- Email: [email protected]
The effects of naloxone on the neural control of the urinary bladder of the cat
355
Brain Research, 264 (1983) 355 358 Elsevier Biomedical Press
The effects of naloxone on the neural control of the urinary bladder of the cat J, R. ROPPOLO, A. M. BOOTH and W.C. DE G R O A T
Department of Pharmacology, School of Medicine, UniversiO"of Pittsburg.h, Pittsburg.h, PA 15261 (U.S.A.) (Accepted November 23rd, 1982)
Kevwords. n a l o x o n e - bladder reflexes
parasympathetic reflexes
enkephalins
Naloxone in doses ranging from 0.5 to 512 ~tg/kg i.v., enhanced reflex contractions of the urinary bladder of the cat. At the lowest doses (threshold, 0.5 5/*g/kg) the drug increased the frequency of spontaneous bladder contractions. In large doses ( 10 100 >g/kg) the drug produced an initial tonic contraction of bladder lasting 15 40 rain followed by a period of high frequency rhythmic activity. Multiunit firing in parasympathetic postganglionic nerves on the surface of the urinary bladder was also enhanced. Bursts of firing which in untreated animals occurred during large bladder contractions occurred continuously during the entire sustained contraction of the bladder following large doses of naloxone. Various evidence indicates that the site of action of naloxone is in the central nervous system, These findings suggest that the parasympathetic reflex pathway to the urinary bladder may be subject to tonic enkephalinergic inhibitory control.
Neurons in the parasympathetic nucleus of the sacral spinal cord innervate the urinary bladder and other organs of the pelvic viscera. Electrophysiological 3.4 and horseradish peroxidase tracing studies 13 in this laboratory have shown that the area of the sacral parasympathetic nucleus (SPN) providing innervation to the urinary bladder of the cat is located in lamina VII at the lateral border of the intermediate gray. This same area has been shown recently 5,9 to contain a dense network of terminal processes and neurons which exhibit leucine enkephalin (L-Enk) immunoreactivity. L-Enk terminals have also been demonstrated in bladder ganglia of the cat? These data coupled with the observation that L-Enk inhibits transmission in bladder ganglia and at other sites in the autonomic nervous system L~L~2, suggest that enkephalins may play an important role in the neural control of bladder function. This possibility was investigated in the present study by examining the effects ofnaloxone, an opiate antagonist, on the activity of urinary bladder. Experiments were performed on 11 cats of either sex anesthetized with chloralose (5(570 mg/kg i.v.). A midline abdominal incision exposed the urinary bladder and its parasympathetic
(pelvic nerves) and sympathetic (hypogastric nerves) innervation. Intravesical pressure was measured using a pressure transducer connected to a saline-filled cannula which was inserted into the bladder lumen through an incision in the urethra. In most experiments the hypogastric nerves were sectioned bilaterally to block the sympathetic inhibitory input to the bladder. In some experiments the pelvic nerves and postganglionic nerve fibers on the surface of the bladder were isolated and prepared for stimulation and recording. In 3 animals a laminectom) was performed at the first lumbar vertebra to permit the induction of spinal anesthesia (lidocaine) during the experiment. In all animals, blood pressure, end-tidal CO2 and body temperature were monitored and maintained at normal levels. Spontaneous bladder contractions recorded under constant volume conditions in this series of experiments consisted of rhythmic contractions occurring at a frequency of 0.5 3 per min, with a duration of l(L30 s and a peak amplitude of 35 60 cm H20. An initial bladder pressure of" 5 10 cm H20 was necessary to trigger the rhythmic contractions which then occurred for many hours with only minimal variation 6.
0006-8993/83/0000~0000/$03.00 ~ 1983 Elsevier Science Publishers
356
250 _ A
B ffl
e~
~
0
O
40
E
1 min
CONTROL
NALOXONE
250 C
® o
[
D
0
......
~4pg/kg
'~8/Jg/kg NALOXONE
- . . . .
_. . . . . . . . . .
£:i "-,._',A'
'~1 6pglkg NALOXONE
Fig. 1. Effects of increasing doses of naloxone on multiunit activity (top trace in each panel) recorded from postganglionic fibers on surface of the bladder and bladder pressure (bottom trace in each panel) recorded through an intraluminal bladder cannula. A: control. B, C, D: increasing accumulated doses of naloxone given i.v. The multiunit activity is a ratemeter output with a time constant of 0.5 s.
An efferent discharge recorded from vesical parasympathetic postganglionic nerve fibers preceded the bladder contractions and continued for the duration of the contraction, subsiding just prior to the decline in bladder pressure (Figs. I A and 2A). The efferent firing was characterized as a sustained period of burst-firing; one burst occurring every 0,3 1.6 s (mean 0.51 s) during the large contractions (Fig. 1). Efferent firing did not occur during the periods between contractions. Naloxone was studied in doses of 0.5-512 ~g/kg in 11 cats. In 5 cats complete cumulative dose-response curves were obtained using doses from 1/~g/kg to 64/~g/kg. The dose ofnaloxone
necessary to produce a detectable change in bladder activity ranged from 0.5 /zg/kg to 2 ~g/kg in different animals. In this low dose range naloxone increased the frequency of the contractions and in some cases also prolonged the contractions (Fig. IB). At doses of2-8/zg/kg bladder contractions occurred at lower amplitude but at a higher frequency (7-8/min) (Fig. 1C). Maximal responses occurring at doses of 8 32 /~g/kg consisted of either high frequency rhythmic activity with a small (5-10 cm H20) shift in baseline intravesical pressure, or a large tonic contraction with a large (20-40 cm H20) shift in pressure, upon which was superimposed high frequency rhythmic activity (Fig. ID. 2B).
357 The latter effect was seen more often when a single large dose of naloxone was given initially rather than with multiple injections of small doses. This difference in response may reflect a form of rapid tolerance that develops to an initial dose of naloxone. This phenomenon has been reported previously "~ with regard to the effect of naloxone on monosynaptic reflexes in the cat. Efferent neural activity was recorded from postganglionic nerve fibers in 4 experiments. Following a single large dose of naloxone (10100 /zg/kg) a continuous series of multiunit bursts were recorded accompanied by a continuous contraction of the bladder (Fig. 2). The bursting activity closely resembled that seen during the peaks of rhythmic bladder contractions. The activity persisted for 15 40 min followed by a period of recovery characterized by the reappearance of intermittent firing and rhythmic bladder contractions. Full recovery usually required 1.5 2 h. Smaller doses of naloxone increased the duration of firing associated with bladder contractions and reduced the quiescent period between contractions (Fig. I B, C). Similar neural activity was noted during recovery after larger doses of the drug.
A
o -!-
CONTROL 40
[.
200jJV
. . . . . . . . . . . . . . . . . . .
i
1
B
In order to show that the enhanced efferent firing and bladder activity following naloxone was dependent on a neuronal pathway in the central nervous system, several types of experiments were performed. In the first study after the effect of naloxone was established, lidocaine was injected into the upper lumbar segments of the spinal cord (two experiments). Within 15 s of the injection the vesical efferent activity was abolished and the bladder pressure decreased from approximately 25 to less than 10 c m H 2 0 . A second group of experiments were performed in order to exclude a possible peripheral site of action of naloxone. Bladder contractions were elicited by electrical stimulation (5 Hz) of efferent axons in the pelvic nerve or in the sacral ventral roots in animals where the spinal cord had been injected with lidocaine or where the sacral ventral roots had been sectioned centrally. Large doses of naloxone up to 1 mg/kg i.v., did not modify the bladder contractions induced by stimulation of the peripheral efferent pathways. We conclude that naloxone enhances the parasympathetic excitatory outflow from the central nervous system to the urinary bladder thereby causing a profound contraction of the bladder smooth muscle. Three pieces of evidence in-
l . A t _~ .Jh j
",
T ~T T 1 T
' _.~,__a
o
0
.
.
.
.
.
.
Illlll
I.
mdl
Ill l
d=
=
.
sec
NALOXONE 4o
.
i
o ~
. . . . . . . . . . . . . . . . . . . . . . . . . . . .
~T ~ T ~ T .
16,ug/kg
200p 1' s e c
Fig. 2. Effects of a single dose of naloxone on bladder pressure (top trace each row) and multiunit activity (bottom trace each row). A and B are film records from the oscilloscope. A: control. B: 3 min after 16/Lg/kg naloxone i.v. The expanded time base shows the burst activity in the multiunit record.
358 dicate that the site of action of naloxone is within the central nervous system: (l) the efferent outflow originating centrally was enhanced by naloxone; (2) naloxone had no effect on bladder contractions induced by stimulation of peripheral parasympathetic nerves to the bladder; and (3) recent work in this laboratory by Simonds et al? 6 showed that naloxone did not modify transmission in parasympathetic ganglia of the bladder. Although the site of naloxone action in the central nervous system was not determined in the present experiments, the most likely site would be in areas which contain endogenous opiate substances and which are part of the micturition reflex pathway, for example the intermediolateral gray of the sacral spinal cord and brainstem micturition center 2-5,9.H.~5. Immunohistochemical studies have shown that the sacral autonomic nucleus of the cat is characterized by
a dense accumulation of enkephalinergic terminals. In addition, recent pharmacological experiments revealed that injections of enkephalins into the sacral subarachnoid space inhibit bladder reflexes, whereas naloxone blocks the inhibition and enhances bladder activity ~, An action of naloxone on the spinal cord was also demonstrated in chronic spinal animals (transection at L 1) where the administration of naloxone intravenously induced micturition and enhanced somatovesical excitatory reflexes ~. These studies suggest that at the level of the spinal cord, the parasympathetic outflow to the urinary bladder is subject to a tonic enkephalinergic inhibitory control, which can in turn be blocked by naloxone.
1 Booth, A. M., Ostrowski, N., McLinden, S., Lowe, 1. and de Groat, W. C., An analysis of the inhibitory effects of leucine enkephalin on transmission in vesical parasympathetic ganglia of the cat, Soc. Neurosci. Abstr., 7 ( 1981 ) 214. 2 de Groat, W. C.. Nervous control of the urinary bladder of the cat, Brain Research, 87 (1975) 201 211. 3 de Groat, W. C., Booth, A. M., Krier, J., Milne, R., Morgan, C. and Nadelhaft, I., Neural control of the urinary bladder and large intestine. In C. McC. Brooks, K. Koizumi and A. Sato (Eds.), Integrative Functions of the A utonomic Nervous System, Tokyo Univ. Press. Tokyo-Elsevier, Amsterdam, 1979, pp. 234 247. 4 de Groat, W. C., Booth, A. M., Milne, R. J. and Roppollo, J. R., Parasympathetic preganglionic neurons in the sacral spinal cord, J. auton. Nerv. Syst., 5 (1982) 23 44. 5 de Groat, W. C., Kawatani, M., Booth, A. M., Lowe, I. P. and Zug, D., Identification of leucine-enkephalin in the sacral parasympathetic outflow to the urinary bladder of the cat, Neuroscience, 7 Suppl. (1982) 550. 6 de Groat, W. C. and Ryall, R. W., Reflexes to sacral parasympathetic neurones concerned with micturition in the cat, J. Physiol. (Lond.), 200 (1969) 87 108. 7 [:aden, A. 1., Jacobs, T. P. and Holaday, J. W., Endorphin parasympathetic interaction in spinal shock. J. auton. Nerv. Svst., 2 (1980) 295 304. 8 Frederickson, R., Enkephalin pentapeptides A review of current evidence for physiological role in vertebrate neurotransmission, Life Sci., 21 (1977) 23- 42. 9 Glazer, E. and Basbaum, A., Leucine enkephalin: localization in and axoplasmic transport by sacral parasympathetic preganglionic neurons. Science. 208 (1980) 1479 1481.
10 Goldfarb, J. and Hu, J. W., Enhancement of reflexes by naloxone in spinal cats, Neuropharmacology, 15 (1976) 785- 792. I1 Hisamitsu, T., Roques, B. P. and de Groat, W.C., The role of enkephalins in the sacral parasympathetic reflex pathways to the urinary bladder of the cat, Soc. NeuroscL Abstr., 8 (1982). 12 Konishi, S., Tsunoo, A. and Otsuka, M.. Enkephalins presynaptically inhibit cholinergic transmission in sympathetic ganglia, Nature (Lond.), 282 (1979) 515 516. 13 Nadelhaft, I., Morgan, C. W. and de Groat, W. C., Localization of the sacral autonomic nucleus in the spinal cord of the cat by the horseradish peroxidase technique, J. comp. Neurol., 193 (1980) 265 ~281. 14 Sar, M., Stump[, W. E., Miller, R.J., Chang, K. and Cuatrecasas, P., Immunohistochemical localization ofenkephalin in rat brain and spinal cord, J. comp. Neurol., 182 (1978) 17 38. 15 Satoh, K., Shimizu, N., l-ohyama, M. and Maeda. T., Localization of the micturition reflex center at dorsolateral pontine tegmentum of the rat, NeuroscL l,ett., 8 (1978) 27 33. 16 Simonds, W. F., Booth, A. M., Thor, K. B.. Ostrowski, N. L., Nagel, J. R. and de Groat. W. C., Parasympathetic ganglia: naloxone antagonizes inhibition by leucine-enkephalin and GABA (unpublished). 17 Thor, K. B., Roppolo, J. R. and de Groat, W. C., Enhancement of vesical and Somatovesical reflexes by naloxone in unanesthetized chronic spinal cats, Pharmacologist. 24 (1982) 122.
Supported by N S F Grant PCM 7900093, NIH Grant NS 18075 and an N I M H CRC Grant MH 30915.
Brain Research, 264 (1983) 355 358 Elsevier Biomedical Press
The effects of naloxone on the neural control of the urinary bladder of the cat J, R. ROPPOLO, A. M. BOOTH and W.C. DE G R O A T
Department of Pharmacology, School of Medicine, UniversiO"of Pittsburg.h, Pittsburg.h, PA 15261 (U.S.A.) (Accepted November 23rd, 1982)
Kevwords. n a l o x o n e - bladder reflexes
parasympathetic reflexes
enkephalins
Naloxone in doses ranging from 0.5 to 512 ~tg/kg i.v., enhanced reflex contractions of the urinary bladder of the cat. At the lowest doses (threshold, 0.5 5/*g/kg) the drug increased the frequency of spontaneous bladder contractions. In large doses ( 10 100 >g/kg) the drug produced an initial tonic contraction of bladder lasting 15 40 rain followed by a period of high frequency rhythmic activity. Multiunit firing in parasympathetic postganglionic nerves on the surface of the urinary bladder was also enhanced. Bursts of firing which in untreated animals occurred during large bladder contractions occurred continuously during the entire sustained contraction of the bladder following large doses of naloxone. Various evidence indicates that the site of action of naloxone is in the central nervous system, These findings suggest that the parasympathetic reflex pathway to the urinary bladder may be subject to tonic enkephalinergic inhibitory control.
Neurons in the parasympathetic nucleus of the sacral spinal cord innervate the urinary bladder and other organs of the pelvic viscera. Electrophysiological 3.4 and horseradish peroxidase tracing studies 13 in this laboratory have shown that the area of the sacral parasympathetic nucleus (SPN) providing innervation to the urinary bladder of the cat is located in lamina VII at the lateral border of the intermediate gray. This same area has been shown recently 5,9 to contain a dense network of terminal processes and neurons which exhibit leucine enkephalin (L-Enk) immunoreactivity. L-Enk terminals have also been demonstrated in bladder ganglia of the cat? These data coupled with the observation that L-Enk inhibits transmission in bladder ganglia and at other sites in the autonomic nervous system L~L~2, suggest that enkephalins may play an important role in the neural control of bladder function. This possibility was investigated in the present study by examining the effects ofnaloxone, an opiate antagonist, on the activity of urinary bladder. Experiments were performed on 11 cats of either sex anesthetized with chloralose (5(570 mg/kg i.v.). A midline abdominal incision exposed the urinary bladder and its parasympathetic
(pelvic nerves) and sympathetic (hypogastric nerves) innervation. Intravesical pressure was measured using a pressure transducer connected to a saline-filled cannula which was inserted into the bladder lumen through an incision in the urethra. In most experiments the hypogastric nerves were sectioned bilaterally to block the sympathetic inhibitory input to the bladder. In some experiments the pelvic nerves and postganglionic nerve fibers on the surface of the bladder were isolated and prepared for stimulation and recording. In 3 animals a laminectom) was performed at the first lumbar vertebra to permit the induction of spinal anesthesia (lidocaine) during the experiment. In all animals, blood pressure, end-tidal CO2 and body temperature were monitored and maintained at normal levels. Spontaneous bladder contractions recorded under constant volume conditions in this series of experiments consisted of rhythmic contractions occurring at a frequency of 0.5 3 per min, with a duration of l(L30 s and a peak amplitude of 35 60 cm H20. An initial bladder pressure of" 5 10 cm H20 was necessary to trigger the rhythmic contractions which then occurred for many hours with only minimal variation 6.
0006-8993/83/0000~0000/$03.00 ~ 1983 Elsevier Science Publishers
356
250 _ A
B ffl
e~
~
0
O
40
E
1 min
CONTROL
NALOXONE
250 C
® o
[
D
0
......
~4pg/kg
'~8/Jg/kg NALOXONE
- . . . .
_. . . . . . . . . .
£:i "-,._',A'
'~1 6pglkg NALOXONE
Fig. 1. Effects of increasing doses of naloxone on multiunit activity (top trace in each panel) recorded from postganglionic fibers on surface of the bladder and bladder pressure (bottom trace in each panel) recorded through an intraluminal bladder cannula. A: control. B, C, D: increasing accumulated doses of naloxone given i.v. The multiunit activity is a ratemeter output with a time constant of 0.5 s.
An efferent discharge recorded from vesical parasympathetic postganglionic nerve fibers preceded the bladder contractions and continued for the duration of the contraction, subsiding just prior to the decline in bladder pressure (Figs. I A and 2A). The efferent firing was characterized as a sustained period of burst-firing; one burst occurring every 0,3 1.6 s (mean 0.51 s) during the large contractions (Fig. 1). Efferent firing did not occur during the periods between contractions. Naloxone was studied in doses of 0.5-512 ~g/kg in 11 cats. In 5 cats complete cumulative dose-response curves were obtained using doses from 1/~g/kg to 64/~g/kg. The dose ofnaloxone
necessary to produce a detectable change in bladder activity ranged from 0.5 /zg/kg to 2 ~g/kg in different animals. In this low dose range naloxone increased the frequency of the contractions and in some cases also prolonged the contractions (Fig. IB). At doses of2-8/zg/kg bladder contractions occurred at lower amplitude but at a higher frequency (7-8/min) (Fig. 1C). Maximal responses occurring at doses of 8 32 /~g/kg consisted of either high frequency rhythmic activity with a small (5-10 cm H20) shift in baseline intravesical pressure, or a large tonic contraction with a large (20-40 cm H20) shift in pressure, upon which was superimposed high frequency rhythmic activity (Fig. ID. 2B).
357 The latter effect was seen more often when a single large dose of naloxone was given initially rather than with multiple injections of small doses. This difference in response may reflect a form of rapid tolerance that develops to an initial dose of naloxone. This phenomenon has been reported previously "~ with regard to the effect of naloxone on monosynaptic reflexes in the cat. Efferent neural activity was recorded from postganglionic nerve fibers in 4 experiments. Following a single large dose of naloxone (10100 /zg/kg) a continuous series of multiunit bursts were recorded accompanied by a continuous contraction of the bladder (Fig. 2). The bursting activity closely resembled that seen during the peaks of rhythmic bladder contractions. The activity persisted for 15 40 min followed by a period of recovery characterized by the reappearance of intermittent firing and rhythmic bladder contractions. Full recovery usually required 1.5 2 h. Smaller doses of naloxone increased the duration of firing associated with bladder contractions and reduced the quiescent period between contractions (Fig. I B, C). Similar neural activity was noted during recovery after larger doses of the drug.
A
o -!-
CONTROL 40
[.
200jJV
. . . . . . . . . . . . . . . . . . .
i
1
B
In order to show that the enhanced efferent firing and bladder activity following naloxone was dependent on a neuronal pathway in the central nervous system, several types of experiments were performed. In the first study after the effect of naloxone was established, lidocaine was injected into the upper lumbar segments of the spinal cord (two experiments). Within 15 s of the injection the vesical efferent activity was abolished and the bladder pressure decreased from approximately 25 to less than 10 c m H 2 0 . A second group of experiments were performed in order to exclude a possible peripheral site of action of naloxone. Bladder contractions were elicited by electrical stimulation (5 Hz) of efferent axons in the pelvic nerve or in the sacral ventral roots in animals where the spinal cord had been injected with lidocaine or where the sacral ventral roots had been sectioned centrally. Large doses of naloxone up to 1 mg/kg i.v., did not modify the bladder contractions induced by stimulation of the peripheral efferent pathways. We conclude that naloxone enhances the parasympathetic excitatory outflow from the central nervous system to the urinary bladder thereby causing a profound contraction of the bladder smooth muscle. Three pieces of evidence in-
l . A t _~ .Jh j
",
T ~T T 1 T
' _.~,__a
o
0
.
.
.
.
.
.
Illlll
I.
mdl
Ill l
d=
=
.
sec
NALOXONE 4o
.
i
o ~
. . . . . . . . . . . . . . . . . . . . . . . . . . . .
~T ~ T ~ T .
16,ug/kg
200p 1' s e c
Fig. 2. Effects of a single dose of naloxone on bladder pressure (top trace each row) and multiunit activity (bottom trace each row). A and B are film records from the oscilloscope. A: control. B: 3 min after 16/Lg/kg naloxone i.v. The expanded time base shows the burst activity in the multiunit record.
358 dicate that the site of action of naloxone is within the central nervous system: (l) the efferent outflow originating centrally was enhanced by naloxone; (2) naloxone had no effect on bladder contractions induced by stimulation of peripheral parasympathetic nerves to the bladder; and (3) recent work in this laboratory by Simonds et al? 6 showed that naloxone did not modify transmission in parasympathetic ganglia of the bladder. Although the site of naloxone action in the central nervous system was not determined in the present experiments, the most likely site would be in areas which contain endogenous opiate substances and which are part of the micturition reflex pathway, for example the intermediolateral gray of the sacral spinal cord and brainstem micturition center 2-5,9.H.~5. Immunohistochemical studies have shown that the sacral autonomic nucleus of the cat is characterized by
a dense accumulation of enkephalinergic terminals. In addition, recent pharmacological experiments revealed that injections of enkephalins into the sacral subarachnoid space inhibit bladder reflexes, whereas naloxone blocks the inhibition and enhances bladder activity ~, An action of naloxone on the spinal cord was also demonstrated in chronic spinal animals (transection at L 1) where the administration of naloxone intravenously induced micturition and enhanced somatovesical excitatory reflexes ~. These studies suggest that at the level of the spinal cord, the parasympathetic outflow to the urinary bladder is subject to a tonic enkephalinergic inhibitory control, which can in turn be blocked by naloxone.
1 Booth, A. M., Ostrowski, N., McLinden, S., Lowe, 1. and de Groat, W. C., An analysis of the inhibitory effects of leucine enkephalin on transmission in vesical parasympathetic ganglia of the cat, Soc. Neurosci. Abstr., 7 ( 1981 ) 214. 2 de Groat, W. C.. Nervous control of the urinary bladder of the cat, Brain Research, 87 (1975) 201 211. 3 de Groat, W. C., Booth, A. M., Krier, J., Milne, R., Morgan, C. and Nadelhaft, I., Neural control of the urinary bladder and large intestine. In C. McC. Brooks, K. Koizumi and A. Sato (Eds.), Integrative Functions of the A utonomic Nervous System, Tokyo Univ. Press. Tokyo-Elsevier, Amsterdam, 1979, pp. 234 247. 4 de Groat, W. C., Booth, A. M., Milne, R. J. and Roppollo, J. R., Parasympathetic preganglionic neurons in the sacral spinal cord, J. auton. Nerv. Syst., 5 (1982) 23 44. 5 de Groat, W. C., Kawatani, M., Booth, A. M., Lowe, I. P. and Zug, D., Identification of leucine-enkephalin in the sacral parasympathetic outflow to the urinary bladder of the cat, Neuroscience, 7 Suppl. (1982) 550. 6 de Groat, W. C. and Ryall, R. W., Reflexes to sacral parasympathetic neurones concerned with micturition in the cat, J. Physiol. (Lond.), 200 (1969) 87 108. 7 [:aden, A. 1., Jacobs, T. P. and Holaday, J. W., Endorphin parasympathetic interaction in spinal shock. J. auton. Nerv. Svst., 2 (1980) 295 304. 8 Frederickson, R., Enkephalin pentapeptides A review of current evidence for physiological role in vertebrate neurotransmission, Life Sci., 21 (1977) 23- 42. 9 Glazer, E. and Basbaum, A., Leucine enkephalin: localization in and axoplasmic transport by sacral parasympathetic preganglionic neurons. Science. 208 (1980) 1479 1481.
10 Goldfarb, J. and Hu, J. W., Enhancement of reflexes by naloxone in spinal cats, Neuropharmacology, 15 (1976) 785- 792. I1 Hisamitsu, T., Roques, B. P. and de Groat, W.C., The role of enkephalins in the sacral parasympathetic reflex pathways to the urinary bladder of the cat, Soc. NeuroscL Abstr., 8 (1982). 12 Konishi, S., Tsunoo, A. and Otsuka, M.. Enkephalins presynaptically inhibit cholinergic transmission in sympathetic ganglia, Nature (Lond.), 282 (1979) 515 516. 13 Nadelhaft, I., Morgan, C. W. and de Groat, W. C., Localization of the sacral autonomic nucleus in the spinal cord of the cat by the horseradish peroxidase technique, J. comp. Neurol., 193 (1980) 265 ~281. 14 Sar, M., Stump[, W. E., Miller, R.J., Chang, K. and Cuatrecasas, P., Immunohistochemical localization ofenkephalin in rat brain and spinal cord, J. comp. Neurol., 182 (1978) 17 38. 15 Satoh, K., Shimizu, N., l-ohyama, M. and Maeda. T., Localization of the micturition reflex center at dorsolateral pontine tegmentum of the rat, NeuroscL l,ett., 8 (1978) 27 33. 16 Simonds, W. F., Booth, A. M., Thor, K. B.. Ostrowski, N. L., Nagel, J. R. and de Groat. W. C., Parasympathetic ganglia: naloxone antagonizes inhibition by leucine-enkephalin and GABA (unpublished). 17 Thor, K. B., Roppolo, J. R. and de Groat, W. C., Enhancement of vesical and Somatovesical reflexes by naloxone in unanesthetized chronic spinal cats, Pharmacologist. 24 (1982) 122.
Supported by N S F Grant PCM 7900093, NIH Grant NS 18075 and an N I M H CRC Grant MH 30915.