We took advantage of thè partial protection exerted by suitable dosages of nicotinamide against thè p-cytotoxic effect of Streptozotocin (STZ) to create a new experimental diabetic syndrome in adult rats that appears closer to NIDDM than other available animai models with regard to insulin responsiveness to gincose and sulfonylureas. Among thè various dosages of nicotinamide tested in 3-month-old Wistar rats (100-350 mg/kg body wt), thè dosage of 230 mg/kg, given intraperitoneally 15 min before STZ administration (65 mg/kg i.v.) yielded a maximum of animals with moderate and stable nonfasting hyperglycemia (155 ± 3 vs. 121 ± 3 mg/dl in controls; P < 0.05) and 40% preservation of pancreatic insulin stores. We also evaluated liceli function both in vitro and in vivo 4-9 weeks after inducing diabetes. In thè isolated perfused pancreas, insulin response to glucose elevation (5-11 mmol/1) was clearly present, although significantly reduced with respect to controls (P < 0.01). Moreover, thè insulin response to tolbutamide (0.19 mmol/1) was similar to that observed in normal pancreases. Perfused pancreases from diabetic animals also exhibited a striking hypersensitivity to arginine infusìon (7 mmol/1). In rats administered STZ plus nicotinamide, intravenous glucose tolerance tests revealed clear abnormalities in glucose tolerance and insulin responsiveness, which were interestingly reversed by tolbutamide administration (40 mg/kg i.v.). In conclusion, this novel NIDDM syndrome with reduced pancreatic insulin stores, which is similar to human NIDDM in that it has a significant response to glucose (although abnormal in kinetics) and preserved sensitivity to tolbutamide, may provide a particularly advantageous tool for pharmacological investigations of new insulinotropic agents.

We took advantage of the partial protection exerted by suitable dosages of nicotinamide against the beta-cytotoxic effect of streptozotocin (STZ) to create a new experimental diabetic syndrome in adult rats that appears closer to NIDDM than other available animal models with regard to insulin responsiveness to glucose and sulfonylureas. Among the various dosages of nicotinamide tested in 3-month-old Wistar rats (100-350 mg/kg body wt), the dosage of 230 mg/kg, given intraperitoneally 15 min before STZ administration (65 mg/kg i.v.) yielded a maximum of animals with moderate and stable nonfasting hyperglycemia (155 +/- 3 vs. 121 +/- 3 mg/dl in controls; P < 0.05) and 40% preservation of pancreatic insulin stores. We also evaluated beta-cell function both in vitro and in vivo 4-9 weeks after inducing diabetes. In the isolated perfused pancreas, insulin response to glucose elevation (5-11 mmol/l) was clearly present, although significantly reduced with respect to controls (P < 0.01). Moreover, the insulin response to tolbutamide (0.19 mmol/l) was similar to that observed in normal pancreases. Perfused pancreases from diabetic animals also exhibited a striking hypersensitivity to arginine infusion (7 mmol/l). In rats administered STZ plus nicotinamide, intravenous glucose tolerance tests revealed clear abnormalities in glucose tolerance and insulin responsiveness, which were interestingly reversed by tolbutamide administration (40 mg/kg i.v.). In conclusion, this novel NIDDM syndrome with reduced pancreatic insulin stores, which is similar to human NIDDM in that it has a significant response to glucose (although abnormal in kinetics) and preserved sensitivity to tolbutamide, may provide a particularly advantageous tool for pharmacological investigations of new insulinotropic agents.

Evidence is presented showing that a neuronal isoform of nitric oxide synthase (NOS) is expressed in rat pancreatic islets and INS-1 cells. Sequencing of the coding region indicated a 99.8% homology with rat neuronal NOS (nNOS) with four mutations, three of them resulting in modifications of the amino acid sequence. Doubleimmunofluorescence studies demonstrated the presence of nNOS in insulin-secreting ␤-cells. Electron microscopy studies showed that nNOS was mainly localized in insulin secretory granules and to a lesser extent in the mitochondria and the nucleus. We also studied the mechanism involved in the dysfunction of the ␤-cell response to arginine and glucose after nNOS blockade with N Gnitro-L-arginine methyl ester. Our data show that miconazole, an inhibitor of nNOS cytochrome c reductase activity, either alone for the experiments with arginine or combined with sodium nitroprusside for glucose, is able to restore normal secretory patterns in response to the two secretagogues. Furthermore, these results were corroborated by the demonstration of a direct enzymesubstrate interaction between nNOS and cytochrome c, which is strongly reinforced in the presence of the NOS inhibitor. Thus, we provide immunochemical and pharmacological evidence that ␤-cell nNOS exerts, like brain nNOS, two catalytic activities: a nitric oxide production and an NOS nonoxidating reductase activity, both of which are essential for normal ␤-cell function. In conclusion, we suggest that an imbalance between these activities might be implicated in ␤-cell dysregulation involved in certain pathological hyperinsulinic states. T he short-lived free radical gas nitric oxide (NO) is synthesized from L-arginine by a family of enzymes known as NO synthases (NOSs). Three NOS isoenzymes, encoded by three separate genes, have been described, including the Ca 2ϩ /calmodulin-dependent and constitutively expressed neuronal NOS (nNOS) and endothelial NOS (eNOS) enzymes and a calmodulin-independent cytokine-inducible NOS (iNOS) enzyme found in various cell types (rev. in 1). The small amounts of NO, produced by the constitutive forms in response to increases in intracellular calcium, play a crucial role in a number of physiological functions, including neurotransmission (2), vascular tone (3) and platelet aggregation (4), whereas the large amounts, produced by iNOS in a calcium-independent manner over prolonged periods of time, are implicated in pathological functions, such as cytotoxicity of activated macrophages (5).Even though the inducible isoform has been cloned in insulin-producing cells after induction by cytokines (6), it is unclear whether pancreatic ␤-cells express a constitutive NOS. Both NADPH-diaphorase (NADPH-d) histochemical staining previously shown to be specific for NOS (7) and immunohistochemical studies using various nNOS antisera yielded apparently conflicting data. Positive NADPH-d and immunoreactive nNOS stainings have been found to be colocalized in most pancreatic endocrine cells (8), a finding not confirmed in other studies...

Strategies based on activating GLP-1 receptor (GLP-1R) are intensively developed for the treatment of type 2 diabetes. The exhaustive knowledge of the signaling pathways linked to activated GLP-1R within the ␤-cells is of major importance. In ␤-cells, GLP-1 activates the ERK1/2 cascade by diverse pathways dependent on either G␣ s /cAMP/cAMP-dependent protein kinase (PKA) or ␤-arrestin 1, a scaffold protein. Using pharmacological inhibitors, ␤-arrestin 1 small interfering RNA, and islets isolated from ␤-arrestin 1 knock-out mice, we demonstrate that GLP-1 stimulates ERK1/2 by two temporally distinct pathways. The PKA-dependent pathway mediates rapid and transient ERK1/2 phosphorylation that leads to nuclear translocation of the activated kinases. In contrast, the ␤-arrestin 1-dependent pathway produces a late ERK1/2 activity that is restricted to the ␤-cell cytoplasm. We further observe that GLP-1 phosphorylates the cytoplasmic proapoptotic protein Bad at Ser-112 but not at Ser-155. We find that the ␤-arrestin 1-dependent ERK1/2 activation engaged by GLP-1 mediates the Ser-112 phosphorylation of Bad, through p90RSK activation, allowing the association of Bad with the scaffold protein 14-3-3, leading to its inactivation. ␤-Arrestin 1 is further found to mediate the antiapoptotic effect of GLP-1 in ␤-cells through the ERK1/2-p90RSK-phosphorylation of Bad. This new regulatory mechanism engaged by activated GLP-1R involving a ␤-arrestin 1-dependent spatiotemporal regulation of the ERK1/2-p90RSK activity is now suspected to participate in the protection of ␤-cells against apoptosis. Such signaling mechanism may serve as a prototype to generate new therapeutic GLP-1R ligands.GLP-1 (glucagon-like peptide-1), produced by post-translational processing of the proglucagon in enteroendocrine L-cells, is a potent gluco-regulatory peptide hormone. GLP-1 is released into the blood stream in response to nutrient ingestion, such as carbohydrates, amino acids, and fats, during the early postprandial period (1, 2). A major target for GLP-1 actions is the pancreatic ␤-cell. One of the main physiological roles of this endocrine hormone is to enhance insulin secretion in a glucose-dependent manner (1-5). Besides its insulinotropic action, GLP-1 also favors the maintenance of a correct ␤-cell glucose sensing, regulates transcriptional synthesis, induces ␤-cell proliferation, and is protective against apoptosis (6 -9). Strategies based on activating GLP-1 receptor 3 are intensively developed for the treatment of type 2 diabetes and studies aiming at a better and exhaustive understanding of GLP-1 actions within the ␤-cells are of great importance (1-5).GLP-1 exerts its intracellular effects through binding to its specific receptor that spans the plasma membrane. The GLP-1R belongs to the class II (or B) secretin/glucagon/vasoactive intestinal peptide superfamily of heptahelical transmembrane G protein-coupled receptors (GPCRs) (10, 11). The GLP-1R is positively coupled to adenylate cyclase, through G␣ s -containing heterotrimeric G-prot...