Mechanisms of Neurotensin Effects on Pancreatic and Duodenal Bicarbonate Secretion in the Rat C. NAGAIN, D. MERLIN, J. CHARIOT, AND C. ROZE INSERM U239 Facultk X . Bichat 16 rue H. Huchard 75018 Paris, France
The 13-amino-acid regulatory peptide neurotensin (NT), released by endocrine ileal N-cells, displays several effects on gastrointestinal motility and secretion, such as inhibition of gastric secretion and stimulation of duodenal bicarbonate and of external pancreatic secretion. Whether these NT effects are physiological, as well as their mechanisms, is debated. The purpose of this study was to analyze in vivo the mechanism o f NT stimulation of external pancreatic and duodenal bicarbonate secretions in the rat.
METHODS Anesthetized male Wistar rats were used. To evaluate pancreatic secretion, bile was diverted, the pylorus was ligated, pure pancreatic juice was collected, and the outputs of sodium (equivalent to volume), bicarbonate, and total proteins were measured.' To evaluate duodenal bicarbonate secretion, a segment (12 mm) of duodenum devoid of Briinner glands, with its blood supply intact, was cannulated between two glass tubes connected to a reservoir. The recirculating luminal perfusate was titrated at pH 7.4.2 NTI-I~ (0.06 to 6 nmol/kg.hr = 0.1 to 10 pg/kg.hr) was intravenously (i.v.) infused for 40 min, alone or associated with various antagonists: atropine sulfate (75 pg/kg 75 pg/kg.hr, i-v.); hexamethonium bromide (6.7 mg/kg 6.7 mg/kg.hr, i.v.); adrenoceptor antagonists: propranolol (1 mg/kg, s . ~ . ) prazosin (0.5 mg/kg, s.c.) idazoxan (0.3 mg/kg, s.c.); CCKAand CCKBantagonists L364718 or L365260 (0.5 mg/ kg, i.v.); methadone (5 mg/kg, s.c.); indomethacin (10 mg/kg, i.v.); naloxone hydrochloride (1 mg/kg, s.c.); bilateral vagotomy ; sensory denervation by chronic capsaicin administration (100 mg/kg, s.c.) 10 days before the experiment.
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RESULTS Dose-Response Curves for Neurotensin NT produced an increase of pancreatic secretion; the peak stimulation was reached 20 to 40 min after the end of the 40-min NT infusion. The dose-response curve was biphasic, with submaximal stimulation at supramaximal doses. The maximal response 359
ANNALS NEW YORK ACADEMY OF SCIENCES
360
was obtained with 1.9 nmol/kg.hr (3.16 pg/kg.hr) and represented about 30% of the maximal response to cholecystokinin. The EDsos were 170 pmol/kg.hr for sodium and bicarbonate and 250 pmol/kg.hr for protein. NT produced a dose-related increase of duodenal bicarbonate secretion. The peak stimulation was reached 30 min after beginning the 40-min NT infusion. The maximal response was obtained with 0.6 nmol/kg-hr (1 Crg/kg.hr) and represented about 80% of the maximal response to the synthetic prostaglandin enprostil. The ED50 was 60 pmol/kg.hr. Data reported in the literature suggest that NT doses found to act on duodenal and pancreatic secretion in this study may induce physiological NT plasma levels.3-5
Mechanism of Action The effect of NT on pancreatic secretion was abolished by atropine or hexamethonium; reduced (-80%) by vagotomy; reduced (-80%: sodium and bicarbonate) or abolished (protein) by L364718; unchanged by L365260 or by the association of a1 a2 fi adrenoceptor antagonists; reduced (-85%) by methadone and reduced (-70%) after capsakin pretreatment. The effect of NT on duodenal bicarbonate secretion was unchanged by atropine, naloxone, indomethacin, or L364718; reduced (-50%) by hexamethonium; reduced (-50%) by vagotomy; and reduced (-60%) after capsaicin pretreatment.
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CONCLUSION NT stimulatesexternal pancreatic secretion in the anesthetized rat with a low efficacy compared to cholecystokinin (CCK), but in a dose range compatible with postprandial plasma levels measured by others. The mechanism of action is indirect, neurally mediated, and involves nicotinic and muscarinic synapses, CCKA receptors, and in part capsakin-sensitive sensory fibers. We suggest that NT activates receptors present on peripheral afferent fibers, that this information is built up in a central reflex loop involving in part CCKA receptors, and conveyed to the pancreas by a classical cholinergic vagal efferent pathway. NT stimulates duodenal bicarbonate secretion in the anesthetized rat with a good efficacy compared to enprostil, and in a dose range compatible with postprandial plasma levels measured by others. The mechanism of action is complex. It depends for about 50% on primary sensory afferent neurons, and on vagal (sensory or motor) fibers, nicotinic synapses, and noncholinergic effector(s). The remainder of the effect is unexplained. REFERENCES 1,
2. 3. 4. 5.
RozB, C., J.
DE LA TOUR, J. CHARIOT, M. SOUCHARD & C. DEBRAY. 1975. Biol. GastroentCrol. (Paris) 8: 291-295. G. & A. GARNER. 1982. Am. J. Physiol. 242: G1834193. FLEMSTR~M, HAMMER, R. A . , R. E. HERNANDEZ & A. SHEPARD. 1990. Dig. Dis. Sci. 35: 439-443. TRIMBLE, E. R.,C. SHAW,R. BRUZZONE, A. GJINOVCI & K. D. BUCHANAN. 1987. Gastroenterology 92: 699-703. B. SYTNIK & J. WALSH.1987. J. Clin. Invest. 80: 1064-1067. MOGARD, M., V. MAXWELL,
The 13-amino-acid regulatory peptide neurotensin (NT), released by endocrine ileal N-cells, displays several effects on gastrointestinal motility and secretion, such as inhibition of gastric secretion and stimulation of duodenal bicarbonate and of external pancreatic secretion. Whether these NT effects are physiological, as well as their mechanisms, is debated. The purpose of this study was to analyze in vivo the mechanism o f NT stimulation of external pancreatic and duodenal bicarbonate secretions in the rat.
METHODS Anesthetized male Wistar rats were used. To evaluate pancreatic secretion, bile was diverted, the pylorus was ligated, pure pancreatic juice was collected, and the outputs of sodium (equivalent to volume), bicarbonate, and total proteins were measured.' To evaluate duodenal bicarbonate secretion, a segment (12 mm) of duodenum devoid of Briinner glands, with its blood supply intact, was cannulated between two glass tubes connected to a reservoir. The recirculating luminal perfusate was titrated at pH 7.4.2 NTI-I~ (0.06 to 6 nmol/kg.hr = 0.1 to 10 pg/kg.hr) was intravenously (i.v.) infused for 40 min, alone or associated with various antagonists: atropine sulfate (75 pg/kg 75 pg/kg.hr, i-v.); hexamethonium bromide (6.7 mg/kg 6.7 mg/kg.hr, i.v.); adrenoceptor antagonists: propranolol (1 mg/kg, s . ~ . ) prazosin (0.5 mg/kg, s.c.) idazoxan (0.3 mg/kg, s.c.); CCKAand CCKBantagonists L364718 or L365260 (0.5 mg/ kg, i.v.); methadone (5 mg/kg, s.c.); indomethacin (10 mg/kg, i.v.); naloxone hydrochloride (1 mg/kg, s.c.); bilateral vagotomy ; sensory denervation by chronic capsaicin administration (100 mg/kg, s.c.) 10 days before the experiment.
+
+
+
+
RESULTS Dose-Response Curves for Neurotensin NT produced an increase of pancreatic secretion; the peak stimulation was reached 20 to 40 min after the end of the 40-min NT infusion. The dose-response curve was biphasic, with submaximal stimulation at supramaximal doses. The maximal response 359
ANNALS NEW YORK ACADEMY OF SCIENCES
360
was obtained with 1.9 nmol/kg.hr (3.16 pg/kg.hr) and represented about 30% of the maximal response to cholecystokinin. The EDsos were 170 pmol/kg.hr for sodium and bicarbonate and 250 pmol/kg.hr for protein. NT produced a dose-related increase of duodenal bicarbonate secretion. The peak stimulation was reached 30 min after beginning the 40-min NT infusion. The maximal response was obtained with 0.6 nmol/kg-hr (1 Crg/kg.hr) and represented about 80% of the maximal response to the synthetic prostaglandin enprostil. The ED50 was 60 pmol/kg.hr. Data reported in the literature suggest that NT doses found to act on duodenal and pancreatic secretion in this study may induce physiological NT plasma levels.3-5
Mechanism of Action The effect of NT on pancreatic secretion was abolished by atropine or hexamethonium; reduced (-80%) by vagotomy; reduced (-80%: sodium and bicarbonate) or abolished (protein) by L364718; unchanged by L365260 or by the association of a1 a2 fi adrenoceptor antagonists; reduced (-85%) by methadone and reduced (-70%) after capsakin pretreatment. The effect of NT on duodenal bicarbonate secretion was unchanged by atropine, naloxone, indomethacin, or L364718; reduced (-50%) by hexamethonium; reduced (-50%) by vagotomy; and reduced (-60%) after capsaicin pretreatment.
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CONCLUSION NT stimulatesexternal pancreatic secretion in the anesthetized rat with a low efficacy compared to cholecystokinin (CCK), but in a dose range compatible with postprandial plasma levels measured by others. The mechanism of action is indirect, neurally mediated, and involves nicotinic and muscarinic synapses, CCKA receptors, and in part capsakin-sensitive sensory fibers. We suggest that NT activates receptors present on peripheral afferent fibers, that this information is built up in a central reflex loop involving in part CCKA receptors, and conveyed to the pancreas by a classical cholinergic vagal efferent pathway. NT stimulates duodenal bicarbonate secretion in the anesthetized rat with a good efficacy compared to enprostil, and in a dose range compatible with postprandial plasma levels measured by others. The mechanism of action is complex. It depends for about 50% on primary sensory afferent neurons, and on vagal (sensory or motor) fibers, nicotinic synapses, and noncholinergic effector(s). The remainder of the effect is unexplained. REFERENCES 1,
2. 3. 4. 5.
RozB, C., J.
DE LA TOUR, J. CHARIOT, M. SOUCHARD & C. DEBRAY. 1975. Biol. GastroentCrol. (Paris) 8: 291-295. G. & A. GARNER. 1982. Am. J. Physiol. 242: G1834193. FLEMSTR~M, HAMMER, R. A . , R. E. HERNANDEZ & A. SHEPARD. 1990. Dig. Dis. Sci. 35: 439-443. TRIMBLE, E. R.,C. SHAW,R. BRUZZONE, A. GJINOVCI & K. D. BUCHANAN. 1987. Gastroenterology 92: 699-703. B. SYTNIK & J. WALSH.1987. J. Clin. Invest. 80: 1064-1067. MOGARD, M., V. MAXWELL,
