioL Med~ Sci., 52 (1974) 127-140. 11 OBERG,B., AND WHITE, S. W., Circulatory effects of interruption and stimulation of cardiac vagal afferents, Acta physiol, scand., 80 (1970) 383-394. 12 OaT, N. T., AND SrmPHERD, J. T., Vasodepressor reflex from lung inflation in the rabbit, Amer. J. Physiol., 221 (1971) 889-895. 13 PAINTAL,A. S., A study of right and left atrial receptors, J. Physiol. (Lond.), 120 (1953) 596-610. 14 WHITE,S. W., ANGUS, J. A., McRITCHIE, R. J., AND PORGES, W. L., Evaluation of the Doppler Flowmeter for measurement of blood flow in small vessels of unanaesthetized animals. In J. R. S. HALES AND S. W. WHITE (Eds.), Proc. Syrup. Clin. exp. Blood Flow Measurement, Clin. exp. Pharm. Physiol., Suppl. 1 (1974) 79-92. 15 WHITE,S. W., AND McRrrCHIE, R. J., Nasopharyngeal reflexes: integrative analysis of evoked respiratory and cardiovascular effects, Aust. J. exp. BioL Med. Sci., 51 (1973) 17-31. 16 WHITE, S. W., McRITCHIE, R. J., AND FRANKLIN,D. L., Autonomic cardiovascular effects of nasal inhalation of cigarette smoke in the rabbit, Aust. J. exp. Biol. Med. Sci., 52 (1974) 111-126. 17 WHITE, S. W., MCRITCHIE, R. J., AND KORNER, P. I., CNS control of cardio-respiratory nasopharyngeal reflexes in the rabbit, Amer. J. PhysioL, In press.
171
© ElsevierScientificPublishing Company, Amsterdam - Printed in The Netherlands
CENTRAL INTEGRATION OF THE AUTONOMIC CARDIORESPIRATORY RESPONSE TO NASOPHARYNGEAL STIMULATION IN THE RABBIT
SAXON W. WHITE School of Medicine, The Flinders University of South Australia, Bedford Park, S.A. 5042 (Australia)
Environmental disturbances provoke in the intact animal a variety of autonomic responses each of which may be shown to be a function of the input profile and its central integration. In the rabbit, the nasal inhalation of small amounts of irritants such as smoke or ammonia vapour is followed by apnoea in expiration, a rise in arterial pressure, vagal bradycardia and widespread sympathetic adrenergic vasoconstriction resulting in a marked restriction of blood flow in over 90 ~ of the soft tissue mass4,9,16. The trigeminal nerve initiates the apnoea, and the arterial baroreceptors and sudden loss of lung inflation both contribute to the cardiac slowing 1°,15. The olfactory nerves do not play a significant role in the cardiovascular disturbance, and the effects are not a function of chemicals absorbed from the respiratory tract or of stimulation of optic nerves or receptors of the face and lower respiratory tract 1°,t4,15. The current studies were undertaken in unanaesthetized rabbits to examine the role of different brain regions in the strong autonomic activation evoked by nasopharyngeal stimulation. In addition, the separate components of the input profile were studied in relation to the heart rate and mesenteric vasoconstrictor responses, which were used as indices of autonomic excitation mechanisms. The effects of nasal inhalation of cigarette smoke were first examined in spontaneously breathing pontine (decerebrate), thalamic and sham-operated rabbits in which the carotid sinus and aortic nerves were intact, and in similar preparations when they were sectioned. Then in the same animals under conditions of controlled ventilation the separate effects of smoke stimulation without apnoea, and of apnoea without smoke, were studied. The results suggest that both the trigeminal and lung inflation components of the input profile cause activation of heart rate and vasoconstrictor neurones through mechanisms that operate at both bulbospinal and suprabulbar levels, and that convergence of these inputs with the arterial baroreceptor input serves to facilitate the autonomic responses. The experiments were carried out using New Zealand white rabbits weighing between 2.4 and 3.5 kg. Blood flow in the superior mesenteric artery was measured using a Doppler ultrasonic transducer, which was implanted at a preliminary laparotomy under halothane anaesthesia 10--14 days before an experiment. When connected to the Doppler flowmeter (Karl Pierson, Pierson Lab., 23032 Via Cereza Street, Mission Viejo, Calif. 92675) transducers chronically implanted in this way provide an
172 output (blood velocity) which is a linear function of blood flow rate 14,16. On the day of the experiment under halothane anaesthesia, the ear artery was cannulated for systemic arterial pressure measurement (Statham P23 Dc strain-gauge). A frontoparietal craniotomy was then performed and sham-operated rabbits, thalamic rabbits (in which cerebral hemispheres and basal ganglia were removed) and pontine rabbits (infracollicular decerebration) were prepared as described previously 7. There were 4 animals in each of these groups, and in 4 additional animals of each group the carotid sinus and aortic nerves (buffer nerves) were cut bilaterally in the neck immediately
Fig. 1. Method of deliveringcigarette smoke to nasal region of unanaesthetized rabbit.
173 •
$
II
$
II
•
AV mm
Hg
~
MF
~
~'F
56~
¢m/se¢
28 0
HR /rain
'L
o
4eo~ 240
_ _
- -
-
-
O I I'~
I 2 mlns
Fig. 2. Rabbit 2.5 kg, unanaesthetized. A continuous record is shown to illustrate both the stability of the circulation and the reproducibility of the response to successive cigarette smoke stimuli (at
arrows) delivered as in Fig. 1. (From ref. 15, with permission.) after the neurosurgical procedure, exactly as described previously 5. Sham-operated and thalamic animals recovered normal movement and posture within 0.5-1 h from the end of anaesthesia. Pontine rabbits lay on their side until 30 min before the start of the experiment; their body temperature was maintained by an electric blanket. Details of the postoperative management and posturing of these animals for experiments were as previously described 7. Nasopharyngeal stimulation was produced by blowing cigarette smoke gently through a narrow PVC tube to a position immediately in front of the animal's nose (Fig. 1), from where it was actively inhaled. The cardiorespiratory response in a given animal was highly reproducible (Fig. 2), and the response of a given animal was calculated as the mean of 3 stimuli given at 3-5 rain intervals. For each test, measurements of arterial pressure, heart rate, mesenteric blood flow and mesenteric 'conductance' (mesenteric flow/arterial pressure) were made just before smoke stimulation, and 7-12 sec later when the response was at its height. The duration of apnoea was also measured using a light AC-coupled carbonin-plastic strain-gauge strapped about the rabbit's thorax. The response to smoke stimulation was first observed during spontaneous ventilation and the animal was then given a short acting anaesthetic (propanidid, Epontol, Bayer) and intubated with a No. 14 Foregger paediatric endotracheal tube lubricated with Xylocaine jelly. It was then connected to a respiratory pump and ventilated at the average minute ventilation (1 1/min) and rate (60/min) of normal rabbits. While still under anaesthesia, muscular relaxation was induced by decamethonium iodide (1 mg/kg i.v., followed by 0.5 mg/kg i.v. every 20 min). The animals recovered from anaesthesia while ventilation was controlled, and periodic blood gas determinations were made and compared with the values for that animal when spontaneously breathing. Any differences were minimized by appropriate adjustments to the respiratory pump. No operative interventions were carried out while ventilation was controlled and decamethonium continued. Fifteen minutes after beginning controlled ventilation the circulatory effects of apnoea, in the absence of smoke stimulation, were studied by disconnecting the respiratory pump for 7-10 sec to simulate the apnoea of the normal response. Five minutes later the circulatory effects of blowing a small amount of smoke into the nose were studied while ventilation was maintained at 1 1/min. The alternate stimuli of apnoea alone and smoke alone were repeated twice more at 5 rain intervals.
174 The role of central mechanisms was assessed by comparing the evoked mean
changes in a given variable in the different preparations and the responses were expressed as per cent change using each animal as its own control. This method was used because in the present study the differences in initial resting values between preparations were generally ~mall and the conclusions reached by using either absolute or percentage changes were similar 17. Resting status of the different neural preparations. Because of the unusual nature of the preparations used in the present study, it is pertinent to review the status of the resting respiratory and circulatory variables before dealing with differences between responses. In rabbits with intact carotid sinus and aortic nerves resting respiration, arterial pressure, heart rate and mesenteric blood velocity differed little between shamoperated, thalamic and pontine animals (Table I). In rabbits with section of the buffer nerves, arterial pressure was higher than in corresponding groups in which the nerves were intact, whilst mesenteric blood velocity was lower. For this reason most of the quantitative comparisons of 'conductance' changes were made within each of the two subgroups of neural preparations with different buffer nerve status. Controlled ventilation had minimal effects on resting circulatory variables in any of the different preparations 17. Suprabulbar and bulbospinal regulation of respiration. In rabbits with intact buffer nerves, apnoea time was similar in sham-operated and pontine rabbits, but was shorter in thalamic animals (P = 0.05, Table II). In rabbits with sectioned buffer nerves, the duration of apnoea was longer than in the corresponding groups in which the nerves were intact, however, as in animals with intact buffer nerves the apnoea time of sham-operated and pontine animals was similar, but in thalamic animals it was shorter (P = 0.02). The longer apnoea of animals with an intact brain but with sectioned buffer nerves has been noted previously 10, and has been attributed to the absence of arterial chemoreceptor drive secondary to apnoea as asphyxial blood gas changes developed. The shorter duration of apnoea in thalamic animals compared TABLE I RESTING CARDIOVASCULARMEASUREMENTSIN SPONTANEOUSLYBREATHING NEURAL PREPARATIONS WITH AND WITHOUT INTACT CAROTID SINUS AND AORTIC NERVES Values a m averages a n d s t a n d a r d error o f m e a n for 4 animals. H R , heart rate, b e a t s / m i n ; AP, arterial pressure, m m H g ; MBV, mesenteric blood velocity, cm/sec.
Sham Thalamic Pontine
lntact carotid sinus and aortic nerves
Section carotid sinus and aortic nerves
HR
AP
MBV
HR
AP
MBV
264 (4.4) 266
94 (2.2) 93
23 (3.0) 21
292 (2.0) 245
104 (8.4) 106
16 (0.9) 14
(18.2)
(5.8)
(2.5)
(5.0)
(7.5)
(2.5)
220 (31.5)
82 (2.7)
22 (0.9)
293 (7.5)
94 (4.2)
13 (1.3)
175 TABLE II DURATION OF APNOEA EOR DIFFERENT NEURAL PREPARATIONS (GROUPS OF 4 ANIMALS) Values are averages 4- S.E.M.
Sham Thalamic Pontine
Intact carotid sinus and aortic nerves
Section carotid sinus and aortic nerves
7.8 ± 0.63 2.8 d: 1.60 9.3 d: 2.59
17.3 ± 1.44 8.5 ± 2.86 15.0 ± 1.50
with the other two neural preparations suggests that normally cerebral structures facilitate the apnoea initiated by trigeminal stimulation. The apnoea time of pontine animals on the other hand was the same as in sham-operated animals, suggesting in turn that diencephalic structures exert inhibitory effects on the reflex suppression of respiration evoked at bulbospinal levels, an effect normally masked by cerebral facilitation. It should be noted that the characteristic differences between the 3 neural preparations are present irrespective of the buffer nerve status, suggesting that chemoreceptor effects on respiration are predominantly exerted at bulbospinal levels. This finding is in agreement with conclusions drawn from other experiments in which respiratory drive was increased rather than inhibited7. Suprabulbar and bulbospinal regulation of heart rate. Trigeminal afferents could evoke their circulatory effects directly through projections to the pontomedullary reticular formation1, and/or indirectly due to changes associated with the sudden inhibition of respiration in expiration, i.e., the sudden reduction in the input via vagal afferents from lung inflation receptors2,3,1% In addition, with the onset of apnoea and bradycardia, there is a marked increase in atrial transmural pressure 13, and previous studies have shown that arterial baroreceptors contribute to the reduction in heart rate due, most probably, to the rise in arterial pressure. Changes in chemoreceptor activity contribute to the termination of the apnoea but appear to play little part in the bradycardia of the single bolus smoke stimulus10. In the current study, bradycardia was evoked by apnoea without smoke stimulation, and also by smoke stimulation without apnoea. The effects of apnoea alone on heart rate appeared to depend largely on the integrity of the buffer nerves, since after their section there was only a small residual bradycardia (P < 0.05, Fig. 3). This in turn was dependent on intact cerebral hemispheres (P < 0.05). It may be suggested, therefore, that the reflex heart rate effects of sudden loss of lung inflation are partly dependent on arterial barorecepter interactions with lung inflation afferents, and partly on arterial baroreceptor independent mechanisms. After buffer nerve section, the afferent drive induced by apnoea alone arises from lung inflation receptors, cardiac receptors, or both, and is transmitted through vagal afferents2,3,11,1L Other studies have indicated that the arterial baroreceptor-heart rate reflex is dependent on supra~pontine regions 6,8, and the present study suggests that the reflex response to
176 ISECTION C.S.+A.N.J
IINTACT C.S.+AN.I MEAN ARTERIAL PRESSURE %
t
Sh
Th
Po
•
II
m
I00
HEART RATE %
%
801 60 40
oO; oE
I '°-°
20
1
Fig. 3. Haemodynamic effects of simulated apnoea in expiration in the absence of cigarette smoke stimulation in groups of 4 sham-operated rabbits (Sh), thalamic rabbits (Th) and pontine rabbits (Po). Left 3 panels, preparations with intact carotid sinus and aortic nerves; fight 3 panels, preparations in which the carotid sinus and aortic nerves have been cut. The symbol on the left = 2 S.E. of mean difference between control and response values for any one of the 3 responses, calculated by analysis of variance. The mean absolute resting data for these rabbits and those in Figs. 4 and 5 are shown in Table I. a p n o e a alone made by vagal afferents on the one hand, and the carotid sinus and aortic nerves on the other are both dependent on suprapontine regions. The bradycardia induced by nasal smoke stimulation without apnoea in rabbits with section of the buffer nerves probably occurs t h r o u g h trigeminal afferents (Fig. 4). In these animals the reflex effects in part depend on intact cerebral hemispheres, since cardiac slowing was greatest in sham-operated animals (P < 0.05). W h e n the buffer nerves were intact, smoke stimulation without apnoea caused greater bradycardia (P < 0.05) which was similar in magnitude in all 3 neurological preparations. The trigeminal input thus appears to have direct effects on heart rate neurones which are dependent on mechanisms involving the cerebral hemispheres. In addition, there appears to be an interaction effect between trigeminal and arterial baroreceptor afferents at bulbospinal levels. iINTACT C.S.+A.N] Sh
ARTERIAL
PRESSURE HEART RATE %
'40
Th
!1
Po
[SECT,ON CS÷AN] Sh
Th
EQ
I l l
80I 6O 40
FLOW
60 F
CONDUCTANCE I % 2O Fig. 4. Effects of nasal stimulation with smoke during controlled ventilation. Notations as in Fig. 3.
177 I INTACT C.S.*A.N. I Sh MEAN ARTERIAL PRESSURE %
140 I00 L
RCTE
f
~
Th
milll
[ SECTION C.S.+A.N, [
Po
Sh ~ ,[
Th
Po
i..m
60f
FLOW CONDUCTANCE % 20
Fig. 5. Effectsof smoke stimulation in spontaneouslybreathing animals. The respiratory (apnoea time) data from these experiments is shown in Table II. Notations as in Fig. 3.
In view of the above findings, in the spontaneously breathing animal with section of the buffer nerves one would expect bradycardia of the smoke reflex to be present but less than normal in all neural preparations. Moreover, in view of the importance of the cerebral hemispheres for reflexes from both the trigeminal and lung inflation inputs, the cardiac slowing might be expected to be greater in the animals with intact brains. These expectations were confirmed experimentally and are shown in Fig. 5. In the intact animal, the bradycardia due to smoke inhalation thus appears to be a complex response depending on at least 3 peripheral inputs: (1) the direct effects of trigeminal stimulation, (2) the effects of apnoea predominantly through various vagal afferents and (3) the arterial baroreceptors. Suprabulbar and bulbospinal regulation of the mesenteric circulation. Central nervous regulation of the mesenteric circulation through adrenergic vasoconstrictor fibres is similar in certain respects to the regulation of heart rate. Thus in animals in which the buffer nerves are cut, the effects of apnoea without smoke stimulation and smoke stimulation without apnoea differ in magnitude between sham-operated, thalamic and pontine animals (Figs. 3 and 4). Mesenteric vasoconstriction evoked by apnoea alone is dependent on cerebral mechanisms, since only in sham-operated animals is there any fall in 'conductance' (P < 0.05, Fig. 3). Likewise, smoke stimulation alone, although reducing 'conductance' in all 3 neural preparations, does so to the greatest extent in sham-operated animals (Fig. 4). These data, as with the regulation of heart rate, suggest a role for the cerebral hemispheres in the regulation of sympathetic vasoconstrictor fibre activity through both the trigeminal and lung inflation inputs which are independent of the arterial baroreceptor input. Clearly, there is also a direct control through the trigeminal input, which appears to act at a bulbospinal level, but there is no equivalent control through the lung inflation input (el Figs. 3 and 4, right panels, pontine animals). When the buffer nerves are intact the reduction in conductance through either input is considerably enhanced. This possibly reflects a greater degree of reflex vasoconstriction; however, this interpretation is uncertain in view of the differences in resting conductance between animals with and
178 without section of the buffer nerves. Moreover, the rise in arterial pressure might be expected to stimulate arterial baroreceptor reflexes to minimize the mesenteric vasoconstriction rather than accentuate it. It is possible therefore that the arterial chemoreceptor input is responsible for the enhanced vasoconstriction, even though its role in the concomitant reflex bradycardia is negligible 10. An alternative explanation is that projections from trigeminal and vagal afferents converge on vasoconstrictor neurones also receiving arterial baroreceptor afferents, producing 'resetting' of barereflex threshold and gain s . In this way, constrictor effects would be enhanced despite the rise in arterial pressure. Of particular interest in thalamic animals with intact buffer nerves is the vasoconstriction evoked by apnoea alone, which is not present in these preparations when the buffer nerves are cut, or in pontine animals (Fig. 3). These data suggest that suprapontine interactions between lung inflation and arterial baroreceptor afferents play an important role in the vasoconstriction evoked by apnoea alone. Conclusions. It is concluded that when nasopharyngeal stimulation occurs in the intact rabbit there is suprabulbar as well as bulbospinal regulation of respiration and the circulation. The return of respiration after reflex suppression of breathing by trigeminal stimulation is partly a function of arterial chemoreceptor activity acting on bulbospinal neurones. Trigeminal, lung inflation and arterial baroreceptor afferents all appear to play a role in exciting heart rate and sympathetic vasoconstrictor pathways, and arterial baroreceptor-lung inflation and arterial baroreceptor-trigeminal interactions appear to mutually enhance autonomic activity at different levels of the central nervous system. An alternative explanation is that brain sections interrupt facilitatory pathways descending to autonomic neurones in bulbospinal regions. In both cases the results demonstrate the role of suprapontine mechanisms in the autonomic response to a complex input profile elicited by a simple environmental stimulus. These studies were carried out in collaboration with Dr. R. J. McRitchie and Prof. P. I. Korner. Mr. D. Lauff gave valuable technical assistance. The study was supported by research grants from the Life Insurance Medical Research Fund of Australia and New Zealand, the National Heart Foundation of Australia, the Australian Tobacco Research Foundation, the National Health and Medical Research Council of Australia and the Postgraduate Medical Foundation of the University of Sydney.
1 BRODAL,A., Neurological Anatomy, Oxford Univ. Press, London, 1969, 415 pp. 2 DALY, M. DE B., AND HAZZLEDINE, J. L., The effects of artificially induced hyperventilation on the
primary cardiac reflex response to stimulation of the carotid bodies in the dog, J. PhysioL (Lend.), 168 (1963) 872-889. 3 DALY, M. DE B., AND ROBINSON, B. H., An analysis of the reflex systemic vasodilator repsonse elicited by lung inflation in the dog, J. Physiol. (Lend.), 195 (1967) 387~106. 4 FORSTER,R. P., ANDNYBOER,J., Effect of induced apnoea on cardiovascular renal functions in the rabbit, Amer. J. Physiol., 183 (1955) 149-154. 5 KORNER, P. I., Effect of section of the carotid sinus and aortic nerves on cardiac output of the rabbit, J. Physiol. (Lend.), 180 (1965) 266-278.
179 6 KORNER,P. I., SHAW, J., WEST, M. J., AND OLIVER, J. R., Central nervous system control of baroreceptor reflexes in the rabbit, Circulat. Res., 31 (1972) 637-652. 7 KORNER,P. I., UTHER, J. B., AND WHITE, S. W., Central nervous integration of the circulatory and respiratory response to arterial hypoxemia in the rabbit, Circulat. Res., 24 (1969) 757-776. 8 KORNER, P. I., WEST, M. J., AND SHAW, J., Central nervous resetting of baroreceptor reflexes, Aust. J. exp. Biol. Med. ScL, 51 (1973) 53-64. 9 KRETSCHMER,F., Ll~oerReflexe vonder Nasenschleimhaut auf Atmung und Kreislauf, Sber. Akad. Wiss. (Wien), 62 (1870) 147. 10 McRITCHIE, R. J., ANO WHITE, S. W., Role of trigeminal, olfactory, carotid sinus and aortic nerves in the respiratory and circulatory response to nasal inhalation of cigarette smoke and other irritants in the rabbit, Aust. J. exp. BioL Med~ Sci., 52 (1974) 127-140. 11 OBERG,B., AND WHITE, S. W., Circulatory effects of interruption and stimulation of cardiac vagal afferents, Acta physiol, scand., 80 (1970) 383-394. 12 OaT, N. T., AND SrmPHERD, J. T., Vasodepressor reflex from lung inflation in the rabbit, Amer. J. Physiol., 221 (1971) 889-895. 13 PAINTAL,A. S., A study of right and left atrial receptors, J. Physiol. (Lond.), 120 (1953) 596-610. 14 WHITE,S. W., ANGUS, J. A., McRITCHIE, R. J., AND PORGES, W. L., Evaluation of the Doppler Flowmeter for measurement of blood flow in small vessels of unanaesthetized animals. In J. R. S. HALES AND S. W. WHITE (Eds.), Proc. Syrup. Clin. exp. Blood Flow Measurement, Clin. exp. Pharm. Physiol., Suppl. 1 (1974) 79-92. 15 WHITE,S. W., AND McRrrCHIE, R. J., Nasopharyngeal reflexes: integrative analysis of evoked respiratory and cardiovascular effects, Aust. J. exp. BioL Med. Sci., 51 (1973) 17-31. 16 WHITE, S. W., McRITCHIE, R. J., AND FRANKLIN,D. L., Autonomic cardiovascular effects of nasal inhalation of cigarette smoke in the rabbit, Aust. J. exp. Biol. Med. Sci., 52 (1974) 111-126. 17 WHITE, S. W., MCRITCHIE, R. J., AND KORNER, P. I., CNS control of cardio-respiratory nasopharyngeal reflexes in the rabbit, Amer. J. PhysioL, In press.