Dentate Gyrus Somatostatin Cells are Required for Contextual Discrimination During Episodic Memory Encoding.

Memory systems ought to store and discriminate representations of similar experiences in order to efficiently guide future decisions. This problem is solved by pattern separation, implemented in the dentate gyrus (DG) by granule cells to support episodic memory formation. Pattern separation is enabled by tonic inhibitory bombardment generated by multiple GABAergic cell populations that strictly maintain low activity levels in granule cells. Somatostatin-expressing cells are one of those interneuron populations, selectively targeting the distal dendrites of granule cells, where cortical multimodal information reaches the DG. Nonetheless, somatostatin cells have very low connection probability and synaptic efficacy with both granule cells and other interneuron types. Hence, the role of somatostatin cells in DG circuitry, particularly in the context of pattern separation, remains uncertain. Here, by using optogenetic stimulation and behavioral tasks in mice, we demonstrate that somatostatin cells are required for the acquisition of both contextual and spatial overlapping memories.

Pancreatic Endocrine Effects of Dopamine Receptors in Human Islet Cells.

OBJECTIVE: To date, there are no reports on the cellular localization of dopamine receptors in the human pancreas. In our study, we determined the localization and expression of 5 dopamine receptors (D(1), D(2), D(3), D(4), and D(5)) in normal human pancreas tissue. METHODS: Human nonpathological pancreas tissues were fixed with 4% paraformaldehyde, paraffin-embedded, and processed for immunohistochemical analysis to detect dopamine receptors in the human pancreas tissue by using double immunofluorescent labeling and confocal microscopy. RESULTS: We found that the D(1) receptor is present in ß cells; the D(2) receptor is expressed by α, δ, and pancreatic polypeptide cells; the D(4) receptor is expressed by ß and polypeptide cells; whereas the D(5) receptor is expressed only by δ cells. CONCLUSIONS: Our results identify the dopamine receptors (D(1)-D(5)) in normal pancreas tissue and provide a morphological basis for studying the pancreatic endocrine effects of dopamine and suggest a new target for the clinical treatment of diabetes.

Endocrine cells in the ileum of patients with irritable bowel syndrome.

AIM: To study the ileal endocrine cell types in irritable bowel syndrome (IBS) patients. METHODS: Ninety-eight patients with IBS (77 females and 21 males; mean age 35 years, range 18-66 years) were included, of which 35 patients had diarrhea (IBS-D), 31 patients had a mixture of both diarrhea and constipation (IBS-M), and 32 patients had constipation (IBS-C) as the predominant symptoms. The controls were 38 subjects (26 females and 12 males; mean age 40 years, range 18-65 years) who had submitted to colonoscopy for the following reasons: gastrointestinal bleeding, where the source of bleeding was identified as hemorrhoids (n = 24) or angiodysplasia (n = 3), and health worries resulting from a relative being diagnosed with colon carcinoma (n = 11). The patients were asked to complete the: Birmingham IBS symptom questionnaire. Ileal biopsy specimens from all subjects were immunostained using the avidin-biotin-complex method for serotonin, peptide YY (PYY), pancreatic polypeptide (PP), enteroglucagon, and somatostatin cells. The cell densities were quantified by computerized image analysis, using Olympus cellSens imaging software. RESULTS: The gender and age distributions did not differ significantly between the patients and the controls (P = 0.27 and P = 0.18, respectively). The total score of Birmingham IBS symptom questionnaire was 21 ± 0.8, and the three underlying dimensions: pain, diarrhea, and constipation were 7.2 ± 0.4, 6.6 ± 0.4, and 7.2 ± 0.4, respectively. The density of serotonin cells in the ileum was 40.6 ± 3.6 cells/mm² in the controls, and 11.5 ± 1.2, 10.7 ± 5.6, 10.0 ± 1.9, and 13.9 ± 1.4 cells/mm² in the all IBS patients (IBS-total), IBS-D, IBS-M, and IBS-C patients, respectively. The density in the controls differed significantly from those in the IBS-total, IBS-D, IBS-M, and IBS-C groups (P < 0.0001, P = 0.0001, P = 0.0001, and P < 0.0001, respectively). There was a significant inverse correlation between the serotonin cell density and the pain dimension of Birmingham IBS symptom questionnaire (r = -0.6, P = 0.0002). The density of PYY cells was 26.7 ± 1.6 cells/mm(2) in the controls, and 33.1 ± 1.4, 27.5 ± 1.4, 34.1 ± 2.5, and 41.7 ± 3.1 cells/mm² in the IBS-total, IBS-D, IBS-M, and IBS-C patients, respectively. This density differed significantly between patients with IBS-total and IBS-C and the controls (P = 0.03 and < 0.0001, respectively), but not between controls and, IBS-D, and IBS-M patients (P = 0.8, and P = 0.1, respectively). The density of PYY cells correlated significantly with the degree of constipation as recorded by the Birmingham IBS symptom questionnaire (r = 0.6, P = 0.0002). There were few PP-, enteroglucagon-, and somatostatin-immunoreactive cells in the biopsy material examined, which made it impossible to reliably quantify these cells. CONCLUSION: The decrease of ileal serotonin cells is associated with the visceral hypersensitivity seen in all IBS subtypes. The increased density of PYY cells in IBS-C might contribute to the constipation experienced by these patients.

Selective expression of TLQP-21 and other VGF peptides in gastric neuroendocrine cells and modulation by feeding.

Although vgf gene knockout mice are hypermetabolic, administration of the VGF peptide TLQP-21 itself increased energy consumption. Agonist-antagonist roles are thus suggested for different VGF peptides, and the definition of their tissue heterogeneity is mandatory. We studied the rat stomach using antisera to C- or N-terminal sequences of known or predicted VGF peptides in immunohistochemistry and ELISA. TLQP (rat VGF(556-565)) peptide/s were most abundant (162±11 pmol/g, mean±s.e.m.) and were brightly immunostained in enterochromaffin-like (ECL) cells and somatostatin cells. A peptide co-eluting with TLQP-21 was revealed in HPLC of gastric and hypothalamic extracts, while the extended TLQP-62 form was restricted to the hypothalamus. Novel PGH (rat VGF(422-430)) peptide/s were revealed in ghrelin cells, mostly corresponding to low MW forms (0.8-1.5  kDa), while VGF C-terminus peptides were confined to neurons. VGF mRNA was present in the above gastric endocrine cell types, and was prominent in chief cells, in parallel with low-intensity staining for further cleaved products from the C-terminal region of VGF (HVLL peptides: VGF(605-614)). In swine stomach, a comparable profile of VGF peptides was revealed by immunohistochemistry. When fed and fasted rats were studied, a clear-cut, selective decrease on fasting was observed for TLQP peptides only (162±11 vs 74±5.3  pmol/g, fed versus fasted rats, mean±s.e.m., P<0.00001). In conclusion, specific VGF peptides appear to be widely represented in different gastric endocrine and other mucosal cell populations. The selective modulation of TLQP peptides suggests their involvement in peripheral neuro-endocrine mechanisms related to feeding responses and/or ECL cell regulation.

Pseudopod-like basal cell processes in intestinal cholecystokinin cells.

Cholecystokinin (CCK) is secreted by neuroendocrine cells comprising 0.1%-0.5% of the mucosal cells in the upper small intestine. Using CCK promoter-driven green fluorescent protein (GFP) expression in transgenic mice, we have applied immunofluorescence techniques to analyze the morphology of CCK cells. GFP and CCK colocalize in neuroendocrine cells with little aberrant GFP expression. CCK-containing cells are either flask- or spindle-shaped, and in some cells, we have found dendritic processes similar to pseudopods demonstrated for gut somatostatin-containing D cells. Most pseudopods are short, the longest process visualized extending across three cells. Pseudopods usually extend to adjacent cells but some weave between neighboring cells. Dual processes have also been observed. Three-dimensional reconstructions suggest that processes are not unidirectional and thus are unlikely to be involved in migration of CCK cells from the crypt up the villus. Abundant CCK immunostaining is present in the pseudopods, suggesting that they release CCK onto the target cell. In order to identify the type of cells being targeted, we have co-stained sections with antibodies to chromogranin A, trefoil factor-3, and sucrase-isomaltase. CCK cell processes almost exclusively extend to sucrase-isomaltase-positive enterocytes. Thus, CCK cells have cellular processes possibly involved in paracrine secretion.

DTA-mediated targeted ablation revealed differential interdependence of endocrine cell lineages in early development of zebrafish pancreas.

Cell lineage analysis is critical in understanding the relationship between progenitors and differentiated cells as well as the mechanism underlying the process of differentiation. In order to study the zebrafish endocrine pancreas cell lineage, transgenic expression of diphtheria toxin gene A chain (DTA) under two cell type-specific promoters derived from the insulin (ins) and somatostatin2 (sst2) genes was used to ablate the two types of endocrine cells: insulin-producing beta-cells and somatostatin-producing delta-cells, respectively. We found that ablation of beta-cells resulted in a reduction of not only beta-cells but also glucagon-producing alpha-cells; in contrast, delta-cells were largely unaffected. Ablation of delta-cells led to reduction of all three types of endocrine cells: alpha-, beta-, and delta. Interestingly, alpha-cells were more profoundly affected in both beta- and delta-cell ablations and were frequently reduced together with beta- and delta-cells. By taking advantage of Tg(ins:gfp) and Tg(sst2:gfp) lines, we also monitored the changes of different types of endocrine cells in vivo after ablation and found that both beta- and delta-cell populations significantly recovered by 3dpf after their ablation and it seemed that delta-cells had a better capability of recovery than beta-cells. Thus, our current observations indicated differential interdependence of these three cell lineages. The development of zebrafish alpha-cells, but not delta-cells, is dependent on beta-cells, while the development of both alpha- and beta-cells is dependent on delta-cells. In contrast, the development of delta-cells was independent of beta-cells.

An immunohistochemical study of endocrine cells in the stomach of hypertensive rats.

Essential hypertension is a complex disease with both genetic and environmental determinants. The effect of spontaneous hypertension on the distribution and occurrence of somatostatin-, gastrin- and serotonin-immunoreactive cells in the fundus and pylorus of the rat stomach was examined by immunohistochemistry. The animals were killed by decapitation at 4 and 16 weeks of age (5 control rats and 5 hypertensive rats). Endocrine cells generally increase in number in hypertensive rats as compared to control rats. However, the detailed responses of endocrine cells to hypertension depend on the cell type, region of gastric mucosa and age of animals. The present results suggest that hypertension has an influence on the intrinsic regulatory system by endocrine cells control in the rat stomach.

Intranuclear rodlets in human pancreatic islet cells.

OBJECTIVES: Intranuclear rodlets (INRs) are rod-shaped intranuclear inclusions that we have described in neurons of the human brain. We recently identified these structures in pancreatic islet cells. The objectives of this study are to describe the light microscopic features and cellular pattern of distribution of INRs in human pancreatic islet cells. METHODS: Double immunofluorescence staining was performed on 5 human pancreatic tissue samples for the detection of class III beta tubulin (C3T) to detect INRs and for promyelocytic leukemia (PML) protein to examine the relationship between PML and INRs. RESULTS: Intranuclear rodlets were detected in 22.99% of pancreatic B cells compared with only 3.11%, 1.80%, and 1.60% of A, D, and PP cells, respectively. Twenty-four percent of C3T-immunoreactive INRs showed partial or complete immunoreactivity for PML. Promyelocytic leukemia staining within the nuclei of B cells was confined to INRs and was not present in the typical PML bodies present in other cell types. Spatially, PML and C3T staining of islet cell INRs appeared to be mutually exclusive within individual INRs. CONCLUSIONS: Intranuclear rodlets are present within the nuclei of pancreatic islet cells, where they reside predominantly but not exclusively in B cells. Immunoreactivity of B-cell INRs for PML suggests that the functional significance of INRs may be related to that of PML and/or PML bodies. Conversely, the exclusive localization of PML staining to INRs in B cells indicates that PML's function in B cells is selectively associated with INRs. The mutually exclusive pattern of PML and C3T staining suggests dynamic interactions between these 2 proteins in B-cell INRs. In light of evidence for the involvement of INRs and of PML bodies in disease, it will be of interest to investigate these structures in animal models of diabetes and in human diabetes.

Noninsulinoma pancreatogenous hypoglycemia syndrome: quantitative and immunohistochemical analyses of islet cells for insulin, glucagon, somatostatin, and pancreatic and duodenal homeobox protein.

OBJECTIVE: To present a case of an elderly man with noninsulinoma pancreatogenous hypoglycemia syndrome (NIPHS) and to determine the pathogenesis of this syndrome. METHODS: The pancreas of our patient with NIPHS was immunocytochemically stained for insulin-, glucagon-, and somatostatin-secreting cells and pancreatic and duodenal homeobox protein (PDX-1). The clinical findings and morphologic and immunocytochemical analyses of the islets of our patient are described, along with a review of related published reports. RESULTS: A 78-year-old man presented with hyperinsulinemic hypoglycemia, with episodes unrelated to meals or fasting. An insulinoma could not be localized by preoperative imaging or by intraoperative ultrasonography or palpation. He underwent a 70% distal pancreatectomy. For assessment of the possibility that a nuclear transcription factor regulating islet beta-cell growth and development is overexpressed in this disease and is responsible for diffuse islet hyperfunction and proliferation of beta-cells, pancreatic sections from our patient were stained immunocytochemically for PDX-1, insulin, glucagon, and somatostatin. Morphologic findings were compared with pancreatic sections from normal control patients and normative data reported in the literature. Clinical findings and morphologic analyses were consistent with NIPHS. Islets were arranged in long clusters, both in the pancreatic tissue and in peripancreatic adipose tissue. Islets were small but increased in number, and insulin, glucagon, and somatostatin were present in the islets. The relative intensity of insulin staining was increased in our patient in comparison with that in the control patients, and PDX-1 was not overexpressed. CONCLUSION: The etiopathogenesis of NIPHS in this patient involved (1) an increased number of islets with development of ectopic islets in the peripancreatic adipose tissue; (2) alpha- and delta- as well as beta-cell proliferation; and (3) an early step in the development of the islet not involving overexpression of PDX-1.

Regulation of amylin release from cultured rabbit gastric fundic mucosal cells.

BACKGROUND: Amylin (islet amyloid polypeptide) is a hormone with suggested roles in the regulation of glucose homeostasis, gastric motor and secretory function and gastroprotection. In the gastric mucosa amylin is found co-localised with somatostatin in D-cells. The factors regulating gastric amylin release are unknown. In this study we have investigated the regulation of amylin release from gastric mucosal cells in primary culture. Rabbit fundic mucosal cells enriched for D-cells by counterflow elutriation were cultured for 40 hours. Amylin and somatostatin release over 2 hours in response to agonists were assessed. RESULTS: Amylin release was significantly enhanced by activation of protein kinase C with phorbol-12-myristate-13-acetate, adenylate cyclase with forskolin and elevation of intracellular calcium with A23187. Cholecystokinin (CCK), epinephrine and glucagon-like peptide-1 (GLP-1) each stimulated amylin release in a dose-dependent manner. Maximal CCK-stimulated release was greater than either epinephrine or GLP-1, even when the effects of the latter two were enhanced by isobutylmethylxanthine. Stimulated amylin release was significantly inhibited by carbachol (by 51-59%) and octreotide (by 33-42%). Somatostatin release paralleled that of amylin. CONCLUSIONS: The cultured D-cell model provides a means of studying amylin release. Amylin secretion is stimulated by receptor-dependent and -independent activation of Ca2+/protein kinase C and adenylate cyclase pathways. Inhibition involves activation of muscarinic receptors and auto-regulation by somatostatin.

CLN-3 protein is expressed in the pancreatic somatostatin-secreting delta cells.

Juvenile neuronal ceroid lipofuscinosis (JNCL) is a severe autosomal recessive neurodegenerative disorder resulting from mutations in the CLN3 gene. The gene product is a 438-amino acid hydrophobic peptide of unknown function containing five transmembrane domains. In order to study the tissue distribution of the peptide, polyclonal antibodies were raised in rabbits to three epitopes and were affinity purified before use. All three antibodies were used together for immunocytochemical staining of human pancreas. This staining showed localization in pancreatic islet cells. Double labelling of the tissue indicated that cells staining for the CLN3 protein were also positive for somatostatin.

Maternal adrenocortical hormones maintain the early development of pancreatic B cells in the fetal rat.

To investigate the effect of maternal adrenocortical hormones on the development of fetal pancreatic islet cells, pregnant rats were adrenalectomised on d 6 of gestation. On d 12-16 the growth patterns of fetal insulin-producing B cells, glucagon-producing A cells, and somatostatin-producing D cells were observed histometrically. Maternal adrenalectomy resulted in growth retardation of fetal B cells on d 12-15. Maternal corticosterone therapy prevented this retardation. Maternal adrenalectomy, however, did not affect the developmental patterns of A and D cells. By Western blotting and immunohistochemistry, glucocorticoid receptors were demonstrated to be present in the islet cells from d 12 to d 15. These results suggest that maternal adrenocortical hormones, glucocorticoids in particular, maintain the early development of fetal pancreatic B cells through their specific intracellular glucocorticoid receptor.

Effect of islet amyloid polypeptide on somatostatin inhibition of insulin secretion from isolated rat pancreatic islets.

In this study, we investigated the presence of islet amyloid polypeptide (IAPP) in somatostatin cells of rat endocrine pancreas and the effect of exogenous IAPP and somatostatin, separate or combined, on in vitro insulin secretion. By immunocytochemistry, IAPP was found in both B and D cells of rat pancreatic islets. Furthermore, the labeling density of IAPP in D cells was nearly four times higher than in B cells. After a 2-day preincubation in RPMI 1640 (11.1 mM glucose), isolated rat pancreatic islets were exposed to IAPP and/or somatostatin for 90 min in modified Krebs-Ringer bicarbonate (KRB) buffers containing 11.1 or 22.2 mM glucose, or 11.1 mM glucose + 10 mM L-arginine, respectively. At 11.1 mM glucose, insulin secretion was not affected by IAPP and/or somatostatin at concentrations investigated. Insulin response to 22.2 mM glucose was inhibited by exogenous somatostatin. Arginine-stimulated insulin secretion was also inhibited by somatostatin, and the effect was significantly potentiated with additional 10(-5) M IAPP. The study shows that rat pancreatic D cells have higher IAPP density than B cells in the same islets and that IAPP and somatostatin may cooperate on rat pancreatic B cells to regulate the insulin secretion in response to potent stimulation.

Ontogeny of somatostatin cells in the rat fetal pancreas.

The growth pattern of somatostatin cells was clarified immunohistochemically from day 12 to day 18 of gestation in the rat fetus. On day 12, somatostatin cells first appeared within the pancreatic anlage. The total number of somatostatin cells was gradually increased from day 12 to day 16 and rapidly increased thereafter. It may be concluded that such a sequence of events of development in somatostatin cells occurs in a fashion similar to that in B cells.