Robert L. Hazelwood

The Pancreatic Polypeptide (PP-Fold) Family: Gastrointestinal, Vascular, and Feeding Behavioral Implications

Abstract Historical Background. Of the four recognized peptide hormones secreted by the endocrine pancreas, pancreatic polypeptide (PP) was the third to be discovered, isolated, and characterized. Insulin (1921-22) and glucagon (1923) preceded and somatostatin (1975) followed the isolation of PP in 1968. Despite this sequence of events—and the time interval since its discovery—PP remains somewhat of an enigma both in name and in function. The reluctance to label this polypeptide with a more descriptive title than the lackluster PP it bears stems largely from the fact that while broad actions may be attributed to it, no outstanding or dominating effect has been identified with its actions. Thus, the only characteristic carried by use of the phrase PP is to signify the species of PP one is discussing, such as avian, bovine, rat, porcine, ovine, and human, for example. The avian structure has 15 identities (of a total of 36) with that of bovine PP, and the latter differs by one or two residues from most other mammalian PP at any of three or four positions within the molecule. Thus, strong structural conservatism exists from an evolutionary point of view.

Spectrum effects of a new polypeptide (third hormone?) isolated from the chicken pancreas.

Abstract A broad evaluation was made of the biological effects of a new polypeptide (APP), 36 amino acid residues, MW 4200, isolated from the chicken pancreas. APP is an effective hepatic glycogenolytic and plasma hypoglycerolemic agent in the absence of plasma glucose perturbation. APP (at much lower doses) is also a powerful gastric secretogogue, inducing within seconds marked increases in proventricular volume, H + , pepsin, and total protein release. Such proventricular response is not mediated through systemic cardiovascular alterations or the vagus nerves, and are quite opposite to those observed with beef or chicken glucagon. It is concluded that the chicken pancreas releases APP to exert a “gastrinlike” secretogogic action on the proventriculus and, at higher concentrations, a metabolic action as well.

In vitro insulin release from chicken pancreas

Abstract Insulin release from the chicken pancreas (adult hens) was studied in vitro by incubating pancreatic pieces (20 ± 5 mg) for four successive 15-min experimental periods preceded by a 30-min preincubation period. Release of immunoreactive insulin (IRI) into the medium was measured in response to glucose (50, 150, 300, 500, and 700 mg/dl), sulfonylureas, and in the presence or absence of extracellular calcium. The results obtained demonstrate that glucose stimulated insulin release only at concentrations of 500 and 700 mg/dl. At these concentrations, the rate of insulin release was maintained at approximately twice basal levels for only 15 to 30 min before returning to the preincubation value (45 ng/g/min) irrespective of the prolonged (1-hr) glucose stimulation. Glucagon (5 μg/ml) effected a twofold increase in the rate of insulin release in the presence of 50 mg of glucose/dl of medium. Three concentrations (3.1, 6.1, and 12.2 mg/dl) of tolbutamide stimulated a twofold increase of insulin release; however, these avian pancreatic pieces were refractory to a second tolbutamide stimulus. In contrast to mammals, the major metabolite of tolbutamide, carboxytolbutamide, was inert in this system. Glucose- and tolbutamide-induced insulin secretion both were inhibited in the absence of extracellular calcium; this inhibition was abolished by replacement of normal amounts of calcium in the incubation medium. The results of this study suggest that the chicken β cell differs from the mammalian counterpart by being insensitive to all but very high glucose concentrations, and by being incapable of a sustained increase in insulin release in response to prolonged or repetitive insulinotropic stimuli. Additionally, the insulin response to tolbutamide in vitro indicates that the insulin releasing action of this compound may underlie the hypoglycemic effect observed in birds. Dependence on extracellular calcium suggests that the functional mechanism for avian insulin release may be similar in birds and mammals, although birds may differ from mammals in their pattern of insulin release.

Effects of diabetes mellitus on lactation in the rat

Abstract Diabetes mellitus adversely affects the process of lactation. Although insulin administration restores lactation, the manner by which it does so is unknown. The goals of this study were to determine which phase of lactation, milk synthesis/supply (MS), and/or milk release (MR) was affected. Inasmuch as insulin, corticosterone, and prolactin, among other hormones, are involved in milk synthesis in vitro, this study investigated their probable roles in the suppression of lactation in diabetes in vivo. Diabetes was induced in rats on Day 3 postpartum with streptozotocin (50-60 mg/kg, ip). Milk synthesis/supply and milk release were indirectly monitored using body weight gain of pups during a timed-feeding period on postnatal Days 8/9 and 13/14. Plasma corticosterone, insulin, C-peptide, and prolactin were measured by radioimmunoassay in control, diabetic, and insulin-replaced diabetic animals. Insulin replacement was provided by means of an osmotic minipump implanted subcutaneously in the nape of the neck. In addition, several parameters of maternal behavior were monitored in order to determine whether diabetes affected maternal behavior and, therefore, whether such changes played a role in the alterations of lactation observed during diabetes. Diabetes significantly suppressed MS and MR. However, the decrease in MR, which was restored after partial insulin replacement, was a reflection of the reduced MS. There were no significant differences between the parameters of maternal behavior monitored in control and diabetic animals. Blood glucose in diabetic dams was significantly increased over that of controls. Although the levels of plasma glucose in the insulin-replaced groups were significantly lower than those of their uncontrolled diabetic counterparts, they also remained higher than that of controls. Corticosterone was not significantly altered after induction of diabetes. Insulin and C-peptide levels were significantly reduced in the uncontrolled diabetics; however, insulin levels were corrected in the insulin-replaced groups. Serum levels of prolactin decreased in all diabetic groups and insulin failed to restore these levels to those of control animals. In conclusion, it appears that diabetes decreases lactation through a suppressive effect on MS rather than on MR, with insulin replacement correcting for such deficiency. In addition, despite the lactogenic importance of glucocorticoids, prolactin and insulin demonstrated in vitro, lactation in vivo can be corrected without returning the levels of all three hormones to normal. The importance of another factor(s), such as normoglycemia, as essential to the observed defects, needs to be investigated.

Pancreatectomy in the chicken: Does an extra-pancreatic source of insulin exist?

Abstract The possibility of an extrapancreatic source of insulin in aves was studied in adult Single Comb White Leghorn chickens. The effect of partial pancreatectomy (99%) on plasma glucose and immunoreactive insulin (IRI) levels, as well as the tolerance of these depancreatized birds to a glucose load was estimated. Evaluation also was made of the action of tolbutamide in partially depancreatized, totally depancreatized, and sham-operated chickens. Results obtained indicate that partial pancreatectomy (99%) has only transient effects (several days) on plasma glucose and IRI levels. Also, surgical extirpation of the pancreas does not prevent tolbutamide-induced hypoglycemia secondary to insulin release, although such surgery does impair glucose tolerance concomitant with a diminished release of assayable IRI in response to an injected glucose load. Although remnant (1%) pancreatic tissue (splenic lobe) was observed to increase (up to 550%) in size 16 days after partial pancreatectomy, removal of this remnant in a second operation neither abolished circulating IRI 20 hr later nor did it prevent the characteristic hypoglycemic response to injected tolbutamide. The data presented herein suggest the existence of accessory β-islet tissue, yet to be located/described or, alternatively, the presence of a nonpancreatic source of insulin in the chicken.

Prolonged Fructose Feeding and Aldose Reductase Inhibition: Effect on the Polyol Pathway in Kidneys of Normal Rats

Abstract The effects of diets with differing carbohydrate composition on the kidney polyol pathway were investigated. The diets employed were F = fructose rich, G = glucose rich, S = cornstarch rich, and were fed for 30 days to six groups of 12 normal male Sprague-Dawley rats with and without addition of the aldose reductase inhibitor tolrestat (T). Fructose feeding resulted in higher kidney sorbitol levels (F = 0.847 ± 0.152, G = 0.354 ± 0.087, S = 0.207 ± 0.041 μM/g wet wt, P < 0.05). This was not observed in the tolrestat-treated animals (F + T = 0.182 ± 0.024, G + T = 0.149 ± 0.021, S + T = 0.152 ± 0.020 μM/g wet wt). Aldose reductase activity was reduced with tolrestat administration (F = 0.0208 ± 0.0023, F + T = 0.0048 ± 0.0005; G = 0.0210 ± 0.0002, G + T = 0.0059 ± 0.0008; S = 0.0227 ± 0.0022, S + T = 0.0062 ± 0.0007 μU). Myoinositol levels did not differ among groups (F = 1.973 ± 0.182, G = 2.291 ±0.307, S = 2.066 ± 0.155 μM/g wet wt), but tended to increase with aldose reductase inhibition (F + T = 2.253 ± 0.186, G + T = 2.713 ± 0.166, S + T = 2.618 ± 0.221 μM/g wet wt). Plasma glucose was higher in the fructose-fed rats (F = 10.78 ± 0.55, G = 9.09 ± 0.058, S = 9.03 ± 0.52, F + T = 9.75 ± 0.61, G + T = 8.42 ± 0.64, S + T = 8.81 ± 0.49 mM/liter). It is concluded that prolonged fructose feeding results in the accumulation of sorbitol in the kidney, caused by increased flux of glucose through the polyol pathway. This can be prevented by aldose reductase inhibition.

Biological evaluation of the third pancreatic hormone (APP): Hepatocyte and adipocyte effects

Abstract The effects of a third pancreatic hormone, avian pancreatic polypeptide (APP), on metabolism in isolated chicken adipocytes and hepatocytes were studied. APP inhibits glucagon-stimulated lipolysis within the chicken adipocyte by as much as 45%. Additionally, APP causes a marked reduction in glucagon-stimulated cyclic AMP accumulation, suggesting that APP in some way interferes with the glucagon-sensitive adenylate cyclase system. APP binds specifically to the chicken adipocyte; two orders of receptor sites were demonstrated: a high affinity ( K d = 5.6 × 10 −9 ), low capacity (6.75 × 10 −20 mol/cell) site and a low affinity ( K d = 8.3 × 10 −7 ), high capacity (3.18 × 10 −18 mol/cell) site. APP does not alter the rate of glycogenolysis or triglyceride synthesis at the chicken hepatocyte; neither does it stimulate glucose utilization by the chicken adipocyte.

Sensitivity of chicken and rat adipocytes and hepatocytes to isologous and heterologous pancreatic hormones

Abstract Isolated chicken and rat adipocytes and hepatocytes were used to compare chicken vs mammalian pancreatic hormones in regulating glucose and lipid metabolism. Porcine glucagon is twice as potent as chicken glucagon in eliciting lipolysis in both rat and chicken adipocytes. A less but marked bignificant difference exists between the two glucagon hormologs when glucose release by isolated hepatocytes is measured. Chicken adipocytes respond to lower concentrations (1 ng/ml) of glucagon than rat adipocytes (5 ng/ml). Hepatocytes from both animal species responded to glucagon at concentrations of 1 ng/ml. Chicken insulin is slightly more potent than porcine insulin both in inducing glucose utilization and in inhibiting glucagon-stimulated lipolysis in rat adipocytes. Glucose utilization, basal lipolysis, and the rate of glucagon-stimulated lipolysis of the chicken adipocyte are unaltered by insulin concentrations which are 1000 times greater than those required to affect rat adipocytes. The antilipolytic action of avian pancreatic polypeptide (APP) was compared with the bovine homolog of APP, namely bovine pancreatic polypeptide (BPP). BPP is not was effective as APP in inhibiting glucagon-stimulated lipolysis in the chicken adipocyte, and neither APP nor BPP inhibit glucagon-stimulated lipolysis in the rat adipocyte.

Insulin, pancreatic polypeptide, and glucagon release from the chicken pancreas in vitro: Responses to changes in medium glucose and free fatty acid content

Abstract The effects of glucose and fatty acids on release of insulin, pancreatic polypeptide (APP), and glucagon by the chicken pancreas were investigated in vitro. Small fragments of chicken pancreas were perifused; pancreatic hormones were measured in the effluent perifusate by radioimmunoassay. Elevation of medium glucose from 200 mg/dl to 500 mg/dl did not affect insulin release while elevation to 700 mg glucose/dl produced a sustained increase in insulin output. Glucagon release was consistently suppressed when glucose was elevated to either level. Pancreatic polypeptide secretion was not reduced by increases in medium glucose. Butyrate and valerate (5 mM) did not affect output of any of the hormones measured. Oleate and linoleate (0.3 and 1.0 mM, respectively) both stimulated insulin secretion; linoleate also stimulated APP and glucagon secretion. None of the fatty acids tested directly reduced glucagon or APP output although partial antagonistic effects of the fatty acids on pancreatic secretion were suggested by “off response” increases in the pancreatic hormone output upon removal of the long-chain fatty acids from the perifusion medium.

Failure of chicken insulin to alter polysaccharide levels of the avian glycogen body

Abstract 1. 1. In vivo and in vitro studies were carreid out comparing the effects of chicken insulin with those of equal amounts of beef insulin on the glycogen body of the domestic chicken. 2. 2. The glycogen body glycogen moiety was unaltered regardless of experimental design indicating that the constancy of this structures polysaccharide levels is neither attributable to use of a non-homologous hormone nor to restriction of the hormone by the avian blood-brain barrier.