h-89 and 2-phenyl-4-4-5-5-tetramethylimidazoline-1-oxyl-3-oxide

We have examined the expression and activity of inducible nitric oxide synthase (iNOS) and the activity of neuronal constitutive NOS (ncNOS) in isolated rat pancreatic islets, stimulated by a "hyperglycaemic" concentration of glucose, and whether the NOS activities could be modulated by activation of the cyclic AMP/protein kinase A (cyclic AMP/PKA) system in relation to the insulin secretory process. Here, we show that glucose stimulation (20 mmol/l) induces iNOS and increases ncNOS activity. No iNOS is detectable at basal glucose levels (3.3 mmol/l). The addition of glucagon-like-peptide 1 (GLP-1) or dibutyryl-cAMP to islets incubated with 20 mmol/l glucose results in a marked suppression of iNOS expression and activity, a reduction in ncNOS activity and increased insulin release. The GLP-1-induced suppression of glucose-stimulated iNOS activity and expression and its stimulation of insulin release is, at least in part, PKA dependent, since the PKA inhibitor H-89 reverses the effects of GLP-1. These observations have been confirmed by confocal microscopy showing the glucose-stimulated expression of iNOS, its suppression by GLP-1 and its reversion by H-89 in beta-cells. We have also found that the NO scavenger cPTIO and the NOS inhibitor L-NAME potentiate the insulin response to glucose, again suggesting that NO is a negative modulator of glucose-stimulated insulin release. We conclude that the induction of iNOS and the increase in ncNOS activity caused by glucose in rat islets is suppressed by the cyclic AMP/PKA system. The inhibition of iNOS expression by the GLP-1/cyclic AMP/PKA pathway might possibly be of therapeutic potential in NO-mediated beta-cell dysfunction and destruction.

Topics: Animals; Bucladesine; Cyclic AMP; Cyclic AMP-Dependent Protein Kinase Type II; Cyclic AMP-Dependent Protein Kinases; Cyclic N-Oxides; Drug Synergism; Enzyme Activation; Enzyme Inhibitors; Free Radical Scavengers; Glucagon; Glucagon-Like Peptide 1; Glucose; Imidazoles; Insulin; Insulin Secretion; Islets of Langerhans; Isoquinolines; Male; Microdissection; Microscopy, Confocal; NG-Nitroarginine Methyl Ester; Nitric Oxide Synthase; Nitric Oxide Synthase Type II; Peptide Fragments; Protein Kinase Inhibitors; Protein Precursors; Rats; Rats, Sprague-Dawley; Sulfonamides