Exogenous Nitric Oxide and Endogenous Glucose-Stimulated β-Cell Nitric Oxide Augment Insulin Release

  1. Simon R. Smukler1,
  2. Lan Tang1,
  3. Michael B. Wheeler12 and
  4. Anne Marie F. Salapatek1
  1. 1Department of Physiology, University of Toronto, Toronto, Ontario, Canada
  2. 2Department of Medicine, University of Toronto, Toronto, Ontario, Canada


    The role nitric oxide (NO) plays in physiological insulin secretion has been controversial. Here we present evidence that exogenous NO stimulates insulin secretion, and that endogenous NO production occurs and is involved in the regulation of insulin release. Radioimmunoassay measurement of insulin release and a dynamic assay of exocytosis using the dye FM1-43 demonstrated that three different NO donors—hydroxylamine (HA), sodium nitroprusside, and 3-morpholinosydnonimine (SIN-1)—each stimulated a marked increase in insulin secretion from INS-1 cells. Pharmacological manipulation of the guanylate cyclase/guanosine 3′,5′-cyclic monophosphate pathway indicated that this pathway was involved in mediating the effect of the intracellular NO donor, HA, which was used to simulate endogenous NO production. This effect was further characterized as involving membrane depolarization and intracellular Ca2+ ([Ca2+]i) elevation. SIN-1 application enhanced glucose-induced [Ca2+]i responses in primary β-cells and augmented insulin release from islets in a glucose-dependent manner. Real-time monitoring of NO using the NO-sensitive fluorescent dye, diaminofluorescein, was used to provide direct and dynamic imaging of NO generation within living β-cells. This showed that endogenous NO production could be stimulated by elevation of [Ca2+]i levels and by glucose in both INS-1 and primary rat β-cells. Scavenging endogenously produced NO-attenuated glucose-stimulated insulin release from INS-1 cells and rat islets. Thus, the results indicated that applied NO is able to exert an insulinotropic effect, and implicated endogenously produced NO in the physiological regulation of insulin release.


    • Address correspondence and reprint requests to Dr. Anne Marie Salapatek, University of Toronto, Dept. of Physiology, Rm. 3336, 1 King’s College Circle, Toronto, ON, Canada M5S 1A8. E-mail: annemarie.salapatek{at}

      Received for publication 5 December 2001 and accepted in revised form 30 August 2002.

      Partial funding for the studies performed was supplied by a Banting & Best Diabetes Centre−Eli Lilly Program grant.

      l-arg, l-arginine; [Ca2+]i, intracellular Ca2+ concentration; l-cit, l-citrulline; cNOS, constitutive NOS; CPT-cGMP, 8-(4-Chlorophenylthio)-guanosine 3′,5′-cyclic monophosphate; cPTIO, 2-(4-carboxyphenyl)-4,4,5,5-tetramethylimidazoline-1-oxyl-3-oxide; Daf2-DA, 4,5-diaminofluorescein diacetate; DiBAC4(3), bis-(1,3-dibutylbarbituric acid)trimethine oxonol; DZ, diazoxide; eNOS, endothelial NOS; FM1-43, N-(3-triethylammoniumpropyl)-4-(4-(dibutylamino)styryl) pyridinium dibromide; Fura2-AM, Fura2-acetoxymethyl ester; GC, guanylate cyclase; cGMP, guanosine 3′,5′-cyclic monophosphate; HA, hydroxylamine; iNOS, inducible NOS; [K+]o, extracellular K+ concentration; KATP channel, ATP-sensitive K+ channel; KRBB, Krebs-Ringer bicarbonate buffer; Kv channel, voltage-dependent K+ channel; l-NAME, NG-nitro-l-arginine methyl ester; l-NMMA, NG-monomethyl-l-arginine; nNOS, neuronal NOS; NO, nitric oxide; NOS, NO synthase; ODQ, 1H-[1,2,4]oxadiazolo[4,3-a]quinoxalin-1-one; l-OH-arg, NG-hydroxy-l-arginine; l-orn, l-ornithine; RIA, radioimmunoassay; SIN-1, 3-morpholinosydnonimine; SNP, sodium nitroprusside; TEA, tetraethylammonium; YC-1, 3-(5′-hydroxymethyl-3′-furyl)-1-benzylindazole.

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