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New long latency bulbospinal evoked potentials blocked by serotonin antagonists
542
Brain Research, 65 (1974) 542-546 © Elsevier Scientific Publishing Company, Amsterdam - Printed in The Netherlands
New long latency bulbospinal evoked po,tentials blocked by serotonin antagonists H E R B E R T K. P R O U D F I T AND E D M U N D
G. A N D E R S O N
University of Illinois Department of Pharmacology, College of Medicbw, Chicago, Ill. 60680 (U.S.A.)
(Accepted October 4th, 1973)
Histochemicala,s, 9 and biochemical z investigations have demonstrated a system of serotonin (5-HT)-containing cells whose somata lie in the caudal brain stem raphe nuclei and whose axons project to the spinal cord where they terminate in both the dorsal and ventral horns. The major source of descending 5-HT axons are the B1, B2, and B3 cells described by Dahlstr6m and Fuxe 8 which are found in the nucleus raphe obscurus, nucleus raphe pallidus and nucleus raphe magnus, respectively. Several attempts have been made to activate these descending serotonergic cells and record their effects on spinal reflexes6,7,12,13. These studies failed to evoke responses which could be attributed to activation of the bulbospinal serotonergic neurons, and those effects which were susceptible to 5-HT antagonists now appear to be explicable in terms of release from the tonic effects of the bulbospinal serotonergic pathway 13. In a more recent communication, Barasi and Roberts 2 reported a long latency activation of motoneurons in rats following raphe stimulation. However, their stereotaxic coordinates place their stimulating electrodes in the B8 group of cells of the nucleus raphe medianus 8 whose primary projection is to the telencephalon via the medial forebrain bundle 1. The present report describes a series of experiments in which stimulation of the more caudal nucleus raphe magnus of the brain stem evoked potentials that could be recorded from both dorsal and ventral roots of the lumbar spinal cord. The B3 cells of the nucleus raphe magnus have been shown by fluorescence histochemical techniques to project axons to the lumbar segments of the spinal cord4, 8. Components of these evoked potentials exhibited properties expected from activation of the descending serotonergic system. These components demonstrated long latencies consistent with the slow conduction velocity of the small diameter serotonergic neurons, and they were blocked by 5-HT antagonists. Unanesthetized decerebrate cats were used. The medial part of the cerebellum, with the associated cerebellar nuclei, was removed. Concentric bipolar stainless steel stimulating electrodes (Kopf, model NE-100) were used to stimulate the brain stem. The inner shaft diameter was 0.2 mm with 0.5 mm of tip separation. The stimulating electrode was visually oriented on the midline 2-7 mm rostral to the obex and lowered 2-5 mm from the surface into the brain stem raphe.
SHORT
543
COMMUNICATIONS
DORSAL ROOT POTENTIAL 120- b- •%'0
PRE-DRUG CONTROL CINANSERIN 4mg/kg
,oo
< ~-
I rain.
: ;
~
r
0_ 8 0 e
; :
o
"
• 60
3 min.
Z
I
o
a
a
I..Z
bJ 40
DRP-I • DRP-2• ....
,
I
n~ Ld O-
5 min.
•
p.•
\
•
" i
20
•
a e
',0
15 min.
2 0 0 p.V
,~
I'0 1'5 T I M E (rain)
2'0
i.J
I O0 msec
Cinonserin 4 mg/kg
Fig. 1. Effect of cinanserin (4 mg/kg) on the short and long latency dorsal root potentials (DRP-I and DRP-2) evoked from the caudal brain stem. Dorsal root potentials evoked by brain stem stimulation are shown on the left and consist of two negative-going waves; one having a short latency (22 msec) and the other having a longer latency (95 msec). The evoked potentials are represented by 10 superimposed oscilloscope tracings. The control potentials are shown in the top panel and those after cinanserin below. The graph on the right shows the increase in the DRP-1 and the decrease in DRP-2 after cinanserin expressed as a percent of the area under the control DRP. After a laminectomy, the most caudal L7 dorsal rootlet and the L7 ventral root were mounted on platinum recording electrodes, while the remaining ipsilateral L7 dorsal root was mounted on platinum stimulating electrodes. A constant temperature oil pool bathed the exposed spinal cord, and spontaneous muscle movement was controlled by the injection of gallamine triethiodide (Flaxedil). All drugs were administered intravenously. Stimulation of the nucleus raphe magnus using 30 msec trains of 0.5 msec, 3 V pulses (0.08-0.25 mA) evoked potentials in both the dorsal and ventral roots o f the lumbar cord consisting, in both cases, of two negative waves. The potentials recorded from the dorsal roots were designated DRP-1 and DRP-2 (Fig. 1), and those recorded from the ventral root as VRP-1 and VRP-2 (Fig. 2). As the stimulating electrodes were lowered into the brain stem only the DRP-1 and VRP-1 were observed, though VRP-1 did not appear with all electrode locations. With further penetration the second negative waves began to appear (DRP-2 and VRP-2). The DRP-2 and VRP-2 reached an optimum size at an electrode depth of between 2 and 5 mm from the surface, while DRP-1 and VRP-1 were minimally influenced by the
544
SHORT COMMUNICATIONS
electrode position. The movement of the stimulating electrodes by as little as 0.51 mm resulted in a disappearance of DRP-2 and VRP-2. This suggests these potentials originate from discrete areas in the raphe nuclei. In contrast the DRP-1 and VRP-1 could be evoked from wide areas of the reticular formation. D R P - I had an onset latency of about 22 msec, and DRP-2 a latency of about 95 msec. VRP-1 and VRP-2 had onset latencies of 5-10 and 50-60 msec, respectively. The long latency of DRP-2 and VRP-2 suggests they are mediated by slow conducting fibers. Dahlstr6m and Fuxe 9 report that the serotonin-containing fibers descending from the brain stem raphe make 'intimate contact' with large motoneurons in the ventral horn. By assuming that the system giving rise to DRP-2 and VRP-2 has a minimal number of synapses, the conduction velocity for the pathways which generate VRP-2 and DRP-2 were calculated to be approximately 3.5 and 2.5 m/see respectively. Because of the long latency o f DRP-2 and VRP-2, more than a dozen synapses would have to be interposed in these pathways to increase these estimates of conduction velocity by
VENTRAL ROOT POTENTIAL PRE-DRUG CONTROL
I00"
CINANSERIN~:~> 4mg/kg
90-
I min.
80-
n.-
5min.
70-
G. ~, 6 0 -
_J 0 I.- 5 0 Z O
lOmin.
15rain. 30min.
~
40"
~
30-
/
20-
! mV[ I00 r n s e c
ib 2'o 3'o ~ TIME ,rain, Cinonserin 4 rng/kg
Fig. 2. Depression by cinanserin (4 mg/kg) of the long latency ventral root potential (VRP-2) evoked by brain stem stimulation. The superimposed oscilloscope tracings on the left show only VRP-2. In this experiment the stimulating electrode was situated 1.5 mm lateral to the midline in a group of 5-HT-containing cell bodies located outside the nucleus raphe magnus where no short latency VRP-1 could be elicited. The top panel shows the control VRP-2 and those below the reduction in VRP-2 area after cinanserin. The graph on the right shows the reduction in VRP-2 area expressed as a percent of the control VRP after cinanserin.
SHORT COMMUNICATIONS
545
10~. This is consistent with the conduction velocity expected in descending 5-HT fibers of 1-2/~m in diameter as described by Dahlstrfm and Fuxe 9. The 5-HT antagonists cinanserin and methysergide were given to evaluate the possibility that the DRP-2 and VRP-2 were serotonergically mediated. Within 1 min after the intravenous injection of 4 mg/kg of cinanserin into 5 cats, DRP-2 was either abolished or greatly reduced while DRP-1 was increased. In the sample experiment illustrated in Fig. 1, the DRP-2 declined to 33 ~ of control value within 1 rain after i.v. injection of cinanserin and was maximally depressed to 16~ of control within 5 rain. Recovery was observed after about 20 rain. The administration of 1 mg/kg of methysergide, another potent 5-HT antagonist, to 4 cats produced results identical to those obtained using cinanserin. The possibility that descending adrenergic systems were involved in generating the DRP-2 is unlikely since administration of the alpha adrenergic antagonist phenoxybenzamine (5 mg/kg) was without effect on DRP-2. The increase in DRP-1 following the administration of the 5-HT antagonists has been previously documented and discussed lz. The long latency ventral root potential (VRP-2) was also either abolished or greatly reduced after cinanserin (Fig. 2) or methysergide. In the preparation illustrated no VRP-1 was observed. However, when a VRP-1 was present no change in its size was observed. The time course of the 5-HT antagonist's action on the long latency ventral root potential was similar to that of the long latency dorsal root potential. The long latency potentials (DRP-2, VRP-2) were evoked primarily from an area of the caudal brain stem identified histologically as the caudal nucleus raphe magnus. However, histological examination of our electrode placements revealed that these potentials could also be evoked from a few sites outside this nucleus. It was observed that these placements were located in areas identified by Pin et al. 11 as having 5-HT-containing cell bodies. The size of the fibers carrying the DRP-1 and DRP-2 was estimated by determining the length constants for these potentials. These measurements were made by moving the recording electrodes in 1 mm increments (with a constant interelectrode distance) toward the distal end of the rootlet and determining the distance required to reduce the DRP to 1/e of its initial value. The length constant of the fibers carrying the segmentally evoked DRP was determined for comparison with the brain stemevoked DRP-1 and DRP-2. In 3 experiments, the segmental DRP evoked at Group I threshold exhibited an average length constant of 6.0 mm, the average DRP-2 length constant was 5.8 mm and the average DRP-1 length constant was 3.5 ram. Since the DRP-2 length constant was essentially the same as that of the segmental DRP, it may be assumed that these two DRPs are carried in fibers of about the same diameter, that is Group I fibers. The smaller length constant of DRP-1 indicates that it is generated in smaller fibers, probably Groups II and III. We have found that in decerebellate cats stimulation of areas in the caudal brain stem having 5-HT-containing cell bodies evokes complex potentials in both the dorsal and ventral roots of the lumbar cord. The first components of these potentials are well knownS,10,12,13. However, the second c o m p o n e n t s (DRP-2 and VRP2) have not been reported by other workers. The long latency of these components is
546
SHORT COMMUNICATIONS
consistent with the slow-conduction velocity expected from the small diameter serotonergic axons which descend from the caudal raphe. The blockade ot these longlatency potentials by the administration of two chemically different 5-HT antagonists, cinanserin and methysergide, provides evidence that these potentials are mediated by 5-HT. As a tentative explanation of our observations, we postulate a descending serotonergic system composed of two basic divisions: one affecting motoneurons and the other contacting interneurons along pathways which ultimately terminate presynaptically on Group I primary afferent terminals. It is not known whether the serotonin pathway affecting motoneurons terminates directly on the motoneuron or on an intercalated neuron. The data suggest that the descending serotonergic system presynaptically inhibits impulses arriving from the periphery while increasing motoneuronal excitability. Thus, the activation of this system would inhibit peripheral input while increasing motoneuronal responsiveness to central activation. As a result, this system would function to bias motoneuronal control in favor of central over segmental input. This study was supported by NIH Grant NS05611.
1 ANDI~N, N.-E., DAHLSTROM,A., FUXE, K., LARSSON,K., OLSON L., AND UNGERSTEDT,U., Ascending monoamine neurons to the telencephalon and diencephalon, Acta physiol, scand., 67 (1966) 313-326. 2 BARASI, S., AND ROBERTS, M. S . T., The action of 5-hydroxytryptamine antagonists and precursors on bulbospinal facilitation of spinal reflexes, Brain Research, 52 (1973) 385-388. 3 CARLSSON, A., MAGNUSSON, T., AND ROSENGREN,E., 5-Hydroxytryptamine in the spinal cord normally and after transection, Experientia (Basel), 19 (1963) 359. 4 CARLSSON,A., FALCK, B., FUXE, K., AND H1LLARP, N.-A., Cellular localization of monoamines in the spinal cord, .4cta physiol, scand., 60 (1964) 112-119. 5 CARPENTER,D, ENGBERG, I., AND LUNDBERG, A., Primary afferent depolarization evoked from the brain stem and the cerebellum, Arch. ital. Biol., 104 (1966) 50-72. 6 CLINESCHMIDT,B. V., AND ANDERSON,E. G., Lysergic acid diethylamide: Antagonism of supraspinal inhibition of spinal reflexes, Brain Research, 16 (1969) 296-300. 7 CLINESCHMIDT,B. V., AND ANDERSON, E. G., The blockade of bulbospinal inhibition by 5hydroxytryptamine antagonists, Exp. Brain Res., 11 (1970) 175-186. 8 DAHLSTROM,A., AND FUXE, K., Evidence for the existence of monoamine-containing neurons in the central nervous system. I. Demonstration of monoamines in the cell bodies of brain stem neurons, Actaphysiol. scand., 62, Suppl. 232 (1964) 1-55. 9 DAHLSTROM,k., AND FUXE, K., Evidence for the existence of monoamine neurons in the central nervous system. II. Experimentally induced changes in the intraneuronal amine levels of bulbospinal neuron systems, Acta physiol, scand., 64, Suppl. 247 (1965) 1-36. 10 LUNDBERG,A., AND VYKLICKY, L., Inhibition of transmission to primary afferents by electrical stimulation of the brain stem, Arch. itaL Biol., 104 (1966) 86-97. 11 PiN, C., JONES,B., ET JOUVET,M., Topographie des neurones monoaminergiques du tronc crrrbral du chat: 6tude par histofluorescence, C. R. Soc. Biol. (Paris), 162 (1968) 2136-2141. 12 PROUDFIT, H. K., AND ANDERSON, E. G., Alteration by serotonin antagonists of the effects of bulbospinal stimulation of spinal reflex pathways, Fed. Proc., 31 (1972) 318. 13 PROUDFIT,H. K., AND ANDERSON, E. G., Influence of serotonin antagonists on bulbospinal systems, Brain Research, 61 (1973) 331-341.
Brain Research, 65 (1974) 542-546 © Elsevier Scientific Publishing Company, Amsterdam - Printed in The Netherlands
New long latency bulbospinal evoked po,tentials blocked by serotonin antagonists H E R B E R T K. P R O U D F I T AND E D M U N D
G. A N D E R S O N
University of Illinois Department of Pharmacology, College of Medicbw, Chicago, Ill. 60680 (U.S.A.)
(Accepted October 4th, 1973)
Histochemicala,s, 9 and biochemical z investigations have demonstrated a system of serotonin (5-HT)-containing cells whose somata lie in the caudal brain stem raphe nuclei and whose axons project to the spinal cord where they terminate in both the dorsal and ventral horns. The major source of descending 5-HT axons are the B1, B2, and B3 cells described by Dahlstr6m and Fuxe 8 which are found in the nucleus raphe obscurus, nucleus raphe pallidus and nucleus raphe magnus, respectively. Several attempts have been made to activate these descending serotonergic cells and record their effects on spinal reflexes6,7,12,13. These studies failed to evoke responses which could be attributed to activation of the bulbospinal serotonergic neurons, and those effects which were susceptible to 5-HT antagonists now appear to be explicable in terms of release from the tonic effects of the bulbospinal serotonergic pathway 13. In a more recent communication, Barasi and Roberts 2 reported a long latency activation of motoneurons in rats following raphe stimulation. However, their stereotaxic coordinates place their stimulating electrodes in the B8 group of cells of the nucleus raphe medianus 8 whose primary projection is to the telencephalon via the medial forebrain bundle 1. The present report describes a series of experiments in which stimulation of the more caudal nucleus raphe magnus of the brain stem evoked potentials that could be recorded from both dorsal and ventral roots of the lumbar spinal cord. The B3 cells of the nucleus raphe magnus have been shown by fluorescence histochemical techniques to project axons to the lumbar segments of the spinal cord4, 8. Components of these evoked potentials exhibited properties expected from activation of the descending serotonergic system. These components demonstrated long latencies consistent with the slow conduction velocity of the small diameter serotonergic neurons, and they were blocked by 5-HT antagonists. Unanesthetized decerebrate cats were used. The medial part of the cerebellum, with the associated cerebellar nuclei, was removed. Concentric bipolar stainless steel stimulating electrodes (Kopf, model NE-100) were used to stimulate the brain stem. The inner shaft diameter was 0.2 mm with 0.5 mm of tip separation. The stimulating electrode was visually oriented on the midline 2-7 mm rostral to the obex and lowered 2-5 mm from the surface into the brain stem raphe.
SHORT
543
COMMUNICATIONS
DORSAL ROOT POTENTIAL 120- b- •%'0
PRE-DRUG CONTROL CINANSERIN 4mg/kg
,oo
< ~-
I rain.
: ;
~
r
0_ 8 0 e
; :
o
"
• 60
3 min.
Z
I
o
a
a
I..Z
bJ 40
DRP-I • DRP-2• ....
,
I
n~ Ld O-
5 min.
•
p.•
\
•
" i
20
•
a e
',0
15 min.
2 0 0 p.V
,~
I'0 1'5 T I M E (rain)
2'0
i.J
I O0 msec
Cinonserin 4 mg/kg
Fig. 1. Effect of cinanserin (4 mg/kg) on the short and long latency dorsal root potentials (DRP-I and DRP-2) evoked from the caudal brain stem. Dorsal root potentials evoked by brain stem stimulation are shown on the left and consist of two negative-going waves; one having a short latency (22 msec) and the other having a longer latency (95 msec). The evoked potentials are represented by 10 superimposed oscilloscope tracings. The control potentials are shown in the top panel and those after cinanserin below. The graph on the right shows the increase in the DRP-1 and the decrease in DRP-2 after cinanserin expressed as a percent of the area under the control DRP. After a laminectomy, the most caudal L7 dorsal rootlet and the L7 ventral root were mounted on platinum recording electrodes, while the remaining ipsilateral L7 dorsal root was mounted on platinum stimulating electrodes. A constant temperature oil pool bathed the exposed spinal cord, and spontaneous muscle movement was controlled by the injection of gallamine triethiodide (Flaxedil). All drugs were administered intravenously. Stimulation of the nucleus raphe magnus using 30 msec trains of 0.5 msec, 3 V pulses (0.08-0.25 mA) evoked potentials in both the dorsal and ventral roots o f the lumbar cord consisting, in both cases, of two negative waves. The potentials recorded from the dorsal roots were designated DRP-1 and DRP-2 (Fig. 1), and those recorded from the ventral root as VRP-1 and VRP-2 (Fig. 2). As the stimulating electrodes were lowered into the brain stem only the DRP-1 and VRP-1 were observed, though VRP-1 did not appear with all electrode locations. With further penetration the second negative waves began to appear (DRP-2 and VRP-2). The DRP-2 and VRP-2 reached an optimum size at an electrode depth of between 2 and 5 mm from the surface, while DRP-1 and VRP-1 were minimally influenced by the
544
SHORT COMMUNICATIONS
electrode position. The movement of the stimulating electrodes by as little as 0.51 mm resulted in a disappearance of DRP-2 and VRP-2. This suggests these potentials originate from discrete areas in the raphe nuclei. In contrast the DRP-1 and VRP-1 could be evoked from wide areas of the reticular formation. D R P - I had an onset latency of about 22 msec, and DRP-2 a latency of about 95 msec. VRP-1 and VRP-2 had onset latencies of 5-10 and 50-60 msec, respectively. The long latency of DRP-2 and VRP-2 suggests they are mediated by slow conducting fibers. Dahlstr6m and Fuxe 9 report that the serotonin-containing fibers descending from the brain stem raphe make 'intimate contact' with large motoneurons in the ventral horn. By assuming that the system giving rise to DRP-2 and VRP-2 has a minimal number of synapses, the conduction velocity for the pathways which generate VRP-2 and DRP-2 were calculated to be approximately 3.5 and 2.5 m/see respectively. Because of the long latency o f DRP-2 and VRP-2, more than a dozen synapses would have to be interposed in these pathways to increase these estimates of conduction velocity by
VENTRAL ROOT POTENTIAL PRE-DRUG CONTROL
I00"
CINANSERIN~:~> 4mg/kg
90-
I min.
80-
n.-
5min.
70-
G. ~, 6 0 -
_J 0 I.- 5 0 Z O
lOmin.
15rain. 30min.
~
40"
~
30-
/
20-
! mV[ I00 r n s e c
ib 2'o 3'o ~ TIME ,rain, Cinonserin 4 rng/kg
Fig. 2. Depression by cinanserin (4 mg/kg) of the long latency ventral root potential (VRP-2) evoked by brain stem stimulation. The superimposed oscilloscope tracings on the left show only VRP-2. In this experiment the stimulating electrode was situated 1.5 mm lateral to the midline in a group of 5-HT-containing cell bodies located outside the nucleus raphe magnus where no short latency VRP-1 could be elicited. The top panel shows the control VRP-2 and those below the reduction in VRP-2 area after cinanserin. The graph on the right shows the reduction in VRP-2 area expressed as a percent of the control VRP after cinanserin.
SHORT COMMUNICATIONS
545
10~. This is consistent with the conduction velocity expected in descending 5-HT fibers of 1-2/~m in diameter as described by Dahlstrfm and Fuxe 9. The 5-HT antagonists cinanserin and methysergide were given to evaluate the possibility that the DRP-2 and VRP-2 were serotonergically mediated. Within 1 min after the intravenous injection of 4 mg/kg of cinanserin into 5 cats, DRP-2 was either abolished or greatly reduced while DRP-1 was increased. In the sample experiment illustrated in Fig. 1, the DRP-2 declined to 33 ~ of control value within 1 rain after i.v. injection of cinanserin and was maximally depressed to 16~ of control within 5 rain. Recovery was observed after about 20 rain. The administration of 1 mg/kg of methysergide, another potent 5-HT antagonist, to 4 cats produced results identical to those obtained using cinanserin. The possibility that descending adrenergic systems were involved in generating the DRP-2 is unlikely since administration of the alpha adrenergic antagonist phenoxybenzamine (5 mg/kg) was without effect on DRP-2. The increase in DRP-1 following the administration of the 5-HT antagonists has been previously documented and discussed lz. The long latency ventral root potential (VRP-2) was also either abolished or greatly reduced after cinanserin (Fig. 2) or methysergide. In the preparation illustrated no VRP-1 was observed. However, when a VRP-1 was present no change in its size was observed. The time course of the 5-HT antagonist's action on the long latency ventral root potential was similar to that of the long latency dorsal root potential. The long latency potentials (DRP-2, VRP-2) were evoked primarily from an area of the caudal brain stem identified histologically as the caudal nucleus raphe magnus. However, histological examination of our electrode placements revealed that these potentials could also be evoked from a few sites outside this nucleus. It was observed that these placements were located in areas identified by Pin et al. 11 as having 5-HT-containing cell bodies. The size of the fibers carrying the DRP-1 and DRP-2 was estimated by determining the length constants for these potentials. These measurements were made by moving the recording electrodes in 1 mm increments (with a constant interelectrode distance) toward the distal end of the rootlet and determining the distance required to reduce the DRP to 1/e of its initial value. The length constant of the fibers carrying the segmentally evoked DRP was determined for comparison with the brain stemevoked DRP-1 and DRP-2. In 3 experiments, the segmental DRP evoked at Group I threshold exhibited an average length constant of 6.0 mm, the average DRP-2 length constant was 5.8 mm and the average DRP-1 length constant was 3.5 ram. Since the DRP-2 length constant was essentially the same as that of the segmental DRP, it may be assumed that these two DRPs are carried in fibers of about the same diameter, that is Group I fibers. The smaller length constant of DRP-1 indicates that it is generated in smaller fibers, probably Groups II and III. We have found that in decerebellate cats stimulation of areas in the caudal brain stem having 5-HT-containing cell bodies evokes complex potentials in both the dorsal and ventral roots of the lumbar cord. The first components of these potentials are well knownS,10,12,13. However, the second c o m p o n e n t s (DRP-2 and VRP2) have not been reported by other workers. The long latency of these components is
546
SHORT COMMUNICATIONS
consistent with the slow-conduction velocity expected from the small diameter serotonergic axons which descend from the caudal raphe. The blockade ot these longlatency potentials by the administration of two chemically different 5-HT antagonists, cinanserin and methysergide, provides evidence that these potentials are mediated by 5-HT. As a tentative explanation of our observations, we postulate a descending serotonergic system composed of two basic divisions: one affecting motoneurons and the other contacting interneurons along pathways which ultimately terminate presynaptically on Group I primary afferent terminals. It is not known whether the serotonin pathway affecting motoneurons terminates directly on the motoneuron or on an intercalated neuron. The data suggest that the descending serotonergic system presynaptically inhibits impulses arriving from the periphery while increasing motoneuronal excitability. Thus, the activation of this system would inhibit peripheral input while increasing motoneuronal responsiveness to central activation. As a result, this system would function to bias motoneuronal control in favor of central over segmental input. This study was supported by NIH Grant NS05611.
1 ANDI~N, N.-E., DAHLSTROM,A., FUXE, K., LARSSON,K., OLSON L., AND UNGERSTEDT,U., Ascending monoamine neurons to the telencephalon and diencephalon, Acta physiol, scand., 67 (1966) 313-326. 2 BARASI, S., AND ROBERTS, M. S . T., The action of 5-hydroxytryptamine antagonists and precursors on bulbospinal facilitation of spinal reflexes, Brain Research, 52 (1973) 385-388. 3 CARLSSON, A., MAGNUSSON, T., AND ROSENGREN,E., 5-Hydroxytryptamine in the spinal cord normally and after transection, Experientia (Basel), 19 (1963) 359. 4 CARLSSON,A., FALCK, B., FUXE, K., AND H1LLARP, N.-A., Cellular localization of monoamines in the spinal cord, .4cta physiol, scand., 60 (1964) 112-119. 5 CARPENTER,D, ENGBERG, I., AND LUNDBERG, A., Primary afferent depolarization evoked from the brain stem and the cerebellum, Arch. ital. Biol., 104 (1966) 50-72. 6 CLINESCHMIDT,B. V., AND ANDERSON,E. G., Lysergic acid diethylamide: Antagonism of supraspinal inhibition of spinal reflexes, Brain Research, 16 (1969) 296-300. 7 CLINESCHMIDT,B. V., AND ANDERSON, E. G., The blockade of bulbospinal inhibition by 5hydroxytryptamine antagonists, Exp. Brain Res., 11 (1970) 175-186. 8 DAHLSTROM,A., AND FUXE, K., Evidence for the existence of monoamine-containing neurons in the central nervous system. I. Demonstration of monoamines in the cell bodies of brain stem neurons, Actaphysiol. scand., 62, Suppl. 232 (1964) 1-55. 9 DAHLSTROM,k., AND FUXE, K., Evidence for the existence of monoamine neurons in the central nervous system. II. Experimentally induced changes in the intraneuronal amine levels of bulbospinal neuron systems, Acta physiol, scand., 64, Suppl. 247 (1965) 1-36. 10 LUNDBERG,A., AND VYKLICKY, L., Inhibition of transmission to primary afferents by electrical stimulation of the brain stem, Arch. itaL Biol., 104 (1966) 86-97. 11 PiN, C., JONES,B., ET JOUVET,M., Topographie des neurones monoaminergiques du tronc crrrbral du chat: 6tude par histofluorescence, C. R. Soc. Biol. (Paris), 162 (1968) 2136-2141. 12 PROUDFIT, H. K., AND ANDERSON, E. G., Alteration by serotonin antagonists of the effects of bulbospinal stimulation of spinal reflex pathways, Fed. Proc., 31 (1972) 318. 13 PROUDFIT,H. K., AND ANDERSON, E. G., Influence of serotonin antagonists on bulbospinal systems, Brain Research, 61 (1973) 331-341.