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Post-stimulus facilitatory and inhibitory effects on respiration induced by chemical and electrical stimulation of thin-fiber muscular afferents in dogs
Neuroscience Letters, 35 (1983) 283-287
283
Elsevier Scientific Publishers Ireland Ltd.
POST-STIMULUS FACILITATORY AND INHIBITORY EFFECTS ON RESPIRATION INDUCED BY CHEMICAL AND ELECTRICAL STIMULATION OF THIN-FIBER MUSCULAR AFFERENTS IN DOGS
T. KUMAZAWA*, E. TADAKI, K. MIZUMURA and K. KIM
Department of Nervous and Sensory Functions, Research Institute of Environmental Medicine, Nagoya University, Nagoya, 464 and Department of Physiology, Nagoya University School of Medicine, Nagoya, 466 (Japan) (Received December 6th, 1982; Accepted December 24th, 1982)
Key words: polymodal receptor - A~5 fiber afferents - C fiber afferents - muscular afferents respiratory reflex - post-stimulus inhibition - post-stimulus facilitation.
In anesthetized, vagotomized, paralyzed, and artificially ventilated dogs, respiratory responses to both electrical stimulation of the muscle nerve and chemical stimulation of muscular (polymodal) receptors by means of intra-arterial injection o f NaC1 solution were studied by recording phrenic nervous discharges. During the period of stimulation both types of stimulation caused intensity-dependent facilitation of neural respiratory outputs. After cessation o f stimulation, facilitation persisted for a long time (more than 5 min) with a lower intensity stimulation; however, suppression was observed with a higher intensity stimulation. The present results suggest that afferent inputs from the muscular polymodal receptors activate longacting central mechanisms for enhancement or suppression of respiration.
It is known that stimulation of thin-fiber muscular afferents reflexively induces respiratory and cardiovascular changes [2-4, 10, 11, 14-16]. Recordings of unitary discharges of muscular afferents revealed that the majority of the At5 and C fiber units responded to mechanical, chemical, and heat stimuli maximally in a noxious level [5-7, 12]. Thus, these units are best described as polymodal receptors [5-7]. Our previous observation demonstrated that arterial injection of different chemicals into the muscle caused a similar degree of increase in minute respiratory volume to the increase in discharge rates of muscular polymodal receptors to the same stimulus, suggesting that polymodal receptors play a role in respiratory responses caused by muscular afferents [14]. In the course of a similar line of investigation performed on artificially ventilated dogs, it was noticed that stimulation of the thin-fiber muscular afferents caused long-persisting post-stimulus effects of either enhancement or suppression of neural * Author for correspondence at first address.
0304-3940/83/0000-0000/$ 03.00 © 1983 Elsevier Scientific Publishers Ireland Ltd.
284
respiratory outputs indicated by the phrenic nervous activity. Our preliminary observations suggested that one factor causing the difference might be the intensity of the stimulation [10]. The present work was undertaken to study the effects of changing the intensities of afferent electrical stimulation of the muscle nerve and of changing the concentrations of NaC1 solution injected intra-arterially into the muscle. NaCl solutions have been found to be one of the most consistent and potent stimulants for the polymodal receptors [6-9]. The experiments were performed on mongrel dogs anesthetized with chloralose (35 mg/kg) and urethane (250 m g / k g ) under preliminary thiopenthal anesthesia. Subsequently, chloralose and urethane were added to maintain the state of anesthesia. The animal was paralyzed and artificially ventilated and end-tidal CO2 and 02 were monitored by a gas analyzer (San-ei Sokki). The mean end-tidal CO2 was 4.9°7o and fluctuation of the end4idal CO2 level in one experimental series was kept within 0. I o70 by adjusting the ventilation volume. The esophageal temperature was monitored and its fluctuation was kept within 0.2°C. The mean arterial pressure was recorded f r o m the left c o m m o n carotid artery. The right carotid sinus nerve and both vagi were cut. For chemical stimulation of the gastrocnemius-soleus muscle, 4 lots of 5 ml of NaCl solution with concentrations of 1.8, 2.7, 3.6 and 4.5°7o, respectively, were retrogradely injected into the muscular branch of the caudal femoral artery at a constant rate of 5 ml/50 sec, while the femoral artery and vein were transiently closed at the femoral ring and at the popliteal fossa from 30 sec before the onset of the arterial injection until 1 min after the end of it. Arteries and veins, and the femoral nerve and branches of the sciatic nerve, except those supplying the gastrocnemius-soleus muscle, were ligated or cut in order to limit the area of stimulation to those muscles. Each of the 4 solutions was injected in a random order with an interval of 10 min, to allow the state of respiration to recover from the effect of the preceding stimulation. The injection of Ringer solution by the same method did not significantly modify the respiration of the animals in any case. For electrical stimulation of the muscular afferents, the gastrocnemius-soleus muscle nerve was isolated and cut close to the muscle. The distal end of the nerve was stimulated, while compound action potentials were monitored at the proximal part of the nerve. The strength of the electrical stimulation was set at multiples of the threshold (T) for the action potential. The intensities of the electrical stimulation used were mainly 20, 100, 200 and 400 T; occasionally l0 and 600 T were also tested. Pulses with a repetition rate of 8 Hz were applied for 1 min with an interval of 10 min. The phrenic nervous activity was recorded in an oil pool at the neck, and was rectified and integrated by means of an RC-circuit with a time constant of 0.1 sec. The peak amplitude and period for each breath were measured by means of a microcomputer. In order to make comparison among experiments possible, each peak value of the integrated phrenic discharges was normalized by dividing it by the mean value during the 1 min period before the first stimulation in the experiment.
285
Breath-by-breath measure of neural respiratory outputs was expressed as the product of the normalized peak integrated value and the instantaneous respiratory rate. The breath-by-breath respiratory outputs were averaged for a 30-sec or a 1-min period as a mean respiratory output in an arbitrary unit. Fig. IA shows the averaged responses in mean respiratory outputs in 20 dogs induced by the 4 different concentrations of NaCI solution. The procedure of stopping blood flow caused the slightest change in mean respiratory outputs (FS in Fig. 1). A W
~=
30
(9 1.8c)(~NaCI (~ • 3.696
2.7~NaCI NaCI • 4.5~ NaCI
I,.-
30
==:
~-
/•
/~~~00
© 10-20T, 8HZ 8HZ
~(0 T,
25
25 20
,,x 20
..=.
15 15
iniection
"~1
stim
TIME(min~
Fig. 1. A: respiratory responses to intra-arterial injection of different concentrations of NaC1 solution into the gastrocnemius-soleus muscle of dogs. Ordinate, averaged respiratory outputs, see text; Abscissa, time. P, before stimulation; FS, stopping the blood flow to the muscle; FO, re-opening the blood flow to the muscle; 1-5, 1-5 rain after re-opening the blood flow. Averaged values from 20 dogs; 4 concentrations of NaCI solution were tested in a random order in each dog. B: respiratory responses to afferent electrical stimulation of the gastrocnemius nerve of dogs. The same presentation as A. Averaged respiratory outputs obtained in 29 dogs. In each dog, both low intensity (10 T or 20 T) and high intensity (100, 200, 400 or 600 T) stimulations were tested.
Injection of the solutions increased the mean respiratory output in parallel with the increase of the concentration of the solutions. A significant correlation was found between the concentration of the solutions and the induced increases in mean respiratory outputs in the latter 30 sec of the injection period (L in Fig. 1A). On the other hand, changes in mean respiratory outputs after the re-opening of blood flow were different depending on the concentration of the solutions. With the 1.8°70 solution, the mean respiratory outputs remained high in the following 5 min; with the 2.7% solution, they returned to almost the same level as the pre-stimulus value; with the 3.6 and 4.5% solutions, they decreased to below the pre-stimulus
286 value. Therefore, it can be said that inputs from muscular receptors which respond to NaCI cause dose-dependent increases in respiratory output during the stimulating period and also cause facilitatory or inhibitory effects in the post-stimulation period, depending on the doses. The respiratory effects of afferent electrical stimulation of the muscle nerve were studied in 29 dogs in which both low intensities (below 20 T) and high intensities (above 100 T) o f stimulation were tested. Fig. 1B shows averaged responses in the mean respiratory outputs to both types of stimulation. Facilitation of respiration lasted for more than 5 min after the end of low intensity electrical stimulation, while post-stimulus suppression of respiration was observed with high intensity stimulation. The present study reveals a dose-dependent increase in phrenic nervous activities during intra-arterial injection of NaC1, which has been shown to be a consistent and potent chemical stimulant for the polymodal receptors. Furthermore, the present experiments performed at constant end-tidal CO2 and 02 levels, reveals that the effects on respiration last long after the cessation of stimulation. These long-lasting poststimulation respiratory effects cannot be interpreted as an effect of the afterdischarges of polymodal receptors [6, 7], since the post-stimulatoin respiratory effects outlast the period of the afterdischarges of the receptors, and are also obtained by electrical stimulation of the nerve. Thus, afferent input from the muscular thin-fiber receptors seem to activate longacting central mechanisms for enhancement or suppression of respiration depending on the level of the inputs. Actual central mechanisms participating in these phenomena are not yet known. The long-lasting nature of the post-stimulation respiratory effects led us to suppose participation of neurohumoral factors in these phenomena. A participation of endogenous opiates in the post-stimulation inhibitory effects has been suggested as one factor among them [10]. Eldridge and his collaborators have reported two types of post-stimulation facilitatory effects on respiration in cats [1, 13]: one lasting for 200 or 300 sec, and the other for more than 1 h after the cessation of stimulation. The former effect was elicited by either carotid sinus nerve stimulation or calf muscle squeezing, while the latter effect was obtained by means of the carotid sinus nerve stimulation but not by muscle squeezing. However, we have observed facilitatory post-stimulation effects lasting more than 5 min in 186 trials out of 713 electrical stimulation trials carried out on 93 dogs (including unpublished data). The effects could be recorded for more than 1 h in some trials. Muscular afferents definitely cause long-lasting facilitatory effects on respiration in dogs. The absence o f long-persisting activation of respiration by muscular afferents in the report of Eldridge's group might be due to different experimental conditions, including the difference in animal species. This work was partly supported by the Naito Research Grant for 1979 and by a grant from the Research Foundation for Oriental Medicine.
287 ! Eldridge, F.L., Central neural respiratory stimulatory effect of active respiration, J. appl. Physiol., 37 (1974) 723-735. 2 Hodgson, H.J.F. and Matthews, P.B.C., The ineffectiveness of excitation of the primary endings of the muscle spindle by vibration as a respiratory stimulant in the decerebrate cat, J. Physiol. (Lond.), 194 (1968) 555-563. 3 Kalia, M., Senapati, J.M., Parida, B. and Panda, A., Reflex increase in ventilation by muscle receptors with non-medullated fibers (C fibers), J. appl. Physiol., 32 (1972) 189-193. 4 Kao, F.F., An experimental study of the pathways involved in exercise hyperpnoea employing crosscirculation techniques. In D.J.C. Cunningham and B.B. Lloyd (Eds.), The Regulation of Human Respiration, Blackwell, Oxford, 1963, pp. 461-502. 5 Kniffki, K.D., Mense, S. and Schmidt, R.F., Responses of group IV afferent units from skeletal muscle to stretch, contraction and chemical stimulation, Exp. Brain Res., 31 (1978) 511-522. 6 Kumazawa, T. and Mizumura, K., The polymodal C-fiber receptor in the muscle of the dog, Brain Res., 101 (1976) 589-593. 7 Kumazawa, T. and Mizumura, K., Thin-fiber receptors responding to mechanical, chemical, and thermal stimulation in the skeletal muscle of the dog, J. Physiol. (Lond.), 273 (1977) 179-194. 8 Kumazawa, T. and Mizumura, K., Chemical responses of polymodal receptors of the scrotal contents in dogs, J. Physiol. (Lond.), 299 (1980) 219-231. 9 Kumazawa, T. and Mizumura, K., Mechanical and thermal responses of polymodal receptors recorded from the superior spermatic nerve of dogs, J. Physiol. (Lond.), 299 (1980) 233-245. 10 Kumazawa, T., Tadaki, E. and Kim, K., A possible participation of endogenous opiates in respiratory reflexes induced by thin-fiber muscular afferents, Brain Res., 199 (1980) 244-248. 11 McCloskey, D.I. and Mitchell, J.H., Reflex cardiovascular and respiratory responses originating in excercising muscle, J. Physiol. (Lond.), 224 (1972) 173-186. 12 Mense, S. and Schmidt, R.F., Activation of group IV afferent units from muscle by algesic agents, Brain Res., 72 (1974) 305-310. 13 Millhorn, D.E., Eldridge, F.L. and Waldrop, T.G., Prolonged stimulation of respiration by a new central neural mechanism, Respirat. Physiol., 41 (1980) 87-103. 14 Mizumura, K. and Kumazawa, T., Reflex respiratory response induced by chemical stimulation of muscle afferents, Brain Res., 109 (1976) 402-406. 15 Perez-Gonzalez, J.F. and Coote, J.H., Activity of muscle afferents and reflex circulatory responses to exercise, Amer. J. Physiol., 223 (1972) 138-143. 16 Sato, A., Sato, Y. and Schmidt, R.F., Changes in heart rate and blood pressure upon injection of algesic agents into skeletal muscle, Pfliigers Arch., 393 (1982) 31-36.
283
Elsevier Scientific Publishers Ireland Ltd.
POST-STIMULUS FACILITATORY AND INHIBITORY EFFECTS ON RESPIRATION INDUCED BY CHEMICAL AND ELECTRICAL STIMULATION OF THIN-FIBER MUSCULAR AFFERENTS IN DOGS
T. KUMAZAWA*, E. TADAKI, K. MIZUMURA and K. KIM
Department of Nervous and Sensory Functions, Research Institute of Environmental Medicine, Nagoya University, Nagoya, 464 and Department of Physiology, Nagoya University School of Medicine, Nagoya, 466 (Japan) (Received December 6th, 1982; Accepted December 24th, 1982)
Key words: polymodal receptor - A~5 fiber afferents - C fiber afferents - muscular afferents respiratory reflex - post-stimulus inhibition - post-stimulus facilitation.
In anesthetized, vagotomized, paralyzed, and artificially ventilated dogs, respiratory responses to both electrical stimulation of the muscle nerve and chemical stimulation of muscular (polymodal) receptors by means of intra-arterial injection o f NaC1 solution were studied by recording phrenic nervous discharges. During the period of stimulation both types of stimulation caused intensity-dependent facilitation of neural respiratory outputs. After cessation o f stimulation, facilitation persisted for a long time (more than 5 min) with a lower intensity stimulation; however, suppression was observed with a higher intensity stimulation. The present results suggest that afferent inputs from the muscular polymodal receptors activate longacting central mechanisms for enhancement or suppression of respiration.
It is known that stimulation of thin-fiber muscular afferents reflexively induces respiratory and cardiovascular changes [2-4, 10, 11, 14-16]. Recordings of unitary discharges of muscular afferents revealed that the majority of the At5 and C fiber units responded to mechanical, chemical, and heat stimuli maximally in a noxious level [5-7, 12]. Thus, these units are best described as polymodal receptors [5-7]. Our previous observation demonstrated that arterial injection of different chemicals into the muscle caused a similar degree of increase in minute respiratory volume to the increase in discharge rates of muscular polymodal receptors to the same stimulus, suggesting that polymodal receptors play a role in respiratory responses caused by muscular afferents [14]. In the course of a similar line of investigation performed on artificially ventilated dogs, it was noticed that stimulation of the thin-fiber muscular afferents caused long-persisting post-stimulus effects of either enhancement or suppression of neural * Author for correspondence at first address.
0304-3940/83/0000-0000/$ 03.00 © 1983 Elsevier Scientific Publishers Ireland Ltd.
284
respiratory outputs indicated by the phrenic nervous activity. Our preliminary observations suggested that one factor causing the difference might be the intensity of the stimulation [10]. The present work was undertaken to study the effects of changing the intensities of afferent electrical stimulation of the muscle nerve and of changing the concentrations of NaC1 solution injected intra-arterially into the muscle. NaCl solutions have been found to be one of the most consistent and potent stimulants for the polymodal receptors [6-9]. The experiments were performed on mongrel dogs anesthetized with chloralose (35 mg/kg) and urethane (250 m g / k g ) under preliminary thiopenthal anesthesia. Subsequently, chloralose and urethane were added to maintain the state of anesthesia. The animal was paralyzed and artificially ventilated and end-tidal CO2 and 02 were monitored by a gas analyzer (San-ei Sokki). The mean end-tidal CO2 was 4.9°7o and fluctuation of the end4idal CO2 level in one experimental series was kept within 0. I o70 by adjusting the ventilation volume. The esophageal temperature was monitored and its fluctuation was kept within 0.2°C. The mean arterial pressure was recorded f r o m the left c o m m o n carotid artery. The right carotid sinus nerve and both vagi were cut. For chemical stimulation of the gastrocnemius-soleus muscle, 4 lots of 5 ml of NaCl solution with concentrations of 1.8, 2.7, 3.6 and 4.5°7o, respectively, were retrogradely injected into the muscular branch of the caudal femoral artery at a constant rate of 5 ml/50 sec, while the femoral artery and vein were transiently closed at the femoral ring and at the popliteal fossa from 30 sec before the onset of the arterial injection until 1 min after the end of it. Arteries and veins, and the femoral nerve and branches of the sciatic nerve, except those supplying the gastrocnemius-soleus muscle, were ligated or cut in order to limit the area of stimulation to those muscles. Each of the 4 solutions was injected in a random order with an interval of 10 min, to allow the state of respiration to recover from the effect of the preceding stimulation. The injection of Ringer solution by the same method did not significantly modify the respiration of the animals in any case. For electrical stimulation of the muscular afferents, the gastrocnemius-soleus muscle nerve was isolated and cut close to the muscle. The distal end of the nerve was stimulated, while compound action potentials were monitored at the proximal part of the nerve. The strength of the electrical stimulation was set at multiples of the threshold (T) for the action potential. The intensities of the electrical stimulation used were mainly 20, 100, 200 and 400 T; occasionally l0 and 600 T were also tested. Pulses with a repetition rate of 8 Hz were applied for 1 min with an interval of 10 min. The phrenic nervous activity was recorded in an oil pool at the neck, and was rectified and integrated by means of an RC-circuit with a time constant of 0.1 sec. The peak amplitude and period for each breath were measured by means of a microcomputer. In order to make comparison among experiments possible, each peak value of the integrated phrenic discharges was normalized by dividing it by the mean value during the 1 min period before the first stimulation in the experiment.
285
Breath-by-breath measure of neural respiratory outputs was expressed as the product of the normalized peak integrated value and the instantaneous respiratory rate. The breath-by-breath respiratory outputs were averaged for a 30-sec or a 1-min period as a mean respiratory output in an arbitrary unit. Fig. IA shows the averaged responses in mean respiratory outputs in 20 dogs induced by the 4 different concentrations of NaCI solution. The procedure of stopping blood flow caused the slightest change in mean respiratory outputs (FS in Fig. 1). A W
~=
30
(9 1.8c)(~NaCI (~ • 3.696
2.7~NaCI NaCI • 4.5~ NaCI
I,.-
30
==:
~-
/•
/~~~00
© 10-20T, 8HZ 8HZ
~(0 T,
25
25 20
,,x 20
..=.
15 15
iniection
"~1
stim
TIME(min~
Fig. 1. A: respiratory responses to intra-arterial injection of different concentrations of NaC1 solution into the gastrocnemius-soleus muscle of dogs. Ordinate, averaged respiratory outputs, see text; Abscissa, time. P, before stimulation; FS, stopping the blood flow to the muscle; FO, re-opening the blood flow to the muscle; 1-5, 1-5 rain after re-opening the blood flow. Averaged values from 20 dogs; 4 concentrations of NaCI solution were tested in a random order in each dog. B: respiratory responses to afferent electrical stimulation of the gastrocnemius nerve of dogs. The same presentation as A. Averaged respiratory outputs obtained in 29 dogs. In each dog, both low intensity (10 T or 20 T) and high intensity (100, 200, 400 or 600 T) stimulations were tested.
Injection of the solutions increased the mean respiratory output in parallel with the increase of the concentration of the solutions. A significant correlation was found between the concentration of the solutions and the induced increases in mean respiratory outputs in the latter 30 sec of the injection period (L in Fig. 1A). On the other hand, changes in mean respiratory outputs after the re-opening of blood flow were different depending on the concentration of the solutions. With the 1.8°70 solution, the mean respiratory outputs remained high in the following 5 min; with the 2.7% solution, they returned to almost the same level as the pre-stimulus value; with the 3.6 and 4.5% solutions, they decreased to below the pre-stimulus
286 value. Therefore, it can be said that inputs from muscular receptors which respond to NaCI cause dose-dependent increases in respiratory output during the stimulating period and also cause facilitatory or inhibitory effects in the post-stimulation period, depending on the doses. The respiratory effects of afferent electrical stimulation of the muscle nerve were studied in 29 dogs in which both low intensities (below 20 T) and high intensities (above 100 T) o f stimulation were tested. Fig. 1B shows averaged responses in the mean respiratory outputs to both types of stimulation. Facilitation of respiration lasted for more than 5 min after the end of low intensity electrical stimulation, while post-stimulus suppression of respiration was observed with high intensity stimulation. The present study reveals a dose-dependent increase in phrenic nervous activities during intra-arterial injection of NaC1, which has been shown to be a consistent and potent chemical stimulant for the polymodal receptors. Furthermore, the present experiments performed at constant end-tidal CO2 and 02 levels, reveals that the effects on respiration last long after the cessation of stimulation. These long-lasting poststimulation respiratory effects cannot be interpreted as an effect of the afterdischarges of polymodal receptors [6, 7], since the post-stimulatoin respiratory effects outlast the period of the afterdischarges of the receptors, and are also obtained by electrical stimulation of the nerve. Thus, afferent input from the muscular thin-fiber receptors seem to activate longacting central mechanisms for enhancement or suppression of respiration depending on the level of the inputs. Actual central mechanisms participating in these phenomena are not yet known. The long-lasting nature of the post-stimulation respiratory effects led us to suppose participation of neurohumoral factors in these phenomena. A participation of endogenous opiates in the post-stimulation inhibitory effects has been suggested as one factor among them [10]. Eldridge and his collaborators have reported two types of post-stimulation facilitatory effects on respiration in cats [1, 13]: one lasting for 200 or 300 sec, and the other for more than 1 h after the cessation of stimulation. The former effect was elicited by either carotid sinus nerve stimulation or calf muscle squeezing, while the latter effect was obtained by means of the carotid sinus nerve stimulation but not by muscle squeezing. However, we have observed facilitatory post-stimulation effects lasting more than 5 min in 186 trials out of 713 electrical stimulation trials carried out on 93 dogs (including unpublished data). The effects could be recorded for more than 1 h in some trials. Muscular afferents definitely cause long-lasting facilitatory effects on respiration in dogs. The absence o f long-persisting activation of respiration by muscular afferents in the report of Eldridge's group might be due to different experimental conditions, including the difference in animal species. This work was partly supported by the Naito Research Grant for 1979 and by a grant from the Research Foundation for Oriental Medicine.
287 ! Eldridge, F.L., Central neural respiratory stimulatory effect of active respiration, J. appl. Physiol., 37 (1974) 723-735. 2 Hodgson, H.J.F. and Matthews, P.B.C., The ineffectiveness of excitation of the primary endings of the muscle spindle by vibration as a respiratory stimulant in the decerebrate cat, J. Physiol. (Lond.), 194 (1968) 555-563. 3 Kalia, M., Senapati, J.M., Parida, B. and Panda, A., Reflex increase in ventilation by muscle receptors with non-medullated fibers (C fibers), J. appl. Physiol., 32 (1972) 189-193. 4 Kao, F.F., An experimental study of the pathways involved in exercise hyperpnoea employing crosscirculation techniques. In D.J.C. Cunningham and B.B. Lloyd (Eds.), The Regulation of Human Respiration, Blackwell, Oxford, 1963, pp. 461-502. 5 Kniffki, K.D., Mense, S. and Schmidt, R.F., Responses of group IV afferent units from skeletal muscle to stretch, contraction and chemical stimulation, Exp. Brain Res., 31 (1978) 511-522. 6 Kumazawa, T. and Mizumura, K., The polymodal C-fiber receptor in the muscle of the dog, Brain Res., 101 (1976) 589-593. 7 Kumazawa, T. and Mizumura, K., Thin-fiber receptors responding to mechanical, chemical, and thermal stimulation in the skeletal muscle of the dog, J. Physiol. (Lond.), 273 (1977) 179-194. 8 Kumazawa, T. and Mizumura, K., Chemical responses of polymodal receptors of the scrotal contents in dogs, J. Physiol. (Lond.), 299 (1980) 219-231. 9 Kumazawa, T. and Mizumura, K., Mechanical and thermal responses of polymodal receptors recorded from the superior spermatic nerve of dogs, J. Physiol. (Lond.), 299 (1980) 233-245. 10 Kumazawa, T., Tadaki, E. and Kim, K., A possible participation of endogenous opiates in respiratory reflexes induced by thin-fiber muscular afferents, Brain Res., 199 (1980) 244-248. 11 McCloskey, D.I. and Mitchell, J.H., Reflex cardiovascular and respiratory responses originating in excercising muscle, J. Physiol. (Lond.), 224 (1972) 173-186. 12 Mense, S. and Schmidt, R.F., Activation of group IV afferent units from muscle by algesic agents, Brain Res., 72 (1974) 305-310. 13 Millhorn, D.E., Eldridge, F.L. and Waldrop, T.G., Prolonged stimulation of respiration by a new central neural mechanism, Respirat. Physiol., 41 (1980) 87-103. 14 Mizumura, K. and Kumazawa, T., Reflex respiratory response induced by chemical stimulation of muscle afferents, Brain Res., 109 (1976) 402-406. 15 Perez-Gonzalez, J.F. and Coote, J.H., Activity of muscle afferents and reflex circulatory responses to exercise, Amer. J. Physiol., 223 (1972) 138-143. 16 Sato, A., Sato, Y. and Schmidt, R.F., Changes in heart rate and blood pressure upon injection of algesic agents into skeletal muscle, Pfliigers Arch., 393 (1982) 31-36.