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The effect of gastric inhibitory polypeptide (GIP) in physiological concentrations (250, 500, 1,000 and 2,000 pg/ml) upon endocrine and exocrine secretion from the isolated perfused porcine pancreas was studied at various glucose concentrations in the perfusate. GIP increased insulin release in a dose-dependent manner. The sensitivity of the beta-cells to GIP was glucose independent. No effect was observed on glucagon or exocrine secretion regardless of the glucose concentration in the perfusate. We conclude that GIP is powerful insulin-stimulator even in low physiological concentrations in the presence of glucose concentrations comparable to those seen during an oral glucose load, which makes GIP to one of the strongest incretin candidates known, i.e. the factor(s) contributing to the augmented insulin response after ingestion of glucose.

The effect of gastrin-releasing peptide (GRP) on pancreatic exocrine secretion was studied by infusing it at four dose levels (0.01, 0.1, 1.0, and 10 nmol/l) into the arterial line of the isolated perfused porcine pancreas. At 1.0 nmol/l GRP stimulated protein (37-fold), fluid (13-fold), and bicarbonate secretion (12-fold). Atropine at 1 mumol/l diminished the protein secretion in response to infusion of GRP at a dose of 1 nmol/l to 45% of control. Fluid and bicarbonate responses were not affected by atropine treatment. Electrical stimulation of the vagus nerves resulted in an increase in pancreatic output of GRP and a concomitant stimulation of exocrine secretion. Infusions of acetylcholine, carbachol, pilocarpine, or dimethylphenylpiperazinium had no effect on the output of GRP, although hexamethonium abolished the response to vagal stimulation. It is concluded that GRP in conjunction with acetylcholine is likely to play a prominent part in parasympathetic regulation of pancreatic exocrine secretion.

The effect of pure natural porcine secretin on endocrine and exocrine pancreatic secretion was studied in the totally isolated perfused porcine pancreas. The exocrine pancreatic responses to secretin correspond well with those obtained in the anesthetized pig. The lowest concentration of secretin observed to increase pancreatic secretion was 2.8 pmol/liter, whereas the maximum pancreatic responses were obtained at a secretin concentration of 92 pmol/liter. The infusion of secretin in concentrations ranging from 2.8 to 278 pmol/liter in the presence of a constant concentration of glucose (7.5, 5.0, or 3.5 mmol/liter) was without effect on the insulin and glucagon release. Infusion of secretin at a concentration of 834 pmol/liter in the presence of glucose at 7.5 mmol/liter provoked a significant (P less than 0.01) short-lived increase in insulin secretion. However, there was no effect on the glucagon secretion. The results of this study indicate that neither the augmented insulin response nor the suppression of glucagon elicited by oral glucose depend on secretin.

THE EFFECT OF EFFERENT, PARASYMPATHETIC STIMULATION UPON PANCREATIC POLYPEPTIDE (PP) SECRETION WAS STUDIED IN THREE WAYS: (a) Plasma PP concentrations increased in response to insulin-induced hypoglycemia in both normal subjects, from 11 pM (9.5-12.5) to 136 pM (118-147), n = 8 (median and interquartile range) and in duodenal ulcer patients, from 33 pM (21-52) to 213 pM (157-233), n = 7. The PP response to hypoglycemia was diminished by atropine in normal subjects (P < 0.005) and completely abolished by vagotomy in the duodenal ulcer patients. (b) Electrical stimulation, 8 Hz, of the vagal nerves in anesthetized pigs induced an increase in portal PP concentrations within 30 s from 32 pM (28-39) to 285 pM (248-294), n = 12. Minimal stimulatory frequency was 0.5 Hz and maximal stimulatory frequency 8-12 Hz. Atropine inhibited the PP response to electrical stimulation. Median inhibition with 0.5 mg of atropine/kg body wt was 74%, range 31-90%, n = 6. The response was eliminated by hexamethonium. Adrenergic alpha and beta blockade did not influence the release of PP in response to vagal stimulation. (c) Acetylcholine stimulated, in a dose-dependent manner, the secretion of PP from the isolated perfused porcine pancreas, half-maximal effective dose being 0.19 muM; maximal PP output in response to 5 min stimulation was 228 pmol, range 140-342 pmol, n = 5. Atropine completely abolished this response.The results of the present study together with the previously demonstrated poor PP response to food in vagotomized patients, indicate that vagal, cholinergic stimulation is a major regulator of PP secretion.

The role of nerves that liberate vasoactive intestinal polypeptide (VIP) in the porcine pancrease as mediators of the atropine-resistant action of the vagus on flow and bicarbonate (HCO3) secretion was examined. Efferent electrical stimulation of the vagus in atropinized pigs produced a profuse flow of pancreatic juice with high HCO3 content concomitantly with a significant increase in pancreatic VIP output from 13 to 113 fmol/min. Intravenous administration of somatostatin (SRIF) during continuous electrical vagal stimulation caused a parallel suppression of the VIP release and the pancreatic fluid and HCO3 secretion to prestimulatory values. The SRIF-induced reduction in fluid and HCO3 secretion seemed to be mediated via an inhibition of the VIP release rather than through a direct effect on the exocrine cells, inasmuch as SRIF did not influence the VIP-provoked exocrine response from the in vitro isolated perfused porcine pancreas. The results support the view that VIP is transmitter in the vagally induced atropine-resistant water and HCO3 secretion from the porcine pancreas.

The secretory effect of sulfated and nonsulfated cholecystokinin octapeptide (CCK-8) was studied on the isolated perfused porcine pancreas. Both CCKs in concentrations from 10(-11) to 10(-8) mol/l in the presence of glucose (7.5, 5.0 or 3.5 mmol/l) were without effect on insulin and glucagon release. The exocrine secretion was stimulated by both CCKs in a dose-dependent manner, but sulfated CCK-8 was considerably more potent. The study shows that CCK-8, a major constituent of endogenous CCK, does not contribute to the incretin mechanism, irrespective of degree of sulfation. In contrast, CCK-8 is a potent stimulator of exocrine pancreatic secretion. For this effect sulfation is crucial.

The effects of the four main forms of gastrin (component I, gastrin-34, gastrin-17, and gastrin-14) on insulin, glucagon, and exocrine secretion were measured on the isolated perfused porcine pancreas. All gastrins were studied in concentrations ranging from 10(-11) to 10(-8) M. Depending on the glucose concentration in the perfusate, all four gastrins increased insulin or glucagon secretion in a dose-dependent manner in concentrations above 10(-10) M. These concentrations are slightly above the arterial concentrations in normal pig and man, but they correspond to gastrin concentrations measured in patients with achlorhydria and gastrinomas. The exocrine secretion was stimulated by all gastrins in a dose-dependent manner. The lowest concentrations that stimulated flow rate significantly were within the physiologic range, 10(-11) and 10(-10) M. All gastrins induced maximal flow rate at a concentration of 10(-9) M. The sulfated form of gastrin-17 had the greatest efficacy. The results indicate that all gastrins may influence the exocrine secretion under normal conditions and the endocrine secretion in diseases with endogenous hypergastrinemia.

We studied the autonomic nervous control of pancreatic somatostatin secretion using isolated perfused pig pancreases prepared with either intact vagal or splanchnic nerve supply. Electrical stimulation of the vagus nerves increased pancreatic protein output 59-fold, whereas somatostatin output decreased to 57% of prestimulatory secretion. Acetylcholine mimicked the somatostatin response to vagal stimulation, and atropine abolished the inhibition. Splanchnic nerve stimulation increased perfusion pressure up to threefold, whereas somatostatin output decreased to 68%. Phenoxybenzamine abolished the pressure response to splanchnic nerve stimulation and reversed the inhibition to a 20% increase in somatostatin output. Propranolol did not influence the inhibitory effect of splanchnic stimulation but abolished the increase seen after phenoxybenzamine. It is concluded that both divisions of the autonomic nerve supply to the pancreas are inhibitory to somatostatin secretion, but increased secretion may be brought about by a beta-adrenergic mechanism.

Three different molecular forms of cholecystokinin (CCK-39, -33, and -8) were used in concentrations from 10(-11) to 19(-8) mol/l to stimulate the endocrine and exocrine secretion from the isolated perfused porcine pancreas. During perfusion with a glucose concentration of 7.5 mmol/l CCK-39 in the highest concentration increased the insulin secretion slightly. No significant effect was observed at lower glucose concentrations (5.0 and 3.5 mmol/l). CCK-33 and -8 did not stimulate the secretion of insulin significantly, and neither of the cholecystokinins increased the secretion of glucagon and bicarbonate. All three molecular forms stimulated the secretion of fluid and protein in concentrations above 10(-11) mol/l in a dose-dependent manner and with equal potency. We conclude that the effect of these three cholecystokinins on the endocrine pancreas is without physiological significance, whereas all three are sufficiently potent to play a role in the control of pancreatic protein secretion.

The human vasoactive intestinal polypeptide (VIP) precursor contains a sequence, a peptide with an N-terminal histidine and C-terminal methionine (PHM), which is 93% homologous to the porcine intestinal peptide, (PHI) a peptide having N-terminal histidine and C-terminal isoleucine amide, suggesting that PHI could be a product of the porcine VIP precursor. If so, VIP-producing nerves would be expected to produce and release PHI in addition to VIP. We studied this in the pig pancreas. By immunohistochemistry, we identified nerve cell bodies in local ganglia and nerve fibers that were both PHI and VIP immunoreactive, innervating exocrine as well as endocrine structures. During electrical stimulation of the vagus nerve supply to the isolated perfused pig pancreas, a synchronous and approximately equimolar release of immunoreactive PHI and VIP was observed. Both responses were abolished by hexamethonium and could be copied by cholinergic agonists. Infusions of PHI and VIP at 10(-9) M stimulated the exocrine secretion of fluid and bicarbonate, and the effect of a combination of the two peptides was additive. In addition, both peptides stimulated insulin as well as glucagon secretion. By gel chromatography the VIP immunoreactivity in the venous effluent corresponded precisely to synthetic VIP, but only 24% of the PHI immunoreactivity corresponded to synthetic PHI; a larger peptide (mol wt 5,000-7,000) was responsible for the majority. The biological activity of this component is unknown. The results suggest that both VIP and PHI, released from intrapancreatic nerve fibers, participate in the parasympathetic control of pancreatic secretion.

We studied the effect of varying oxygen supply on the endocrine and exocrine secretions of the isolated perfused porcine pancreas utilizing completely synthetic perfusion medium with and without the addition of erythrocytes. With synthetic medium oxygenated to a Po2 of 500 mmHg, oxygen consumption was constant for flow rates at or above 0.5 ml x min-1 x g-1 (wet weight). Addition of erythrocytes to the medium did not increase oxygen consumption at flow rates above this level. Furthermore, the secretion of fluid, bicarbonate and protein in response to secretin and acetylcholine was not influenced by addition of erythrocytes. Similarly, the secretion of insulin, glucagon, pancreatic polypeptide, and somatostatin in response to arginine and acetylcholine was unchanged; arginine stimulated the secretion of all four peptides, whereas acetylcholine stimulated the secretion of insulin and pancreatic polypeptide and inhibited glucagon and somatostatin secretion. The results indicate that the porcine pancreas is respiring adequately, when perfused with a completely synthetic perfusion medium at flows above about 0.5 ml x min-1 x g-1 and Po2 about 500 mmHg, and that addition of erythrocytes is not necessary for the study of its secretory functions.

We studied the secretion of insulin, glucagon, and the exocrine secretion of the isolated perfused porcine pancreas in response to vasoactive intestinal polypeptide (VIP) in concentrations ranging from 30 to 18,750 pmol/liter at various concentrations of glucose in the perfusion medium. VIP stimulated the insulin and glucagon secretion in a dose-dependent manner. The response pattern was critically dependent on the glucose concentration. In the presence of a glucose concentration of 7.5 mmol/liter, VIP enhanced insulin release without affecting glucagon release. Maximal insulin release was obtained at a VIP concentration of 3,750 pmol/liter. At a glucose concentration of 5.0 or 3.5 mmol/liter, VIP enhanced glucagon release but not insulin release. VIP stimulated the exocrine secretion in a secretin-like manner. The lowest concentration of VIP observed to increase pancreatic exocrine secretion was 30 pmol/liter, whereas the maximal pancreatic exocrine responses were not obtained.

Severe fasting hypoglycemia and hypoinsulinemia occurred in a sixty-six year old woman with an intrahepatic mesenchymal tumor. An extract from the tumor was potent in inhibiting the arginine-induced glucagon secretion from the isolated perfused porcine pancreas. This observation supports the theory recently advanced that mesenchymal tumor hypoglycemia in some cases is due to a still unknown factor secreted from the tumor, which directly inhibits pancreatic glucagon secretion.

Recent investigations have shown that the functions of the gastrointestinal organs may be regulated by a complex system of paracrine cells and peptidergic nerves, in addition to the established humoral and autonomic nervous systems. In this article we discuss the possible paracrine regulation of the secretory functions of the stomach and the pancreas by somatostatin-producing D-cells. Secondly, we discuss the distribution, localization, effects and secretion of the neuropeptides VIP and 'gastrin-releasing polypeptide' (GRP or 'mammalian bombesin') applied to the stomach (GRP) and pancreas (GRP and VIP) of the pig. It is concluded, that all three peptides are capable of influencing powerfully the secretory state of these organs, and that these newer regulatory peptides probably play an important role in the integrate neurohormonal control of gastrointestinal secretion.

The presence of gastrin-releasing peptide (GRP) was studied in extracts of porcine pancreata. Gel filtration and high-pressure liquid chromatographic profiles of these extracts as monitored with both C-terminally and N-terminally directed radioimmunoassays against GRP showed pancreatic GRP to consist of one main form, namely the 27-amino acid peptide originally extracted from porcine stomach, and small amounts of a C-terminal fragment identical with the C-terminal 10-amino acid peptide. Gastrin-releasing peptide-like immunoreactivity released from the isolated perfused porcine pancreas during electrical vagal stimulation was shown by gel filtration to consist of the same two forms. By use of immunocytochemical techniques employing an antiserum directed against its N terminus, GRP was localized to varicose nerve fibers in close association with the exocrine tissue of the porcine pancreas in particular. Some fibers were found penetrating into pancreatic islets also. Immunoreactive nerve cell bodies as well as fibers were found within intrapancreatic ganglia. The potency of GRP in stimulating exocrine as well as endocrine secretion from the porcine pancreas, its presence in close contact with both acini and islets, and its release during vagal stimulation indicate that GRP may have a role in the parasympathetic regulation of endocrine and exocrine secretion from the pig pancreas.

Vasoactive intestinal polypeptide (VIP) in the pig pancreas is localized to nerves, many of which travel along the pancreatic ducts. VIP stimulates pancreatic fluid and bicarbonate secretion like secretin. Electrical vagal stimulation in the pig causes an atropine-resistant profuse secretion of bicarbonate-rich pancreatic juice. In an isolated perfused preparation of the pig pancreas with intact vagal nerve supply, electrical vagal stimulation caused an atropine-resistant release of VIP, which accurately parallelled the exocrine secretion of juice and bicarbonate. Perfusion of the pancreas with a potent VIP-antiserum inhibited the effect of vagal stimulation on the exocrine secretion. It is concluded, that VIP is responsible for (at least part of) the neurally controlled fluid and bicarbonate secretion from the pig pancreas.

The isolation of a porcine pancreas preparation without the attached duodenum is described. The preparation is suitable for study of the secretion of insulin and pancreatic glucagon. The surgical procedure by which the pancreas is removed from tha animal is decribed as well as the perfusion system. The effect of various concentrations of glucose (63, 90, 135 and 180 mg per 100 ml) in the perfusion medium on the secretion of insulin and pancreatic glucagon was investigated in order to observe the performance characteristics of this preparation. Increasing concentrations of glucose caused a significantly greater insulin release, whereas the glucagon response was significantly reduced.