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Insulin induces the release of vasodilator compounds from platelets by a nitric oxide-G kinase-VAMP-3-dependent pathway.

Randriamboavonjy V, Schrader J, Busse R, Fleming I - J. Exp. Med. (2004)

Bottom Line: Insulin failed to relax endothelium-intact rings of porcine coronary artery.Insulin induced the release of adenosine trisphosphate (ATP), adenosine, and serotonin from platelet-dense granules in a NO-dependent manner.The mechanism of dense granule secretion involves the G kinase-dependent association of syntaxin 2 with vesicle-associated membrane protein 3.

View Article: PubMed Central - PubMed

Affiliation: Institut für Kardiovaskuläre Physiologie, Klinikum der J.W.Goethe-Universität, D-60590 Frankfurt am Main, Germany.

ABSTRACT
Insulin-induced vasodilatation is sensitive to nitric oxide (NO) synthase (NOS) inhibitors. However, insulin is unable to relax isolated arteries or to activate endothelial NOS in endothelial cells. Since insulin can enhance platelet endothelial NOS activity, we determined whether insulin-induced vasodilatation can be attributed to a NO-dependent, platelet-mediated process. Insulin failed to relax endothelium-intact rings of porcine coronary artery. The supernatant from insulin-stimulated human platelets induced complete relaxation, which was prevented by preincubation of platelets with a NOS inhibitor, the soluble guanylyl cyclase inhibitor, NS 2028, or the G kinase inhibitor, KT 5823, and was abolished by an adenosine A2A receptor antagonist. Insulin induced the release of adenosine trisphosphate (ATP), adenosine, and serotonin from platelet-dense granules in a NO-dependent manner. This response was not detected using insulin-stimulated platelets from endothelial NOS-/- mice, although a NO donor elicited ATP release. Insulin-induced ATP release from human platelets correlated with the association of syntaxin 2 with the vesicle-associated membrane protein 3 but was not associated with the activation of alphaIIbbeta3 integrin. Thus, insulin elicits the release of vasoactive concentrations of ATP and adenosine from human platelets via a NO-G kinase-dependent signaling cascade. The mechanism of dense granule secretion involves the G kinase-dependent association of syntaxin 2 with vesicle-associated membrane protein 3.

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Effect of insulin on serotonin release from α-granules, the phosphorylation of VASP, and the association of VAMP-3 with syntaxin 2. (a) Effect of L-NA (300 μmol/L) on the insulin-induced release of serotonin from washed human platelets. Values are presented relative to the serotonin levels detected in the supernatant from platelets treated with solvent. The supernatant-induced relaxation of porcine coronary artery rings was assessed, and the results were divided into two categories, i.e., responders and nonresponders. (b) Effect of solvent (CTL), thrombin (Thr, 0.1 U/ml), and the NO donor DETA-NONOate (NO,1 μmol/L) on the association of VAMP-3 with syntaxin 2, immunoprecipitated from washed human platelets. (c) Effect of solvent (CTL) and insulin (1 μmol/L) on the association of VAMP-3 with syntaxin 2, immunoprecipitated from washed human platelets. Experiments were performed in the absence and presence of L-NA and the G kinase inhibitor Rp-8CPT-cGMPs (Rp, 10 μmol/L). The phosphorylation of VASP (Ser239 VASP), as a marker of NOS activation, was also determined in the Triton X-100–soluble fraction of the same platelets preparations. To demonstrate the equal loading of each lane, the blots were reprobed with antibodies recognizing either syntaxin 2 or total VASP protein. The bar graph (d) summarizes the results of VAMP-3 association with syntaxin 2 obtained in seven independent experiments. *P < 0.05 and **P < 0.01 versus unstimulated platelets (CTL and solvent).
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fig5: Effect of insulin on serotonin release from α-granules, the phosphorylation of VASP, and the association of VAMP-3 with syntaxin 2. (a) Effect of L-NA (300 μmol/L) on the insulin-induced release of serotonin from washed human platelets. Values are presented relative to the serotonin levels detected in the supernatant from platelets treated with solvent. The supernatant-induced relaxation of porcine coronary artery rings was assessed, and the results were divided into two categories, i.e., responders and nonresponders. (b) Effect of solvent (CTL), thrombin (Thr, 0.1 U/ml), and the NO donor DETA-NONOate (NO,1 μmol/L) on the association of VAMP-3 with syntaxin 2, immunoprecipitated from washed human platelets. (c) Effect of solvent (CTL) and insulin (1 μmol/L) on the association of VAMP-3 with syntaxin 2, immunoprecipitated from washed human platelets. Experiments were performed in the absence and presence of L-NA and the G kinase inhibitor Rp-8CPT-cGMPs (Rp, 10 μmol/L). The phosphorylation of VASP (Ser239 VASP), as a marker of NOS activation, was also determined in the Triton X-100–soluble fraction of the same platelets preparations. To demonstrate the equal loading of each lane, the blots were reprobed with antibodies recognizing either syntaxin 2 or total VASP protein. The bar graph (d) summarizes the results of VAMP-3 association with syntaxin 2 obtained in seven independent experiments. *P < 0.05 and **P < 0.01 versus unstimulated platelets (CTL and solvent).

Mentions: To determine whether or not the ATP/adenosine was derived from dense granules, we determined whether serotonin, a classical marker of dense granules, was also released from insulin-stimulated platelets. Again, insulin elicited the release of serotonin from those platelets that also generated a relaxing factor but not from platelets which failed to generate a relaxing factor (Fig. 5 a).


Insulin induces the release of vasodilator compounds from platelets by a nitric oxide-G kinase-VAMP-3-dependent pathway.

Randriamboavonjy V, Schrader J, Busse R, Fleming I - J. Exp. Med. (2004)

Effect of insulin on serotonin release from α-granules, the phosphorylation of VASP, and the association of VAMP-3 with syntaxin 2. (a) Effect of L-NA (300 μmol/L) on the insulin-induced release of serotonin from washed human platelets. Values are presented relative to the serotonin levels detected in the supernatant from platelets treated with solvent. The supernatant-induced relaxation of porcine coronary artery rings was assessed, and the results were divided into two categories, i.e., responders and nonresponders. (b) Effect of solvent (CTL), thrombin (Thr, 0.1 U/ml), and the NO donor DETA-NONOate (NO,1 μmol/L) on the association of VAMP-3 with syntaxin 2, immunoprecipitated from washed human platelets. (c) Effect of solvent (CTL) and insulin (1 μmol/L) on the association of VAMP-3 with syntaxin 2, immunoprecipitated from washed human platelets. Experiments were performed in the absence and presence of L-NA and the G kinase inhibitor Rp-8CPT-cGMPs (Rp, 10 μmol/L). The phosphorylation of VASP (Ser239 VASP), as a marker of NOS activation, was also determined in the Triton X-100–soluble fraction of the same platelets preparations. To demonstrate the equal loading of each lane, the blots were reprobed with antibodies recognizing either syntaxin 2 or total VASP protein. The bar graph (d) summarizes the results of VAMP-3 association with syntaxin 2 obtained in seven independent experiments. *P < 0.05 and **P < 0.01 versus unstimulated platelets (CTL and solvent).
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Related In: Results  -  Collection

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fig5: Effect of insulin on serotonin release from α-granules, the phosphorylation of VASP, and the association of VAMP-3 with syntaxin 2. (a) Effect of L-NA (300 μmol/L) on the insulin-induced release of serotonin from washed human platelets. Values are presented relative to the serotonin levels detected in the supernatant from platelets treated with solvent. The supernatant-induced relaxation of porcine coronary artery rings was assessed, and the results were divided into two categories, i.e., responders and nonresponders. (b) Effect of solvent (CTL), thrombin (Thr, 0.1 U/ml), and the NO donor DETA-NONOate (NO,1 μmol/L) on the association of VAMP-3 with syntaxin 2, immunoprecipitated from washed human platelets. (c) Effect of solvent (CTL) and insulin (1 μmol/L) on the association of VAMP-3 with syntaxin 2, immunoprecipitated from washed human platelets. Experiments were performed in the absence and presence of L-NA and the G kinase inhibitor Rp-8CPT-cGMPs (Rp, 10 μmol/L). The phosphorylation of VASP (Ser239 VASP), as a marker of NOS activation, was also determined in the Triton X-100–soluble fraction of the same platelets preparations. To demonstrate the equal loading of each lane, the blots were reprobed with antibodies recognizing either syntaxin 2 or total VASP protein. The bar graph (d) summarizes the results of VAMP-3 association with syntaxin 2 obtained in seven independent experiments. *P < 0.05 and **P < 0.01 versus unstimulated platelets (CTL and solvent).
Mentions: To determine whether or not the ATP/adenosine was derived from dense granules, we determined whether serotonin, a classical marker of dense granules, was also released from insulin-stimulated platelets. Again, insulin elicited the release of serotonin from those platelets that also generated a relaxing factor but not from platelets which failed to generate a relaxing factor (Fig. 5 a).

Bottom Line: Insulin failed to relax endothelium-intact rings of porcine coronary artery.Insulin induced the release of adenosine trisphosphate (ATP), adenosine, and serotonin from platelet-dense granules in a NO-dependent manner.The mechanism of dense granule secretion involves the G kinase-dependent association of syntaxin 2 with vesicle-associated membrane protein 3.

View Article: PubMed Central - PubMed

Affiliation: Institut für Kardiovaskuläre Physiologie, Klinikum der J.W.Goethe-Universität, D-60590 Frankfurt am Main, Germany.

ABSTRACT
Insulin-induced vasodilatation is sensitive to nitric oxide (NO) synthase (NOS) inhibitors. However, insulin is unable to relax isolated arteries or to activate endothelial NOS in endothelial cells. Since insulin can enhance platelet endothelial NOS activity, we determined whether insulin-induced vasodilatation can be attributed to a NO-dependent, platelet-mediated process. Insulin failed to relax endothelium-intact rings of porcine coronary artery. The supernatant from insulin-stimulated human platelets induced complete relaxation, which was prevented by preincubation of platelets with a NOS inhibitor, the soluble guanylyl cyclase inhibitor, NS 2028, or the G kinase inhibitor, KT 5823, and was abolished by an adenosine A2A receptor antagonist. Insulin induced the release of adenosine trisphosphate (ATP), adenosine, and serotonin from platelet-dense granules in a NO-dependent manner. This response was not detected using insulin-stimulated platelets from endothelial NOS-/- mice, although a NO donor elicited ATP release. Insulin-induced ATP release from human platelets correlated with the association of syntaxin 2 with the vesicle-associated membrane protein 3 but was not associated with the activation of alphaIIbbeta3 integrin. Thus, insulin elicits the release of vasoactive concentrations of ATP and adenosine from human platelets via a NO-G kinase-dependent signaling cascade. The mechanism of dense granule secretion involves the G kinase-dependent association of syntaxin 2 with vesicle-associated membrane protein 3.

Show MeSH
Related in: MedlinePlus