Localization of thyrotropin-releasing hormone-and insulin-immunoreactivity in the pancreas of neonatal rats after injection of streptozotocin at birth

Summary Streptozotocin treatment at birth induces, in the pancreas of rats, first depletion of insulin and thyrotropin-releasing hormone and then early regeneration of cells and insulin, but not TRH. This study was undertaken to investigate whether the reduction in pancreatic TRH content can be associated with changes in the intensity and the distribution of TRH-immunoreactivity, and to follow the pattern of regeneration of cells through insulin- and TRH-immunoreactivity.In control animals, strong TRH-immunoreactivity was seen in insulin-containing cells on days 1–4 after birth. At day 7, the TRH-immunoreactivity was already decreased. In contrast, insulin-immunoreactivity was present throughout the neonatal period. A sparse population of cells near ducts also contained both TRH- and insulin-immunoreactivity at 1–2 days age.In streptozotocin-treated animals, TRH-immunoreactivity is found only in a few scattered insulin-containing cells in altered islets on days 1–4. Near the ducts, there were new insulin-containing cells which did not contain TRH. From day 7 regeneration of endocrine cells was characterized by new, typical islets, but these contained insulin-, but not TRH-immunoreactivity. These findings suggest a differential control of the biosynthesis of insulin and TRH within the pancreas.     

Expression of cell type-specific markers during pancreatic development in the mouse: implications for pancreatic cell lineages

Summary The islet cells of the mammalian pancreas are comprised of four different endocrine cell types, each containing a specific hormone. Islet cells also contain two enzymes of the catecholamine biosynthetic pathway: tyrosine hydroxylase (TH) and aromatic L-amino acid decarboxylase (AADC). The cell lineage relationships of these different cell types have not been examined and it is not known whether, during development, they originate from the same or from different precursor populations. In this study we used immunocytochemical procedures to determine whether developing pancreatic cells express markers common to endocrine and exocrine cell types. We found that acinar cell precursors express AADC prior to the appearance of an exocrine marker and that the expression of AADC in acinar cells persists throughout embryogenesis to the first month of postnatal life. At this time, acinar cells do not contain AADC. We also found that exocrine cells containing AADC never express other islet-cell markers. These findings suggest that while acinar and islet cells both arise from precursor cells containing AADC, these progenitor cells do not express a combined endocrine-exocrine phenotype.     

Localization and characterization of neuropeptide Y-like peptides in the brain and islet organ of the anglerfish (Lophius americanus)

Summary Results from a previous report demonstrate that more than one molecular form of neuropeptide Y-like peptide may be present in the islet organ of the anglerfish (Lophius americanus). Most of the neuropeptide Y-like immunoreactive material was anglerfish peptide YG, which is expressed in a subset of islet cells, whereas an additional neuropeptide Y-like peptide(s) was localized in islet nerves. To learn more about the neuropeptide Y-like peptides in islet nerves, we have employed immunohistochemical and biochemical methods to compare peptides found in anglerfish islets and brain. Using antisera that selectively react with either mammalian forms of neuropeptide Y or with anglerfish peptide YG, subsets of neurons were found in the brain that labelled with only one or the other of the antisera. In separate sections, other neurons that were labelled with either antiserum exhibited similar morphologies. Peptides from brains and islets were subjected to gel filtration and reverse-phase high performance liquid chromatography. Radioimmunoassays employing either the neuropeptide Y or peptide YG antisera were used to examine chromatographic eluates. Immunoreactive peptides having retention times of human neuropeptide Y and porcine neuropeptide Y were identified in extracts of both brain and islets. This indicates that peptides structurally similar to both of these peptides from the neuropeptide Y-pancreatic polypeptide family are expressed in neurons of anglerfish brain and nerve fibers of anglerfish islets. The predominant form of neuropeptide Y-like peptide in islets was anglerfish peptide YG. Neuropeptide Y-immunoreactive peptides from islet extracts that had chromatographic retention times identical to human neuropeptide Y and porcine neuropeptide Y were present in much smaller quantities. These results are consistent with the hypothesis that peptides having significant sequence homology with human neuropeptide Y and porcine neuropeptide Y are present in the nerve fibers that permeate the islet.     

Inhibition of isoproterenol-induced DNA and RNA synthesis in rat parotid and pancreas following removal of submandibular-sublingual glands

Summary [³H] thymidine incorporation into DNA of the parotid (PA) gland of adult and 20-day-old rats and into DNA of the pancreas (PANC) of 20-day-old rats was increased markedly following a 2-day regimen of isoproterenol (ISO) administration. However, when the submandibular-sublingual (SM-SL) glands had been removed just prior to initiation of the ISO injections, the [³H] thymidine incorporation into PA and PANC was inhibited, and cpm/mg protein of these organs was even lower than that of organs of untreated rats with SM-SL glands present. Removal of the PA glands just prior to initiation of the ISO regimen had no effect on the ISO-induced [³H] thymidine incorporation into DNA of PANC but partially inhibited that of the submandibular (SM) gland. It is suggested that the inhibitory effects on DNA and RNA synthesis that follow removal of SM-SL glands are attributable to the growth factors (epidermal growth factor and nerve growth factor) found in the rat SM gland. These factors appear to regulate normal DNA synthetic activity of exocrine glands as well as ₁-adrenoceptor mediated DNA synthesis. Cellular hypertrophy induced by the ISO was less markedly affected by absence of the SM glands, but a partial inhibition of [³H] uridine incorporation into RNA of PA of adult rats also occurred when SM-SL glands were removed prior to initiation of the ISO-regimen.     

Cellular localization of intrinsic factor in pancreas and stomach of the dog

Summary A cobalamin (vitamin B₁₂)-binding protein has recently been identified in canine pancreatic juice which is biochemically, immunochemically and functionally similar to canine gastric intrinsic factor. However, the cellular sources of both this pancreatic intrinsic factor and gastric intrinsic factor in the dog are not known. Antisera raised against canine gastric intrinsic factor have been used to examine the distribution of intrinsic factors in the canine pancreas and stomach. Immunoreactivity was demonstrated in duct cells but not acinar or endocrine cells in the pancreas, and in fundic peptic and pyloric gastric pit cells in stomach. All immunostaining was abolished by preabsorption of the antisera with purified canine gastric and pancreatic intrinsic factors. A cellular source of pancreatic intrinsic factor has not been previously described, and the demonstration of intrinsic factor-like immunoreactivity in two cell types in the canine stomach contrasts with its localization in a single cell type in the gastric mucosa of other mammalian species. Furthermore, immunoreactivity in pancreatic duct cells was detected at much higher dilutions of antisera than those required for staining of peptic and gastric pit cells. This suggests a higher concentration of antigen, and supports previous evidence that the pancreas is a major source of intrinsic factor in the dog.     

Lysosomes and pancreatic islet function

Summary Ultrastructural studies of pancreatic islets have suggested that crinophagy provides a possible mechanism for intracellular degradation of insulin in the insulin-producing B-cells. In the present study, a quantitative estimation of crinophagy in mouse pancreatic islets was attempted by morphometric analysis of lysosomes containing immunoreactive insulin. Isolated islets were incubated in tissue culture for one week in 3.3, 5.5 or 28 mmol/l glucose. The lysosomes of the pancreatic B-cells were identified by morphological and enzyme-cytochemical criteria and divided into three subpopulations comprising primary lysosomes and insulin-positive or insulin-negative secondary lysosomes. Both the volume and numerical density of the primary lysosomes increased with increasing glucose concentration. The proportion of insulin-containing secondary lysosomes was highest at 5.5 and lowest at 3.3 mmol/l glucose. Insulin-negative secondary lysosomes predominated at 3.3 mmol/l glucose. Studies of the dose-response relationships of glucose-stimulated insulin biosynthesis and insulin secretion of the pancreatic islets showed that biosynthesis had an apparent K_m-value for glucose of 7.0 mmol/l, whereas it was 14.5 mmol/l for secretion. The pronounced crinophagic activity at 5.5 mmol/l glucose may thus be explained by the difference in glucose sensitivity between insulin biosynthesis and secretion resulting in an intracellular accumulation of insulin-containing secretory granules. The predominance of insulin-negative secondary lysosomes at 3.3 mmol/l glucose may reflect an increased autophagy, whereas the predominance of primary lysosomes at 28 mmol/l glucose may reflect a generally low activity of intracellular degradative processes.     

Immunohistochemical localization of porcine diazepam-binding inhibitor (DBI) to rat endocrine pancreas

Summary The occurrence of diazepam-binding inhibitor (DBI), isolated and characterized from porcine upper intestine, was examined in the pancreas of Sprague-Dawley albino rats using indirect immunofluorescence. The polypeptide was found in the endocrine Langerhans islets and, utilizing double-labelling controls, it was shown to be present within the peripherally located glucagon-containing cells. Regulation of islet hormone production may therefore be under DBI control.     

Effect of initial developmental stage on morphology of transplanted embryonic chick pancreas

Summary It has been reported that only certain types of pancreatic parenchymal cells survive transplantation. This study examines whether the extent of differentiation of the pancreas at the time of transplantation affects the resulting morphology or viability of its components. Segments of chick pancreas or its primordia from stages preceding formation of dorsal bud (60 h) through hatching (day 21) were implanted in the abdominal region of three-day chick embryos. After various periods of growth, grafts were examined by light- and electron microscopy. In all transplants, individual endocrine cells (A, B, D, PP) and islet structure were identical to those of normal embryos of comparable age. The exocrine portion also appeared normal in implants from embryos younger than seven days. In grafts from older donors, however, normal acini and ducts were replaced by dilated, irregularly shaped tubules from which new islets appeared to develop. These results suggest that transplantation causes structural modification of exocrine tissue, which may reflect its initial functional capabilities. These observations are compared with similar structural alterations that occur following experimental obstruction of ducts and in human pancreatic pathology.     

Chronic administration of an organophosphorus insecticide to rats alters cholinergic muscarinic receptors in the pancreas

Male rats were treated for 10 days with the organophosphorus insecticide, acetylcholinesterase inhibitor, O,O-diethyl S-[2-(ethylthio)ethyl]phosphorodithioate (disulfoton, 2 mg/kg/day by gavage). At the end of the treatment, binding of [³H]quinuclidinyl benzilate ([³H]QNB) to cholinergic muscarinic receptors and cholinesterase (ChE) activity were assayed in the pancreas. Functional activity of pancreatic muscarinic receptor was investigated by determining carbachol-stimulated secretion of α-amylase in vitro. ChE activity and [³H]QNB binding were significantly decreased in the pancreas from disulfoton-treated rats. The alteration of [³H]QNB binding was due to a decrease in muscarinic receptor density with no change in the affinity. Basal secretion of amylase from pancreas in vitro was not altered, but carbachol-stimulated secretion was decreased. The effect appeared to be specific since pancreozymin was able to induce the same amylase release from pancreases of control and treated rats. The results suggest that repeated exposures to sublethal doses of an organophosphorus insecticide lead to a biochemical and functional alteration of cholinergic muscarinic receptors in the pancreas.     

Enhancement of the repair of carcinogen-induced DNA damage in the hamster pancreas by dietary selenium

We measured single strand breaks (SSB) in pancreas DNA produced by N-nitrosobis(2-oxopropyl)amine (BOP) in hamsters fed purified diets containing added sodium selenite (Se) at 0.0, 0.1 and 5.0 ppm. There were fewer SSB in those given the 5.0 ppm Se diet throughout the experiment. One hour after dosing with BOP (20 mg/kg), there were 2.26 ± 0.47, 2.83 ± 0.43 and 1.74 ± 0.43 SSB per 10⁸ daltons (mean ± S.E.M.) respectively in the three groups. The SSB were repaired faster in the 5.0 ppm Se-fed group. The approximate half-lives of the SSB were 33, 30 and 8 days, respectively. In the hamsters fed 5.0 ppm Se there was a small, statistically significant increase in pancreatic DNA synthesis. Autoradiographic analysis indicated that this was repair synthesis. In a second experiment, hamsters were fed one of the three diets prior to and for 2 days after administration of a single dose of BOP (20 mg/kg). They were then fed the 5.0 ppm Se diet for 5 days. The number of SSB was compared with those in hamsters fed their original diet for 7 days after BOP dosing. There was a statistically significant difference in the number of SSB in the hamsters fed 0.1 ppm Se before and for 2 days after BOP. In these hamsters there were 1.21 ± 0.24 SSB per 10⁸ daltons compared with 3.19 ± 0.4 (mean ± S.E.M.). These results suggest high levels of dietary Se stimulate the repair of carcinogen-induced DNA damage.