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Effect of bombesin infused intrapancreatically on glucagon and insulin secretion
Effect of Bombesin Infused Intrapancreatically on Glucagon and Insulin Secretion Akio Kaneto, Toshio
Kaneko, Seishi Nakaya,
Synthetic bombesin was infused at a dose of 20 pmoler/kg/min for 10 min into the cranial pancreaticoduodenal artery of anesthetized dogs. Plasma immunoreactive glucagon concentrations in the cranial pancreaticoduodenal vein as well as in the femoral artery were concurrently and slowly elevated. However, the net release of glucagon from the pancreas did not increase significantly during infusion of bombesin. Plasma immunoreactive insulin
Hiroshi
Kajinuma,
and Kinori
Kosaka
concentrations in the pancreatic vein were transiently mised, and a delayed rise was noted in arterial plasma IRI. Net release of insulin was significantly augmented during infusion of the tetradecapeptide. Plasma glucose levels did not change after bombesin. These results indicate that the gastrointestinal tetradecapeptide may stimulate secretion of both insulin and gut glucagonlike immunoreactivity in the dog.
EPTIDES OF THE BOMBESIN FAMILY have recently been isolated I from the skin of two European frogs, Discoglossidae (Bombina bombina and Bombina variegata variegata). They have been shown to stimulate secretion of pancreatic juice rich in hydrolases in man’ and the dog,2 and gall bladder emptying2*’ and gastrin release 4s in the dog. Erspamer and Melchiorri6 discovered that some cells in the antral and duodenal mucosa of dogs and pigs contain substances that react with antibombesin antibodies, suggesting that bombesin and its relatives may be candidate hormones of the gut.6,’ According to Deschodt-Lanckman et al.,* amylase secretion and calcium efflux from the rat pancreas are stimulated in vitro by peptides of the bombesin family. The authors have already observed some pancreatotropic action of several gastrointestinal polypeptides or their derivatives such as tetragastrin,’ vasoactive intestinal polypeptide (VIP),‘O and substance P.” We therefore decided to explore the direct action of synthetic bombesin infused into the pancreatic artery upon the function of the islets of Langerhans. D
MATERIALS
AND
METHODS
Six mongrel dogs weighing 16.2-24.6 kg were used under pentobarbital anesthesia after an overnight fast. After midline laparotomy, an extracorporeal pancreatic blood circuit was made between the cranial pancreaticoduodenal vein and portal vein. as previously described.12 Cannulation of the cranial pancreaticoduodenal artery and femoral artery was performed 3 hr prior to the initial blood sampling, as previously reported.13 The right gastroepiploic vessels were ligated at the pyloric level, and the abdominal wall was kept closed until completion of the experiment. Pancreatic vein blood flow was measured by collecting timed blood samples after interrupting the extracorporeal shunt and adjusting pressure in the pancreatic catheter back to portal venous pressure using the portal cannula as a manometer.
From the Third Departmeni of Internal Medicine and the First Deparrment of Internal Medicine, Faculty of Medicine, University of Tokyo. Tokyo, and the First Department of Surgery, Kobe University School of Medicine, Kobe, Japan. Received/or publication June 9. 1977. Address reprinl requests to Dr. Akio Kaneto, Third Department of Internal Medicine, Faculty of Medicine, University of Tokyo, Tokyo, Japan 113. 0 1978 by Prune & Stratton. Inc. 00260495/78/2705-0008$01.00/0 Metabolism, Vol. 27, No. 5 (May), 1978
549
550
Physiologic
KANETO
ET AL.
saline solution
was infused (1 ml/ min) as a control substance, over a period of 3 hr, pancreaticoduodenal artery. At 30 min and again at 1 IO min after initiation of the infusion,synthetic bombesin* was added to the infusion for a period of 10 min at a dose of 20 pmoles/kg/min. The bombesin solution was prepared immediately before use. Blood samples were obtained -6, -3(F) or F2). 0, 3, 6, and 10 min after the start of each infusion of bombesin, and 5, 10, 30, and 60 min after the cessation. The sampling at 0 min was done during the initial 15-30 set of the infusion.
into the cranial
Plasma immunoreactive glucagon (IRG) levels were assayed by a slight modification’4 of the chromatographic procedure of Orskov et al.” Two antiglucagon sera were used. One antiserum reacted with pork gut glucagonlike immunoreactivity (GLI) as well as with pancreatic glucagon. The other was the highly specific antiserum 30 K from the laboratory of RH. Unger (pool 2, lot 38). Plasma immunoreactive insulin (IRI) levels were measured by a modification of the using dog insulin (Novo Research Institute, Copenhagen; method of Morgan and Lazarow16 lot H6264) as a standard. The addition of bombesin had no effect on the standard immunoassay curves for glucagon and insulin. The hormone output in pancreatic venous plasma was calculated as the product of individual hormone concentrations and plasma flow in the samples. The net release of pancreatic glucagon and insulin was calculated as the product of the individual differences of femoral arterial plasma hormone concentrations from those in pancreatic venous plasma and corresponding plasma flow. Plasma glucose concentrations in the femoral artery were determined by a glucose oxidase method.17 The Wilcoxon signed-rank test was employed for the statistical analysis of the difference of means in paired samples.
RESULTS
The mean pancreatic venous blood flow showed a transitory fall immediately after initiation of the first infusion of bombesin and a significant increase during the second infusion (Fig. 1). Average pancreatic venous hematocrit did not change during each infusion, and mean pancreatic plasma flow changed along with the blood flow. Mean pancreatic venous plasma concentrations of IRG assayed with the nonspecific antiserum were elevated during and after both infusions, not declining to the baselines within 1 hr. These values fluctuated in parallel with the plasma concentrations of IRG in the femoral artery. Mean pancreatic venous plasma IRG determined with the 30 K antiserum showed a delayed significant rise at the end of and after the infusions. Administration of bombesin, however, failed to enhance the average net release of glucagon measured with each antiserum into the cranial pancreaticoduodenal vein (Fig. 2). Basal net release increased gradually during the course of the experiment. Mean pancreatic effluent plasma concentrations of IRI rose significantly during both infusions; its output also increased, returning to the preinfusion levels within 10 min after cessation of the infusions (Fig. 1). Average net release of insulin into the pancreatic vein was significantly augmented during the infusions, in contrast to that of glucagon (Fig. 2). The femoral arterial plasma IRI was raised significantly to a peak at the end of each infusion of bombesin. Femoral arterial plasma glucose did not change throughout the experiment. DISCUSSION
The present data show that bombesin, a new tetradecapeptide of the gut, is able to stimulate insulin secretion without augmenting release of pancreatic glucagon when it is directly infused into the pancreatic artery. In this respect, *Batch YH83, synthesized by Prof. Noboru Yanaibara, Shizuoka College of Pharmacy, Shizuoka. Japan.
Department
of Bio-organic
Chemistry,
INTRAPANCREATICALLY-INFUSED
BOMBESIN
2000 _:
1500-
x,,-
Fig. 1. Effect of bombesin infused into the cranial pancreaticoduodenal artery at a dose of 20 pmoles/kg/min on the levels of femoral arterial plasma glucose, pancreatic vein blood Row, and plasma IRG and IRI. * , Statistically significant (p < 0.05), preinfusion values, designated FI or Fz, versus postinfusion values; o, IRG concentrations measured with the nonspecific antiserum; t, sign&ant ( p < 0.05) IRG concentrations determined with the 30 K antiserum ( l) .
Pancreatic
Venous
Plasma
! i +
Pancreatic
Venous
Plasma
bombesin is different from other gastrointestinal polypeptides such as gastrin18 or its relative, tetragastriq9 pancreozymin-cholecystokinin,lg and VIP,‘“,20 all of which have been shown to be capable of stimulating both pancreatic glucagon and insulin in the dog. The significant elevation of the arterial plasma IRG assayed with the non-
Fig. 2. Effect of bombesin infused intrapancreatically on net release of glucagon and insulin into the cranial pancreaticoduodenal vein. *, Statistically significant (p i 0.05), preinfusion values, designated Ft or Fs, versus postinfusion values of IRI and IRG (0) (measured with the nonspecific antiglucagon serum).
552
KANETO
ET AL.
specific antiserum seen during infusions of bombesin in the absence of any augmentation in net release of pancreatic glucagon strongly indicates an increase of plasma GLI of extrapancreatic, probably gastrointestinal, origin. Glucagon could not be detected in the blood of dogs that had undergone comTherefore, the origins of extrapancreatic gluplete abdominal evisceration.2’ cagon are probably abdominal organs; other significant sources of glucagon, such as the salivary glands (as reported in rodents22,23), are unlikely to exist in this species. None of the gut hormones have been documented as being able to stimulate release of gut GLI. The absence of any significant increase in net release of pancreatic glucagon during bombesin infusions was confirmed by measurement with the highly specific antiglucagon serum. However, a further delayed rise of plasma IRG in both the pancreatic vein and the femoral artery was seen after the infusions, even as determined with the 30 K antiserum. This finding might well be explained by the possible participation of some gastrointestinal sources, such as the gastric oxyntic mucosa releasing IRG reacting with highly specific antisera,24-28 and by the minor cross-reactivity of the 30 K antiserum with the elevated gut GLI. The gradual increase in the basal release of pancreatic glucagon throughout the course of the experiment may be a stress reaction to the longlasting experimental procedures. The insulin release did not increase immediately upon local administration of bombesin. This suggests that the bombesin effect upon insulin secretion may be an indirect one. It is possible that the fi cell is affected through the mediation of some gut hormones or factors released by bombesin. The arterial plasma glucose level did not change, irrespective of this gradual enhancement of glucagon secretion. It is not clear whether the increase in insulin secretion that occurred in response to administration of bombesin may contribute to maintaining normoglycemia. Most gastrointestinal hormones with a cytotropic effect on the islets of Langerhans have been reported to have common vasoactive properties, raising the possibility that the effect is mediated through increased blood flow per unit mass of the pancreas rather than a direct effect on the cy and p cells. It might be worthy of mention that secretion of insulin was stimulated in the absence of any increase in pancreatic blood flow, as seen during the first infusion of bombesin, and that secretion of pancreatic glucagon was not stimulated even in the presence of the flow augmentation, as seen during the second infusion. ACKNOWLEDGMENT The authors are grateful to Prof. Noboru Yanaibara for his generous donation of synthetic bombesin. We are indebted to Yukichi Kohga, Shimizu Pharmaceutical Company, and Toshiko Kojima for their technical assistance. We wish to think Dr. Yasunori Kanazawa and Dr. Masaki Hayashi for their valued advice.
REFERENCES I. Basso N. Lezoche
E. Percoco M, et al: Secrezione pancreatica esterna dopo infusione di bombesina nell’uomo. Rend Gastroenterol [Suppl] 7:72, 1975 2. Erspamer V, lmprota G, Melchiorri P. et
al: Evidence of cholecystokinin release by bombesin in the dog. Br J Pharmacol 52:227232. 1974 3. Corazziari E, Delle Fave GF, Melchiorri P. et al: Effects of a new polypeptide, bombesin,
INTRAPANCREATICALLY-INFUSED
BOMBESIN
on gall bladder and duodenojejunal mechanical activity in man. Rend Gastroenterol 5:140. 1973 4. Bertaccini G, Erspamer V, Melchiorri P, et al: Gastrin release by bombesin in the dog. Br J Pharmacol52:219-225. 1974 5. Impicciatore M, Debas H. Walsh JH, et al: Release of gastrin and stimulation of acid secretion by bombesin in dog. Rend Gastroenterol 6:99-101, 1974 6. Erspamer V. Melchiorri P: Actions of bombesin on secretions and motility of the gastrointestinal tract. in Thompson JC (ed): Gastrointestinal Hormones. Austin. University of Texas Press, 1975. pp 575-589 7. Grossman MI: Additional candidate hormones of the gut. Gastroenterology 69: 570 571,1975 8. Deschodt-Lanckman M. Robberecht P, DeNeef P, et al: In vitro action of bombesin and bombesin-like peptides on amylase secrecyclase tion, calcium efflux, and adenylate activity in the rat pancreas. J Clin Invest 58: 891 898, 1976 9. Kaneto A. Mizuno Y, Tasaka Y, et al: Stimulation of glucagon secretion by tetragastrin. Endocrinology 86: I I75 1180,1970 IO. Kaneto A, Kaneko T. Kajinuma H. et al: Effect of VIP infused intrapancreatically on glucagon and insulin secretion. Metabolism 26:781 ~786. 1977 I I. Kaneto A, Kaneko T, Kajinuma H, et al: Effect of substance P and neurotensin infused intrapancreatically on glucagon and insulin secretion. Endocrinology (in press) 12. Kaneto A, Kosaka K: Stimulation of glucagon and insulin secretion by acetylcholine infused intrapancreatically. Endocrinology 95: 676-68 I, 1974 13. Kaneto A, Kajinuma H, Kosaka K, et al: Stimulation of insulin secretion by parasympathomimetic agents. Endocrinology 83:65ll 658, 1968 14. Kaneto A. Kosaka K: Stimulation of glucagon secretion by oxytocin. Endocrinology 87:439-444, 1970 15. @rskov H. Thomsen HG, Yde H: Wick chromatography for rapid and reliable immunoassay of insulin, glucagon and growth hormone. Nature 219:193-195. 1968
553
16. Morgan CR, Lazarow A: Immunoassay of insulin: Two antibody system. Plasma insulin levels of normal, subdiabetic and diabetic rats. Diabetes l2:I 15-126, 1963 17. Washko ME, Rice EW: Determination of glucose by an improved enzymatic procedure. Clin Chem 7:5422545, 1961 IS. Iversen J: Secretion of glucagon and insulin from the isolated, perfused canine pancreas. J Clin Invest SO:2 12332 136, 197 I 19. Unger RH, Ketterer H, Duprt J, et al: The effects of secretin, pancreozymin, and gastrin on insulin and glucagon secretion in anesthetized dogs. J Clin Invest 46:630-645. 1967 20. Schebalin M. Said SI, Makhlouf GM: Stimulation of insulin and glucagon secretion by vasoactive intestinal polypeptide. Am J Physiol232:El97-E200, 1977 21. Lefebvre PJ, Luyckx AS: Plasma glucagon after kidney exclusion: Experiments in somatostatin-infused and in eviscerated dogs. Metabolism 25:761l768. 1976 22. Lawrence AM, Kirsteins L. Hojvat S. et al: Submaxillary gland hyperglycemic factor in man and animals; an extrapancreatic glucagon. Clin Res 24:364A. 1976 (abstract) 23. Dunbar JC, Silverman H. Kirman E, et al: Salivary gland and kidney glucagon in the rat. Fed Proc 35:218, 1976 (abstract) 24. Vranic M, Pek S, Kawamori R: In“glucagon immunoreactivity” in creased plasma of totally depancreatized dogs. Diabetes 23:905-912. 1974 25. Matsuyama T, FOB PP: Plasma glucose, insulin, pancreatic, and enteroglucagon levels in normal and depancreatized dogs. Proc Sot Exp Biol Med 147:97-102, 1974 26. Mashiter K. Harding PE. Chou M. et al: Persistent pancreatic glucagon but not insulin response to arginine in pancreatectomized dogs. Endocrinology 96:678-693, 1975 27. Sasaki H. Rubalcava B. Baetens D, et al: Identification of glucagon in the gastrointestinal tract. J Clin Invest 56:135-145. 1975 28. Baetens D. Rufener C, Srikant CB. et al: Identification of glucagon-producing cells (A cells) in dog gastric mucosa. J Cell Biol 69: 455-464. 1976
Kaneko, Seishi Nakaya,
Synthetic bombesin was infused at a dose of 20 pmoler/kg/min for 10 min into the cranial pancreaticoduodenal artery of anesthetized dogs. Plasma immunoreactive glucagon concentrations in the cranial pancreaticoduodenal vein as well as in the femoral artery were concurrently and slowly elevated. However, the net release of glucagon from the pancreas did not increase significantly during infusion of bombesin. Plasma immunoreactive insulin
Hiroshi
Kajinuma,
and Kinori
Kosaka
concentrations in the pancreatic vein were transiently mised, and a delayed rise was noted in arterial plasma IRI. Net release of insulin was significantly augmented during infusion of the tetradecapeptide. Plasma glucose levels did not change after bombesin. These results indicate that the gastrointestinal tetradecapeptide may stimulate secretion of both insulin and gut glucagonlike immunoreactivity in the dog.
EPTIDES OF THE BOMBESIN FAMILY have recently been isolated I from the skin of two European frogs, Discoglossidae (Bombina bombina and Bombina variegata variegata). They have been shown to stimulate secretion of pancreatic juice rich in hydrolases in man’ and the dog,2 and gall bladder emptying2*’ and gastrin release 4s in the dog. Erspamer and Melchiorri6 discovered that some cells in the antral and duodenal mucosa of dogs and pigs contain substances that react with antibombesin antibodies, suggesting that bombesin and its relatives may be candidate hormones of the gut.6,’ According to Deschodt-Lanckman et al.,* amylase secretion and calcium efflux from the rat pancreas are stimulated in vitro by peptides of the bombesin family. The authors have already observed some pancreatotropic action of several gastrointestinal polypeptides or their derivatives such as tetragastrin,’ vasoactive intestinal polypeptide (VIP),‘O and substance P.” We therefore decided to explore the direct action of synthetic bombesin infused into the pancreatic artery upon the function of the islets of Langerhans. D
MATERIALS
AND
METHODS
Six mongrel dogs weighing 16.2-24.6 kg were used under pentobarbital anesthesia after an overnight fast. After midline laparotomy, an extracorporeal pancreatic blood circuit was made between the cranial pancreaticoduodenal vein and portal vein. as previously described.12 Cannulation of the cranial pancreaticoduodenal artery and femoral artery was performed 3 hr prior to the initial blood sampling, as previously reported.13 The right gastroepiploic vessels were ligated at the pyloric level, and the abdominal wall was kept closed until completion of the experiment. Pancreatic vein blood flow was measured by collecting timed blood samples after interrupting the extracorporeal shunt and adjusting pressure in the pancreatic catheter back to portal venous pressure using the portal cannula as a manometer.
From the Third Departmeni of Internal Medicine and the First Deparrment of Internal Medicine, Faculty of Medicine, University of Tokyo. Tokyo, and the First Department of Surgery, Kobe University School of Medicine, Kobe, Japan. Received/or publication June 9. 1977. Address reprinl requests to Dr. Akio Kaneto, Third Department of Internal Medicine, Faculty of Medicine, University of Tokyo, Tokyo, Japan 113. 0 1978 by Prune & Stratton. Inc. 00260495/78/2705-0008$01.00/0 Metabolism, Vol. 27, No. 5 (May), 1978
549
550
Physiologic
KANETO
ET AL.
saline solution
was infused (1 ml/ min) as a control substance, over a period of 3 hr, pancreaticoduodenal artery. At 30 min and again at 1 IO min after initiation of the infusion,synthetic bombesin* was added to the infusion for a period of 10 min at a dose of 20 pmoles/kg/min. The bombesin solution was prepared immediately before use. Blood samples were obtained -6, -3(F) or F2). 0, 3, 6, and 10 min after the start of each infusion of bombesin, and 5, 10, 30, and 60 min after the cessation. The sampling at 0 min was done during the initial 15-30 set of the infusion.
into the cranial
Plasma immunoreactive glucagon (IRG) levels were assayed by a slight modification’4 of the chromatographic procedure of Orskov et al.” Two antiglucagon sera were used. One antiserum reacted with pork gut glucagonlike immunoreactivity (GLI) as well as with pancreatic glucagon. The other was the highly specific antiserum 30 K from the laboratory of RH. Unger (pool 2, lot 38). Plasma immunoreactive insulin (IRI) levels were measured by a modification of the using dog insulin (Novo Research Institute, Copenhagen; method of Morgan and Lazarow16 lot H6264) as a standard. The addition of bombesin had no effect on the standard immunoassay curves for glucagon and insulin. The hormone output in pancreatic venous plasma was calculated as the product of individual hormone concentrations and plasma flow in the samples. The net release of pancreatic glucagon and insulin was calculated as the product of the individual differences of femoral arterial plasma hormone concentrations from those in pancreatic venous plasma and corresponding plasma flow. Plasma glucose concentrations in the femoral artery were determined by a glucose oxidase method.17 The Wilcoxon signed-rank test was employed for the statistical analysis of the difference of means in paired samples.
RESULTS
The mean pancreatic venous blood flow showed a transitory fall immediately after initiation of the first infusion of bombesin and a significant increase during the second infusion (Fig. 1). Average pancreatic venous hematocrit did not change during each infusion, and mean pancreatic plasma flow changed along with the blood flow. Mean pancreatic venous plasma concentrations of IRG assayed with the nonspecific antiserum were elevated during and after both infusions, not declining to the baselines within 1 hr. These values fluctuated in parallel with the plasma concentrations of IRG in the femoral artery. Mean pancreatic venous plasma IRG determined with the 30 K antiserum showed a delayed significant rise at the end of and after the infusions. Administration of bombesin, however, failed to enhance the average net release of glucagon measured with each antiserum into the cranial pancreaticoduodenal vein (Fig. 2). Basal net release increased gradually during the course of the experiment. Mean pancreatic effluent plasma concentrations of IRI rose significantly during both infusions; its output also increased, returning to the preinfusion levels within 10 min after cessation of the infusions (Fig. 1). Average net release of insulin into the pancreatic vein was significantly augmented during the infusions, in contrast to that of glucagon (Fig. 2). The femoral arterial plasma IRI was raised significantly to a peak at the end of each infusion of bombesin. Femoral arterial plasma glucose did not change throughout the experiment. DISCUSSION
The present data show that bombesin, a new tetradecapeptide of the gut, is able to stimulate insulin secretion without augmenting release of pancreatic glucagon when it is directly infused into the pancreatic artery. In this respect, *Batch YH83, synthesized by Prof. Noboru Yanaibara, Shizuoka College of Pharmacy, Shizuoka. Japan.
Department
of Bio-organic
Chemistry,
INTRAPANCREATICALLY-INFUSED
BOMBESIN
2000 _:
1500-
x,,-
Fig. 1. Effect of bombesin infused into the cranial pancreaticoduodenal artery at a dose of 20 pmoles/kg/min on the levels of femoral arterial plasma glucose, pancreatic vein blood Row, and plasma IRG and IRI. * , Statistically significant (p < 0.05), preinfusion values, designated FI or Fz, versus postinfusion values; o, IRG concentrations measured with the nonspecific antiserum; t, sign&ant ( p < 0.05) IRG concentrations determined with the 30 K antiserum ( l) .
Pancreatic
Venous
Plasma
! i +
Pancreatic
Venous
Plasma
bombesin is different from other gastrointestinal polypeptides such as gastrin18 or its relative, tetragastriq9 pancreozymin-cholecystokinin,lg and VIP,‘“,20 all of which have been shown to be capable of stimulating both pancreatic glucagon and insulin in the dog. The significant elevation of the arterial plasma IRG assayed with the non-
Fig. 2. Effect of bombesin infused intrapancreatically on net release of glucagon and insulin into the cranial pancreaticoduodenal vein. *, Statistically significant (p i 0.05), preinfusion values, designated Ft or Fs, versus postinfusion values of IRI and IRG (0) (measured with the nonspecific antiglucagon serum).
552
KANETO
ET AL.
specific antiserum seen during infusions of bombesin in the absence of any augmentation in net release of pancreatic glucagon strongly indicates an increase of plasma GLI of extrapancreatic, probably gastrointestinal, origin. Glucagon could not be detected in the blood of dogs that had undergone comTherefore, the origins of extrapancreatic gluplete abdominal evisceration.2’ cagon are probably abdominal organs; other significant sources of glucagon, such as the salivary glands (as reported in rodents22,23), are unlikely to exist in this species. None of the gut hormones have been documented as being able to stimulate release of gut GLI. The absence of any significant increase in net release of pancreatic glucagon during bombesin infusions was confirmed by measurement with the highly specific antiglucagon serum. However, a further delayed rise of plasma IRG in both the pancreatic vein and the femoral artery was seen after the infusions, even as determined with the 30 K antiserum. This finding might well be explained by the possible participation of some gastrointestinal sources, such as the gastric oxyntic mucosa releasing IRG reacting with highly specific antisera,24-28 and by the minor cross-reactivity of the 30 K antiserum with the elevated gut GLI. The gradual increase in the basal release of pancreatic glucagon throughout the course of the experiment may be a stress reaction to the longlasting experimental procedures. The insulin release did not increase immediately upon local administration of bombesin. This suggests that the bombesin effect upon insulin secretion may be an indirect one. It is possible that the fi cell is affected through the mediation of some gut hormones or factors released by bombesin. The arterial plasma glucose level did not change, irrespective of this gradual enhancement of glucagon secretion. It is not clear whether the increase in insulin secretion that occurred in response to administration of bombesin may contribute to maintaining normoglycemia. Most gastrointestinal hormones with a cytotropic effect on the islets of Langerhans have been reported to have common vasoactive properties, raising the possibility that the effect is mediated through increased blood flow per unit mass of the pancreas rather than a direct effect on the cy and p cells. It might be worthy of mention that secretion of insulin was stimulated in the absence of any increase in pancreatic blood flow, as seen during the first infusion of bombesin, and that secretion of pancreatic glucagon was not stimulated even in the presence of the flow augmentation, as seen during the second infusion. ACKNOWLEDGMENT The authors are grateful to Prof. Noboru Yanaibara for his generous donation of synthetic bombesin. We are indebted to Yukichi Kohga, Shimizu Pharmaceutical Company, and Toshiko Kojima for their technical assistance. We wish to think Dr. Yasunori Kanazawa and Dr. Masaki Hayashi for their valued advice.
REFERENCES I. Basso N. Lezoche
E. Percoco M, et al: Secrezione pancreatica esterna dopo infusione di bombesina nell’uomo. Rend Gastroenterol [Suppl] 7:72, 1975 2. Erspamer V, lmprota G, Melchiorri P. et
al: Evidence of cholecystokinin release by bombesin in the dog. Br J Pharmacol 52:227232. 1974 3. Corazziari E, Delle Fave GF, Melchiorri P. et al: Effects of a new polypeptide, bombesin,
INTRAPANCREATICALLY-INFUSED
BOMBESIN
on gall bladder and duodenojejunal mechanical activity in man. Rend Gastroenterol 5:140. 1973 4. Bertaccini G, Erspamer V, Melchiorri P, et al: Gastrin release by bombesin in the dog. Br J Pharmacol52:219-225. 1974 5. Impicciatore M, Debas H. Walsh JH, et al: Release of gastrin and stimulation of acid secretion by bombesin in dog. Rend Gastroenterol 6:99-101, 1974 6. Erspamer V. Melchiorri P: Actions of bombesin on secretions and motility of the gastrointestinal tract. in Thompson JC (ed): Gastrointestinal Hormones. Austin. University of Texas Press, 1975. pp 575-589 7. Grossman MI: Additional candidate hormones of the gut. Gastroenterology 69: 570 571,1975 8. Deschodt-Lanckman M. Robberecht P, DeNeef P, et al: In vitro action of bombesin and bombesin-like peptides on amylase secrecyclase tion, calcium efflux, and adenylate activity in the rat pancreas. J Clin Invest 58: 891 898, 1976 9. Kaneto A. Mizuno Y, Tasaka Y, et al: Stimulation of glucagon secretion by tetragastrin. Endocrinology 86: I I75 1180,1970 IO. Kaneto A, Kaneko T. Kajinuma H. et al: Effect of VIP infused intrapancreatically on glucagon and insulin secretion. Metabolism 26:781 ~786. 1977 I I. Kaneto A, Kaneko T, Kajinuma H, et al: Effect of substance P and neurotensin infused intrapancreatically on glucagon and insulin secretion. Endocrinology (in press) 12. Kaneto A, Kosaka K: Stimulation of glucagon and insulin secretion by acetylcholine infused intrapancreatically. Endocrinology 95: 676-68 I, 1974 13. Kaneto A, Kajinuma H, Kosaka K, et al: Stimulation of insulin secretion by parasympathomimetic agents. Endocrinology 83:65ll 658, 1968 14. Kaneto A. Kosaka K: Stimulation of glucagon secretion by oxytocin. Endocrinology 87:439-444, 1970 15. @rskov H. Thomsen HG, Yde H: Wick chromatography for rapid and reliable immunoassay of insulin, glucagon and growth hormone. Nature 219:193-195. 1968
553
16. Morgan CR, Lazarow A: Immunoassay of insulin: Two antibody system. Plasma insulin levels of normal, subdiabetic and diabetic rats. Diabetes l2:I 15-126, 1963 17. Washko ME, Rice EW: Determination of glucose by an improved enzymatic procedure. Clin Chem 7:5422545, 1961 IS. Iversen J: Secretion of glucagon and insulin from the isolated, perfused canine pancreas. J Clin Invest SO:2 12332 136, 197 I 19. Unger RH, Ketterer H, Duprt J, et al: The effects of secretin, pancreozymin, and gastrin on insulin and glucagon secretion in anesthetized dogs. J Clin Invest 46:630-645. 1967 20. Schebalin M. Said SI, Makhlouf GM: Stimulation of insulin and glucagon secretion by vasoactive intestinal polypeptide. Am J Physiol232:El97-E200, 1977 21. Lefebvre PJ, Luyckx AS: Plasma glucagon after kidney exclusion: Experiments in somatostatin-infused and in eviscerated dogs. Metabolism 25:761l768. 1976 22. Lawrence AM, Kirsteins L. Hojvat S. et al: Submaxillary gland hyperglycemic factor in man and animals; an extrapancreatic glucagon. Clin Res 24:364A. 1976 (abstract) 23. Dunbar JC, Silverman H. Kirman E, et al: Salivary gland and kidney glucagon in the rat. Fed Proc 35:218, 1976 (abstract) 24. Vranic M, Pek S, Kawamori R: In“glucagon immunoreactivity” in creased plasma of totally depancreatized dogs. Diabetes 23:905-912. 1974 25. Matsuyama T, FOB PP: Plasma glucose, insulin, pancreatic, and enteroglucagon levels in normal and depancreatized dogs. Proc Sot Exp Biol Med 147:97-102, 1974 26. Mashiter K. Harding PE. Chou M. et al: Persistent pancreatic glucagon but not insulin response to arginine in pancreatectomized dogs. Endocrinology 96:678-693, 1975 27. Sasaki H. Rubalcava B. Baetens D, et al: Identification of glucagon in the gastrointestinal tract. J Clin Invest 56:135-145. 1975 28. Baetens D. Rufener C, Srikant CB. et al: Identification of glucagon-producing cells (A cells) in dog gastric mucosa. J Cell Biol 69: 455-464. 1976