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Neuropeptides in afferent pathways to the urinary bladder and colon
$152 EFFECTS
OF
GALANIN
ON ENDOCRINE
FUNCTIONS
OF
STOMACH
AND
PANCREAS
IN THE
RAT
NOBUYUKI YAGI*, CHIZUKO YANAIHARA*, TOHRU MOCHIZUKI*, JUNJI ISHIKAWA*, and NOBORU YANAIHARA Laboratory of Bioorganic Chemistry, Shizuoka College of Pharmacy, 2-2-10shika, Shizuoka 422 G a l a n i n is a n e w b r a i n - g u t peptide consisting o f 29 a m i n o a c i d r e s i d u e s . In the present study, the effects of galanin on gastric and pancreatic endocrine functions were examined with synthetic replicate in isolated rat stomach and pancreas perfusion systems. Synthetic galanin was prepared by a solid phase technique. In an isolated stomach perfusion system, synthetic galanin (10 - 9 10-7M) had little effect on the basal secretion of somatostatin and gastrin, while it suppressed gastrin release induced by methacholine (10-SM) . The stimulated g a s t r i n r e l e a s e w a s r e d u c e d b y g a l a n i n to 4 9 . 1 + 2 . 1 6 % at 10-9M, to 35.0±2.47%(p<0.05) at 10-SM and to 23.510.84%(p<0.01) at 10-7M. In a similar manner, the gastrin release stimulated by porcine GRP(14-27) (10-7M) was also suppressed by galanin (I0-9-I0-7M) dose-dependently. During the course of the experiments, somatostatin levels, suppressed by methacholine or stimulated by porcine GRP(14-27), were not altered by the presence of galanin (I0-9-I0-7M) . In r a t s t o m a c h t i s s u e a n a m i n o - t e r m i n a l specific anti-galanin monoclonal antibody B4G9 immunostained a dense population o f n e r v e s in t h e p y l o r i c s p h i n c t e r a n d lamina muscularis mucosa and scattered nerves in the pyloric mucosa. Synthetic galanin (10-10-10-7M) also showed a suppressing effect on glucose (150, 2 5 0 a n d 300 m g % ) - i n d u c e d insulin release in an isolated pancreas perfusion system. The suppressing effects of galanin on glucose-induced insulin release increased dose-dependently at concentrations ranging from 10-10M to 10-7M. Somatostatin concentrations in the perfusates d i d n o t c h a n g e in t h e p r e s e n c e o f g a l a n i n . Imamunostaining of rat pancreatic tissue revealed galanin immunoreactivity in nervous elements, but not in the islet cells. The present results support the concept that galanin participates in an inhibitory system in the peripheral neural regulation of endocrine functions of rat stomach and pancreas, at least not through the release of somatostatin. DIFFERENCES IN THE SYNAFTIC STRUCTURE OF" ?EPTIDE NERVE TERMINALS DERIVED FROM THE LUMBAR SPLANCHNIC AND COLONIC NERVES IN THE INFERIOR MESENTERIC GANGLION SADAHIKO MASUKO* and TANEMICHI CHIBA Department of Anatomy, Saga Medical School, Nabeshima, Saga 840-01, Japan The inferior mesenteric ganglion (IMG) receives substance P (SP)-immunoreactive (IR) primary afferent fibers as we]] as enkepha]in (ENK)-IR pregang]ionic fibers from the lumbar sp]anchnie nerves and dynorphin (DYN)-, vasoaetive intestinal po]ypeptide (VIP)-, cho]ecystokinin (CCK)- and bombesin-IR fibers from the colonic nerves. In the physiological studies of the IMG, an excitatory action of SP, an inhibitory action of ENK and modu]atory actions of VIP and CCK have been suggested. However, the synaptic structures of these peptidergic nerve terminals have not been studied sufficiently. Male adult guinea pigs were anesthetized with Nembutal and the IMGs were partially denervated leaving the lumbar sp]anchnic nerves or the colonic nerves. After a postoperative survival time of 2-4 days, the IMGs were fixed with Zamboni's fixative. The u]trastruetura] organization of SP-, ENK-, DYN-, VIP- and CCK-IR nerve terminals was studied by preembedding staining with the PAP method for electron microscopy. In the ganglia denervated leaving the lumbar sp]anchnic nerves intact, the ENK-IR fibers were highest in density and the SP- or VIP-IR fibers were moderate. The SP-IR and ENK-IB fibers showed varicosities of the sma]] c]ear vesicle predominant type (SV type), whereas the VIP-IR fibers showed varieosities of the large vesicle predominant type (LV type). On]y 3 of 45 SP-IR varieosities and ii of 7 1 E N K - I R varicosities showed syanptic contacts, and a]] of them were axo-dendritic synapses with asymmetrical specialization. In a tots] of 42 VIP-IR fibers 2 axo-dendritie symmetrical synapses were found. In the gang]is denervatied leaving the colonic nerves intact, DYN-IR, VIP-IR and CCK-IR fibers were densely distributed and their varicosities were mainly of the LV type. In a total of 75 DYN-IB, 75 VIP-IR and 60 CCK-IR varicosities, iO, 12 and la axo-dendritic synapses were found, respectively. Only one axo-somatic synapse of CCK-IR varicosity was found. Most of these syanptic specializations were symmetrica]. The light microscopic immunocytochemistry of serial semi-thin sections (0.5 ~m) revealed that DYN-IR~ VIP-IR and CCK-IR fibers distributed in a similar pattern and were frequently found in identical nerve fibers, suggesting the coexistence of these three peptides in the single varicosity. The difference of synaptic structures between the SV type var~cosities with asymmetrical synapses (SP-IR and ENK-IR terminals) and the LV type varicosities with symmetrical synapses (DYN-IR, VIP-IR and CCK-IR terminals) might relate to the difference of the modes of action among these peptides.
OF
GALANIN
ON ENDOCRINE
FUNCTIONS
OF
STOMACH
AND
PANCREAS
IN THE
RAT
NOBUYUKI YAGI*, CHIZUKO YANAIHARA*, TOHRU MOCHIZUKI*, JUNJI ISHIKAWA*, and NOBORU YANAIHARA Laboratory of Bioorganic Chemistry, Shizuoka College of Pharmacy, 2-2-10shika, Shizuoka 422 G a l a n i n is a n e w b r a i n - g u t peptide consisting o f 29 a m i n o a c i d r e s i d u e s . In the present study, the effects of galanin on gastric and pancreatic endocrine functions were examined with synthetic replicate in isolated rat stomach and pancreas perfusion systems. Synthetic galanin was prepared by a solid phase technique. In an isolated stomach perfusion system, synthetic galanin (10 - 9 10-7M) had little effect on the basal secretion of somatostatin and gastrin, while it suppressed gastrin release induced by methacholine (10-SM) . The stimulated g a s t r i n r e l e a s e w a s r e d u c e d b y g a l a n i n to 4 9 . 1 + 2 . 1 6 % at 10-9M, to 35.0±2.47%(p<0.05) at 10-SM and to 23.510.84%(p<0.01) at 10-7M. In a similar manner, the gastrin release stimulated by porcine GRP(14-27) (10-7M) was also suppressed by galanin (I0-9-I0-7M) dose-dependently. During the course of the experiments, somatostatin levels, suppressed by methacholine or stimulated by porcine GRP(14-27), were not altered by the presence of galanin (I0-9-I0-7M) . In r a t s t o m a c h t i s s u e a n a m i n o - t e r m i n a l specific anti-galanin monoclonal antibody B4G9 immunostained a dense population o f n e r v e s in t h e p y l o r i c s p h i n c t e r a n d lamina muscularis mucosa and scattered nerves in the pyloric mucosa. Synthetic galanin (10-10-10-7M) also showed a suppressing effect on glucose (150, 2 5 0 a n d 300 m g % ) - i n d u c e d insulin release in an isolated pancreas perfusion system. The suppressing effects of galanin on glucose-induced insulin release increased dose-dependently at concentrations ranging from 10-10M to 10-7M. Somatostatin concentrations in the perfusates d i d n o t c h a n g e in t h e p r e s e n c e o f g a l a n i n . Imamunostaining of rat pancreatic tissue revealed galanin immunoreactivity in nervous elements, but not in the islet cells. The present results support the concept that galanin participates in an inhibitory system in the peripheral neural regulation of endocrine functions of rat stomach and pancreas, at least not through the release of somatostatin. DIFFERENCES IN THE SYNAFTIC STRUCTURE OF" ?EPTIDE NERVE TERMINALS DERIVED FROM THE LUMBAR SPLANCHNIC AND COLONIC NERVES IN THE INFERIOR MESENTERIC GANGLION SADAHIKO MASUKO* and TANEMICHI CHIBA Department of Anatomy, Saga Medical School, Nabeshima, Saga 840-01, Japan The inferior mesenteric ganglion (IMG) receives substance P (SP)-immunoreactive (IR) primary afferent fibers as we]] as enkepha]in (ENK)-IR pregang]ionic fibers from the lumbar sp]anchnie nerves and dynorphin (DYN)-, vasoaetive intestinal po]ypeptide (VIP)-, cho]ecystokinin (CCK)- and bombesin-IR fibers from the colonic nerves. In the physiological studies of the IMG, an excitatory action of SP, an inhibitory action of ENK and modu]atory actions of VIP and CCK have been suggested. However, the synaptic structures of these peptidergic nerve terminals have not been studied sufficiently. Male adult guinea pigs were anesthetized with Nembutal and the IMGs were partially denervated leaving the lumbar sp]anchnic nerves or the colonic nerves. After a postoperative survival time of 2-4 days, the IMGs were fixed with Zamboni's fixative. The u]trastruetura] organization of SP-, ENK-, DYN-, VIP- and CCK-IR nerve terminals was studied by preembedding staining with the PAP method for electron microscopy. In the ganglia denervated leaving the lumbar sp]anchnic nerves intact, the ENK-IR fibers were highest in density and the SP- or VIP-IR fibers were moderate. The SP-IR and ENK-IB fibers showed varicosities of the sma]] c]ear vesicle predominant type (SV type), whereas the VIP-IR fibers showed varieosities of the large vesicle predominant type (LV type). On]y 3 of 45 SP-IR varieosities and ii of 7 1 E N K - I R varicosities showed syanptic contacts, and a]] of them were axo-dendritic synapses with asymmetrical specialization. In a tots] of 42 VIP-IR fibers 2 axo-dendritie symmetrical synapses were found. In the gang]is denervatied leaving the colonic nerves intact, DYN-IR, VIP-IR and CCK-IR fibers were densely distributed and their varicosities were mainly of the LV type. In a total of 75 DYN-IB, 75 VIP-IR and 60 CCK-IR varicosities, iO, 12 and la axo-dendritic synapses were found, respectively. Only one axo-somatic synapse of CCK-IR varicosity was found. Most of these syanptic specializations were symmetrica]. The light microscopic immunocytochemistry of serial semi-thin sections (0.5 ~m) revealed that DYN-IR~ VIP-IR and CCK-IR fibers distributed in a similar pattern and were frequently found in identical nerve fibers, suggesting the coexistence of these three peptides in the single varicosity. The difference of synaptic structures between the SV type var~cosities with asymmetrical synapses (SP-IR and ENK-IR terminals) and the LV type varicosities with symmetrical synapses (DYN-IR, VIP-IR and CCK-IR terminals) might relate to the difference of the modes of action among these peptides.