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Substance P effects on the dorsal motor nucleus of the vagus
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Brain ResearchBulletin, Vol. 23, PP. 149-153. Q Pergamon Press pk. 1989. Printed in the U.S.A
BRIEF COMMUNICATION
Substance P Effects on the Dorsal Motor Nucleus of the Vagus CARLOS
R. PLATA-SALAMAN,’
ATSUO FUKUDA,* TAKETSUGU MINAMI”
AND YUTAKA
OOMURA*
Department ofP~y~h~~~gyand Institute furNeuroscience, Universes of Delaware, Newark, DE 19716 and *Department of Physiology, Faculty of Medicine, Kyushu University, Fukuoka 812, Japan Received 15 March 1989 PLATA-SALAMAN, C. R., A. PLJKUDA, T. MINAh41AND Y. OOMURA. Substance P eflects on the dorsal motor nucleus of the vague. BRAIN RES BULL 23(1/Z) 149-153, 1989.~-Substance P (SP) was applied by superfusion (0.8 to 3.4 X 10e6 M) to neurons of the dorsal motor nucleus of the vagus {DMV) in slice preparations of the rat medulla oblongata. In~acelluI~ recordings showed that 18 of 43 (42%) neurons were depolarized and the depolarization was associated with an increase in membrane input resistance; five of 43 (12%) neurons were hyperpolarized and tire hyperpolarization was associated with a decrease in membrane input resistance. Both effects were reversible and persisted after blockade of synaptic transmission by Ca*’ free/high Mg*+ solution. These data show that: 1) vagal neurons in the DMV have receptors for SP, 2) SP may modulate vagal output specially by increasing neuronal excitability; 3) although elec~ophysiologic~ effects of SP have been studied in a variety of central and periphera1 neurons, this is the first evidence of SP effects on DMV; 4) DMV is a particular brain target site in which SP may induce depo- or hyperpolarization. Substance P
Intracellular recording
Rat
Brainstem slice
Dorsal motor nucleus of the vagus
was 1 to 2 ml/min. Recording glass micropipettes were filled with 3 M potassium acetate (DC resistance, 60 to 150 MII1). The identi~cation of DMV and recording procedure were the same as previously described (18). The data are presented as means + SD.
SUBSTANCE P (SP) is considered to be a neuroregulator (transmitter and/or modulator) in the central and peripheral nervous system. Excitatory effects of SP have been shown in locus coemleus (Z), myenteric (8), sympathetic (5), spinal dorsal horn (17), and nucleus tractus solitarius (NTS) (15) neurons. On the other hand, nuclei which contain SP-like immunoreactive cell bodies and fibers (1, 6, 9-11, 13, 14, 16, 20) project directly to the DMV, and some of the vagal afferent fibers which also contain SP (6,12) establish synaptic connections with DMV neurons (7). Thus, SP could play a role in the regulation of vagal input and output. However, the effects of SP on the DMV have not yet been studied. In the present study we investigated the effects of SP on DMV neurons recorded in~cellul~ly .
RESULTS
Only neurons that had membrane resting potentials exceeding - 50 mV and action potential with overshoot were considered. Membrane depolarization was measured by comparing the resting potential before SP application with that at the maximal response after application. Membrane testing potential and membrane input mV and 109.92 19.2 Mfl, (n=43), resistance were -61~7.8 respectively. Action potentials ranged from 60 to 90 mV. Superfusion with SP (0.8, 1.7 and 3.4X 10e6 M) depolarized 18 of 43 DMV neurons (42%). The depolarization by SP (7.3 * 2.4 mV by l.7~lO~~M,n=ll,f=lO.l,~
METHOD
Transverse brainstem slices (400 &rn thick) containing the DMV were obtained from adult male Wistar rats, and were incubated for 90 min in oxygenated Krebs Ringer solution at room temperature, as previously described ( 18,19). The composition of Krebs Ringer solution (in mM) was: NaCl, 124; KCI, 5; NaH,PO,, 1.24; CaCl,, 2.4; MgSO,, 1.3; NaHCO,, 26; and glucose, 10 (pH about 7.4). A single slice was transferred to a recording chamber and submerged in a continuously flowing solution equilibrated with 95% 0, and 5% CO, at 37°C. In all cases SP (Peptide Institute, Osaka, Japan) was applied by super-fusion; the flow rate
‘Requests for reprints should be addressed to Carlos R. Plats-Salaman.
149
150
PLATA-SALAMAN
ET AL.
1 OmV
, .
,. 2 min IO.11 nA 2 min
ca
CONTROL
SP (1.7
X10-6M)
Cb
*CONTROL nSP
50 ms
PK. 1. Depolarization induced by SP on DMV neuron, (A) Superfusion with SP (indicated by overbar) caused a 9.2 mV depolarization and bursting of action potentials with about 13% increase in membrane input resistance as indicated by an increase in the amplitude of electrotonic potentials represented as downward deflections which correspond to hyperpolarizing pulses (resting membrane potential, - 68 mV; action potential, 88 mV). (B) When a negative current was applied through the recording electrode during SP depolarization to restore the membrane potential to the value before SP application, membrane resistance retained the control values (depolarization, 8.4 mV; resting membrane potential, - 57 mV; action potential, 86 mV). Upper traces, membrane potential; lower traces, applied current. The responses induced by SP began 2 to 4 min after starting perfusion because of superfusion time. (C) Current-voltage relationship of SP depolarization. (a) voltage deflections (upper) and rectangular depo- and hyperpolarizing current pulses (lower) before (control) and during SP application; (b) plot of the current-voltage relationship. Note the increase in membrane input resistance during SP application (1.7 x 10e6 M, filled squares) as compared with control (filled circles). Apparent reversal potential (arrow), - 66 mV. With long hyperpolarizing current pulses, a reduction in the potential change was observed (anomalous rectification, dashed lines). A, B, and C, different neurons.
Different concentrations of SP (4.2 x lo-’ to 3.4 x 1O-6 M) induced dose-dependent depolarizations (Fig. 2A). Repeated application of SP did not cause desensitization. Suferfusion with a solution containing 12 mM Mg*+ and 0 mM Ca + for more than 15 min, did not affect the depolarizing effect of SP accompanied by increased membrane input resistance (Fig. 2B). This indicates that the depolarization induced by SP was a direct effect on the postsynaptic membrane. Membrane hyperpolarization by SP was also produced in five neurons (11.6%) at a concentration of 1.7~ 10K6 M (8.423.6 mV hyperpolarization, t= 5.21, p
This study shows that SP depolarizes approximately 40% of DMV neurons by a direct effect on the postsynaptic membrane accompanied by an increased firing rate and membrane input resistance. SP hyperpolarized 12% of DMV neurons also by a direct effect on the postsynaptic membrane. In a previous study,
cholecystokinin octapeptide induced similar effects on DMV neurons (19). Our data support previous intracellular recordings which showed that SP produced neuronal depolarization associated with an increase in membrane input resistance (4,17). In our studies, the membrane depolarization and increased membrane input resistance caused by SP were eliminated by bringing back the membrane potential to the preapplication level with administered current (Figs. 1B and 2A); this supports the notion that the increased membrane input resistance may be due to the closing of a population of voltage-dependent channels. It is also possible that part of the effects observed in this study could be mediated through various tachykinin receptor subtypes because of the high doses of SP used. Thus, SP effects on DMV neurons together with the evidence that SP binding sites (21) and SP-like immunoreactive cell bodies and fibers (22) are present in the DMV suggest a role for SP as a neuroregulator in this area. SP-containing fibers to the DMV may directly modulate visceral afferent and vago-vagal reflexes (3) and DMV neuronal output through vagal efferents influencing cardiovascular (22) and subdiaphragmatic visceral functions such as gastric acid secretion (20) and motility (20). and pancreatic hormone secretion (20).
SUBSTANCE
151
P AND THE VAGAL NUCLEUS
SP (3.4 x10%9
SP
(0.8 XlO+M)
.,
SP (4.2X
Normal sp
,
,,.‘,
,._
.
.
1 O-TM)
Ringer
(1.7 XlO+M)
OmMCa+~ SP (1.7
12mMMg~~ X1(T%l)
2 min PIG. 2. (A) Dose-dependent depolarizing effect by SP on DMV. SP (0.8, 1.7 and 3.4 X 10m6 M) caused 2.4, 5.4 and 13.2 mV depoharization, respectively, in the same neuron (resting membrane potential, - 72 mV; action potential, 74 mV). Increase in input membrane resistance was about 14% and 40% for 1.7 and 3.4 x 10e6 M, respectively. Dashed line, level of original resting membrane potential. (B) Depolarization by SP after elimination of synaptic inputs, Superfusion with a solution containing 12 mM Mg’+, 0 mM Ca2+ did not alter SP-induced ~~l~~tion (7.6 mV), or the ~om~ying 21% increase in membrane input resistance (resting membrane potential, -63 mV; action potential, 76 mV). B upper and lower, same neuron.
152
PLATA-SALAMAN
ET AL.
SP (1.7 XlO+M)
SP (1.7 X lO+M)
OmMCa++ 12mMMg++ SP (1.7 XdM)
I1OmV 2 min FIG. 3. Hypetpolarization induced by SP on DMV neuron. (A) Superfusion with SP caused a 7.2 mV hyperpolarization with about 35% decrease in membrane input resistance (resting membrane potential, - 5 1 mV; action potential, 75 mV). (B) When a positive current was applied through the recording electrode during SP hyperpolarization, membrane input resistance retained the SP-induced decrease of 53% (hyperpolarization, 14 mV; resting membrane potential, -55 mV). (C) Hypetpolarization by SP after elimination of synaptic inputs. Superfusion with a solution containing 12 mM MgZc, 0 mM Caa+ did not prevent SP-induced hyperpolarization (6.8 mV), or the accompanying decrease (26%) in membrane input resistance (resting membrane potential, - 68 mV). A, B, and C, different neurons.
ACKNOWLEDGEMENTS This work was supported by grants-in-aid for scientific research from the Ministry of Education, Science and Culture of Japan, when research was done at Kyushu University.
REFERENCES 1. Carter, D. A.; Lightman, S. L. Comparative distribution and cardiovascular actions of substance P and substance K within the nucleus tractus solitarius of rats. Neuropeptides 8:295-304; 1986. 2. Cheeseman, H. J.; Pinnock, R. D.; Henderson, G. Substance P excitation of rat locus coeruleus neurones. Eur. J. Pharmacol. 94: 93-99; 1983. 3. Colhnan, P. I.; Grundy, D.; Scratcherd, T.; Wach, R. A. Vago-vagal reflexes to the colon of the anaesthetized ferret. J. Physiol. 352: 395-402; 1984. V. A. Substance P depolarizes nerve 4. Dryer, S. E.; Chiappinelli, terminals in an autonomic ganglion. Brain Res. 336:190-194; 1985. 5. Dun, N. J.; MO, N. In vitro effects of substance P on neonatal rat sympathetic preganglionic neurones. J. Physiol. 399:321-333; 1988. 6. Gillis, R. A.; Helke, C. J.; Hamilton, B. L.; Norman, W. P.; Jacobowitz, D. Evidence that substance P is a neurotransmitter of baro- and chemoreceptor afferents in the nucleus of tractus solitarius. Brain Res. 181:476-181; 1980.
7. Kalia, M.; Sullivan, J. M. Brainstem projections of sensory and motor components of the vagus nerve in the rat. J. Comp. Neurol. 211: 248-264; 1982. 8. Katayama, Y.; North, R. A.; Williams, J. T. The action of substance P on neurons of the myentetic plexus of the guinea-pig small intestine. Proc. R. Sot. Lond. [Biol.] 206:191-208: 1979. 9. Kubota, Y.; Takagi, H.; Morishima, Y.; Powell, J. F.; Smith, A. D. Synaptic interaction between catecholaminergic neurons and substance P-immunoteactive axons in the caudal part of the nucleus of the solitary tract of the rat: demonstration by the electron microscopic mirror technique. Brain Res. 333:188-192; 1985. 10. Lawrence, D.; Pittman, Q. J. Interaction between descending paraventricular neurons and vagal motor neurons. Brain Res. 332:158-160; 1985. Il. Leibstein, A. G.; Dermietzel, R.; Willenberg, I. M.; Pauschert, R. Mapping of different neuropeptides in the lower brainstem of the rat:
SUBSTANCE P AND THE VAGAL NUCLEUS
12.
13. 14.
15.
16.
with special reference to the ventral surface. J. Auton. Nerv. Syst. 14:299-313; 1985. MacLean, D. B. Substance P and somatostatin content and transport in vagus and sciatic nerves of the streptozocin-induced diabetic rat. Diabetes 36390-395; 1987. Mai, J. K.; Stephens, P. H.; Hopf, A.; Cuello, A. C, Substance P in the human brain. Neuroscience 17:709-739; 1986. Maley, B. E. The ultrastructural localization of eukephalin and substance P immunoreactivities in the nucleus tractus solitarii of the cat. J. Comp. Neurol. 233:490-496; 1985. Morin-Surun, M. P.; Jordan, D.; Champagnat, J.; Spyer, K. M.; Denavit-Saubie, M. Excitatory effects of iontophoretically applied substance P on neurons in the nucleus tractus solitarius of the cat: lack of interaction with opiates and opioids. Brain Res. 307:388-392; 1984. Morishima, Y .; Takagi, H.; Kawai, Y .; Emson, P. C.; Hillyard, C. J.; Girgis, S. I.; MacIntyre. I. Ultrastructural observation of nerve fibers containing both substance P and calcitonin-gene related peptide in me
153
nucleus tractus solitarii of the rat. Brain Res. 379:157-161; 1986. 17. Murase, K.; Randie, M. Actions of substance P on the spinal dorsal horn neurons. J. physiol. 346:203-217; 1984. 18. Oomura, Y.; Mizuno, Y. Effect of somatostatin on the vagal motor neuron in the rat. Brain Res. Bull. 17:397-401; 1986. 19. Pla~-S~~~, C. R.; Fukuda, A.; Oomura, Y.; Minami, T. Effects of sulphated cholecystokinin octapeptide (CCK-8) on the dorsal motor nucleus of the vagus. Brain Res. Bull. 21:839-842; 1988. 20. Rogers, R. C.; Kita, H.; Butcher, L. L.; Novin, D. Afferent projections to the dorsal motor nucleus of the vagus. Brain Res. Bull, 5365-373; 1980. 21. Shigematsu, K.; Niwa, M.; Saavedra, J. M. Increased density of substance P binding sites in specific brainstem nuclei of spontaneously hypertensive rats. Brain Res. 370:383-387; 1986. 22. Triepel, J.; Weindl, A.; Kiemle, I.; Mader, J. ; VoIz, H. P. ; Reinecke, M.; Forssmann, W. G. Substance P-immunoreactive neurons in the brainstem of the cat related to cardiovascular centers. Cell Tissue Res. 241:3141; 1985.
Brain ResearchBulletin, Vol. 23, PP. 149-153. Q Pergamon Press pk. 1989. Printed in the U.S.A
BRIEF COMMUNICATION
Substance P Effects on the Dorsal Motor Nucleus of the Vagus CARLOS
R. PLATA-SALAMAN,’
ATSUO FUKUDA,* TAKETSUGU MINAMI”
AND YUTAKA
OOMURA*
Department ofP~y~h~~~gyand Institute furNeuroscience, Universes of Delaware, Newark, DE 19716 and *Department of Physiology, Faculty of Medicine, Kyushu University, Fukuoka 812, Japan Received 15 March 1989 PLATA-SALAMAN, C. R., A. PLJKUDA, T. MINAh41AND Y. OOMURA. Substance P eflects on the dorsal motor nucleus of the vague. BRAIN RES BULL 23(1/Z) 149-153, 1989.~-Substance P (SP) was applied by superfusion (0.8 to 3.4 X 10e6 M) to neurons of the dorsal motor nucleus of the vagus {DMV) in slice preparations of the rat medulla oblongata. In~acelluI~ recordings showed that 18 of 43 (42%) neurons were depolarized and the depolarization was associated with an increase in membrane input resistance; five of 43 (12%) neurons were hyperpolarized and tire hyperpolarization was associated with a decrease in membrane input resistance. Both effects were reversible and persisted after blockade of synaptic transmission by Ca*’ free/high Mg*+ solution. These data show that: 1) vagal neurons in the DMV have receptors for SP, 2) SP may modulate vagal output specially by increasing neuronal excitability; 3) although elec~ophysiologic~ effects of SP have been studied in a variety of central and periphera1 neurons, this is the first evidence of SP effects on DMV; 4) DMV is a particular brain target site in which SP may induce depo- or hyperpolarization. Substance P
Intracellular recording
Rat
Brainstem slice
Dorsal motor nucleus of the vagus
was 1 to 2 ml/min. Recording glass micropipettes were filled with 3 M potassium acetate (DC resistance, 60 to 150 MII1). The identi~cation of DMV and recording procedure were the same as previously described (18). The data are presented as means + SD.
SUBSTANCE P (SP) is considered to be a neuroregulator (transmitter and/or modulator) in the central and peripheral nervous system. Excitatory effects of SP have been shown in locus coemleus (Z), myenteric (8), sympathetic (5), spinal dorsal horn (17), and nucleus tractus solitarius (NTS) (15) neurons. On the other hand, nuclei which contain SP-like immunoreactive cell bodies and fibers (1, 6, 9-11, 13, 14, 16, 20) project directly to the DMV, and some of the vagal afferent fibers which also contain SP (6,12) establish synaptic connections with DMV neurons (7). Thus, SP could play a role in the regulation of vagal input and output. However, the effects of SP on the DMV have not yet been studied. In the present study we investigated the effects of SP on DMV neurons recorded in~cellul~ly .
RESULTS
Only neurons that had membrane resting potentials exceeding - 50 mV and action potential with overshoot were considered. Membrane depolarization was measured by comparing the resting potential before SP application with that at the maximal response after application. Membrane testing potential and membrane input mV and 109.92 19.2 Mfl, (n=43), resistance were -61~7.8 respectively. Action potentials ranged from 60 to 90 mV. Superfusion with SP (0.8, 1.7 and 3.4X 10e6 M) depolarized 18 of 43 DMV neurons (42%). The depolarization by SP (7.3 * 2.4 mV by l.7~lO~~M,n=ll,f=lO.l,~
METHOD
Transverse brainstem slices (400 &rn thick) containing the DMV were obtained from adult male Wistar rats, and were incubated for 90 min in oxygenated Krebs Ringer solution at room temperature, as previously described ( 18,19). The composition of Krebs Ringer solution (in mM) was: NaCl, 124; KCI, 5; NaH,PO,, 1.24; CaCl,, 2.4; MgSO,, 1.3; NaHCO,, 26; and glucose, 10 (pH about 7.4). A single slice was transferred to a recording chamber and submerged in a continuously flowing solution equilibrated with 95% 0, and 5% CO, at 37°C. In all cases SP (Peptide Institute, Osaka, Japan) was applied by super-fusion; the flow rate
‘Requests for reprints should be addressed to Carlos R. Plats-Salaman.
149
150
PLATA-SALAMAN
ET AL.
1 OmV
, .
,. 2 min IO.11 nA 2 min
ca
CONTROL
SP (1.7
X10-6M)
Cb
*CONTROL nSP
50 ms
PK. 1. Depolarization induced by SP on DMV neuron, (A) Superfusion with SP (indicated by overbar) caused a 9.2 mV depolarization and bursting of action potentials with about 13% increase in membrane input resistance as indicated by an increase in the amplitude of electrotonic potentials represented as downward deflections which correspond to hyperpolarizing pulses (resting membrane potential, - 68 mV; action potential, 88 mV). (B) When a negative current was applied through the recording electrode during SP depolarization to restore the membrane potential to the value before SP application, membrane resistance retained the control values (depolarization, 8.4 mV; resting membrane potential, - 57 mV; action potential, 86 mV). Upper traces, membrane potential; lower traces, applied current. The responses induced by SP began 2 to 4 min after starting perfusion because of superfusion time. (C) Current-voltage relationship of SP depolarization. (a) voltage deflections (upper) and rectangular depo- and hyperpolarizing current pulses (lower) before (control) and during SP application; (b) plot of the current-voltage relationship. Note the increase in membrane input resistance during SP application (1.7 x 10e6 M, filled squares) as compared with control (filled circles). Apparent reversal potential (arrow), - 66 mV. With long hyperpolarizing current pulses, a reduction in the potential change was observed (anomalous rectification, dashed lines). A, B, and C, different neurons.
Different concentrations of SP (4.2 x lo-’ to 3.4 x 1O-6 M) induced dose-dependent depolarizations (Fig. 2A). Repeated application of SP did not cause desensitization. Suferfusion with a solution containing 12 mM Mg*+ and 0 mM Ca + for more than 15 min, did not affect the depolarizing effect of SP accompanied by increased membrane input resistance (Fig. 2B). This indicates that the depolarization induced by SP was a direct effect on the postsynaptic membrane. Membrane hyperpolarization by SP was also produced in five neurons (11.6%) at a concentration of 1.7~ 10K6 M (8.423.6 mV hyperpolarization, t= 5.21, p
This study shows that SP depolarizes approximately 40% of DMV neurons by a direct effect on the postsynaptic membrane accompanied by an increased firing rate and membrane input resistance. SP hyperpolarized 12% of DMV neurons also by a direct effect on the postsynaptic membrane. In a previous study,
cholecystokinin octapeptide induced similar effects on DMV neurons (19). Our data support previous intracellular recordings which showed that SP produced neuronal depolarization associated with an increase in membrane input resistance (4,17). In our studies, the membrane depolarization and increased membrane input resistance caused by SP were eliminated by bringing back the membrane potential to the preapplication level with administered current (Figs. 1B and 2A); this supports the notion that the increased membrane input resistance may be due to the closing of a population of voltage-dependent channels. It is also possible that part of the effects observed in this study could be mediated through various tachykinin receptor subtypes because of the high doses of SP used. Thus, SP effects on DMV neurons together with the evidence that SP binding sites (21) and SP-like immunoreactive cell bodies and fibers (22) are present in the DMV suggest a role for SP as a neuroregulator in this area. SP-containing fibers to the DMV may directly modulate visceral afferent and vago-vagal reflexes (3) and DMV neuronal output through vagal efferents influencing cardiovascular (22) and subdiaphragmatic visceral functions such as gastric acid secretion (20) and motility (20). and pancreatic hormone secretion (20).
SUBSTANCE
151
P AND THE VAGAL NUCLEUS
SP (3.4 x10%9
SP
(0.8 XlO+M)
.,
SP (4.2X
Normal sp
,
,,.‘,
,._
.
.
1 O-TM)
Ringer
(1.7 XlO+M)
OmMCa+~ SP (1.7
12mMMg~~ X1(T%l)
2 min PIG. 2. (A) Dose-dependent depolarizing effect by SP on DMV. SP (0.8, 1.7 and 3.4 X 10m6 M) caused 2.4, 5.4 and 13.2 mV depoharization, respectively, in the same neuron (resting membrane potential, - 72 mV; action potential, 74 mV). Increase in input membrane resistance was about 14% and 40% for 1.7 and 3.4 x 10e6 M, respectively. Dashed line, level of original resting membrane potential. (B) Depolarization by SP after elimination of synaptic inputs, Superfusion with a solution containing 12 mM Mg’+, 0 mM Ca2+ did not alter SP-induced ~~l~~tion (7.6 mV), or the ~om~ying 21% increase in membrane input resistance (resting membrane potential, -63 mV; action potential, 76 mV). B upper and lower, same neuron.
152
PLATA-SALAMAN
ET AL.
SP (1.7 XlO+M)
SP (1.7 X lO+M)
OmMCa++ 12mMMg++ SP (1.7 XdM)
I1OmV 2 min FIG. 3. Hypetpolarization induced by SP on DMV neuron. (A) Superfusion with SP caused a 7.2 mV hyperpolarization with about 35% decrease in membrane input resistance (resting membrane potential, - 5 1 mV; action potential, 75 mV). (B) When a positive current was applied through the recording electrode during SP hyperpolarization, membrane input resistance retained the SP-induced decrease of 53% (hyperpolarization, 14 mV; resting membrane potential, -55 mV). (C) Hypetpolarization by SP after elimination of synaptic inputs. Superfusion with a solution containing 12 mM MgZc, 0 mM Caa+ did not prevent SP-induced hyperpolarization (6.8 mV), or the accompanying decrease (26%) in membrane input resistance (resting membrane potential, - 68 mV). A, B, and C, different neurons.
ACKNOWLEDGEMENTS This work was supported by grants-in-aid for scientific research from the Ministry of Education, Science and Culture of Japan, when research was done at Kyushu University.
REFERENCES 1. Carter, D. A.; Lightman, S. L. Comparative distribution and cardiovascular actions of substance P and substance K within the nucleus tractus solitarius of rats. Neuropeptides 8:295-304; 1986. 2. Cheeseman, H. J.; Pinnock, R. D.; Henderson, G. Substance P excitation of rat locus coeruleus neurones. Eur. J. Pharmacol. 94: 93-99; 1983. 3. Colhnan, P. I.; Grundy, D.; Scratcherd, T.; Wach, R. A. Vago-vagal reflexes to the colon of the anaesthetized ferret. J. Physiol. 352: 395-402; 1984. V. A. Substance P depolarizes nerve 4. Dryer, S. E.; Chiappinelli, terminals in an autonomic ganglion. Brain Res. 336:190-194; 1985. 5. Dun, N. J.; MO, N. In vitro effects of substance P on neonatal rat sympathetic preganglionic neurones. J. Physiol. 399:321-333; 1988. 6. Gillis, R. A.; Helke, C. J.; Hamilton, B. L.; Norman, W. P.; Jacobowitz, D. Evidence that substance P is a neurotransmitter of baro- and chemoreceptor afferents in the nucleus of tractus solitarius. Brain Res. 181:476-181; 1980.
7. Kalia, M.; Sullivan, J. M. Brainstem projections of sensory and motor components of the vagus nerve in the rat. J. Comp. Neurol. 211: 248-264; 1982. 8. Katayama, Y.; North, R. A.; Williams, J. T. The action of substance P on neurons of the myentetic plexus of the guinea-pig small intestine. Proc. R. Sot. Lond. [Biol.] 206:191-208: 1979. 9. Kubota, Y.; Takagi, H.; Morishima, Y.; Powell, J. F.; Smith, A. D. Synaptic interaction between catecholaminergic neurons and substance P-immunoteactive axons in the caudal part of the nucleus of the solitary tract of the rat: demonstration by the electron microscopic mirror technique. Brain Res. 333:188-192; 1985. 10. Lawrence, D.; Pittman, Q. J. Interaction between descending paraventricular neurons and vagal motor neurons. Brain Res. 332:158-160; 1985. Il. Leibstein, A. G.; Dermietzel, R.; Willenberg, I. M.; Pauschert, R. Mapping of different neuropeptides in the lower brainstem of the rat:
SUBSTANCE P AND THE VAGAL NUCLEUS
12.
13. 14.
15.
16.
with special reference to the ventral surface. J. Auton. Nerv. Syst. 14:299-313; 1985. MacLean, D. B. Substance P and somatostatin content and transport in vagus and sciatic nerves of the streptozocin-induced diabetic rat. Diabetes 36390-395; 1987. Mai, J. K.; Stephens, P. H.; Hopf, A.; Cuello, A. C, Substance P in the human brain. Neuroscience 17:709-739; 1986. Maley, B. E. The ultrastructural localization of eukephalin and substance P immunoreactivities in the nucleus tractus solitarii of the cat. J. Comp. Neurol. 233:490-496; 1985. Morin-Surun, M. P.; Jordan, D.; Champagnat, J.; Spyer, K. M.; Denavit-Saubie, M. Excitatory effects of iontophoretically applied substance P on neurons in the nucleus tractus solitarius of the cat: lack of interaction with opiates and opioids. Brain Res. 307:388-392; 1984. Morishima, Y .; Takagi, H.; Kawai, Y .; Emson, P. C.; Hillyard, C. J.; Girgis, S. I.; MacIntyre. I. Ultrastructural observation of nerve fibers containing both substance P and calcitonin-gene related peptide in me
153
nucleus tractus solitarii of the rat. Brain Res. 379:157-161; 1986. 17. Murase, K.; Randie, M. Actions of substance P on the spinal dorsal horn neurons. J. physiol. 346:203-217; 1984. 18. Oomura, Y.; Mizuno, Y. Effect of somatostatin on the vagal motor neuron in the rat. Brain Res. Bull. 17:397-401; 1986. 19. Pla~-S~~~, C. R.; Fukuda, A.; Oomura, Y.; Minami, T. Effects of sulphated cholecystokinin octapeptide (CCK-8) on the dorsal motor nucleus of the vagus. Brain Res. Bull. 21:839-842; 1988. 20. Rogers, R. C.; Kita, H.; Butcher, L. L.; Novin, D. Afferent projections to the dorsal motor nucleus of the vagus. Brain Res. Bull, 5365-373; 1980. 21. Shigematsu, K.; Niwa, M.; Saavedra, J. M. Increased density of substance P binding sites in specific brainstem nuclei of spontaneously hypertensive rats. Brain Res. 370:383-387; 1986. 22. Triepel, J.; Weindl, A.; Kiemle, I.; Mader, J. ; VoIz, H. P. ; Reinecke, M.; Forssmann, W. G. Substance P-immunoreactive neurons in the brainstem of the cat related to cardiovascular centers. Cell Tissue Res. 241:3141; 1985.