Endothelin antagonism uncovers insulin-mediated vasorelaxation in vitro and in vivo.

The endothelial actions of insulin remain an area of intense research because they relate to both insulin sensitivity and vascular tone. Physiological doses of insulin evoke endothelium-dependent vasorelaxation in humans; however, this remains a pharmacological phenomenon in rat aortas. Because insulin may stimulate the divergent production of both nitric oxide and endothelin-1, we hypothesized that the lack of insulin-induced vasorelaxation at low/subthreshold concentrations may be due to the concurrent production of endothelin-1, which in turn serves to inhibit nitric oxide-dependent, insulin-mediated dilation. To investigate this, we studied the effects of subthreshold concentrations of insulin (100 mU/L) on norepinephrine-induced contraction in rat aortas following short-term and long-term endothelin blockade. In addition, the effects of tetrahydrobiopterin inhibition (with diaminohydroxyprimidine) on norepinephrine-induced contraction in the presence of insulin and endothelin receptor blockade were investigated. Subthreshold concentrations of insulin failed to evoke vasorelaxation in rat aortas. Strikingly, short-term endothelin A/B receptor blockade with bosentan (10(-2) mmol/L) uncovered insulin-mediated dilation; the percent maximum contraction and sensitivity of aortas to norepinephrine were attenuated (% maximum relaxation: bosentan+insulin 74+/-4%* versus bosentan 92+/-3%, insulin 107+/-5% P:<0.002; pD(2) values: bosentan+insulin 6.87+/-0.14* versus bosentan 7.40+/-0.15, insulin 7.63+/-0.11, *P:<0.002). This effect was mediated through endothelin A receptors because bosentan and BQ-123 (10(-2) mmol/L) attenuated norepinephrine-induced contraction to a similar degree. In addition, insulin evoked vasorelaxation in aortas isolated from rats after long-term bosentan treatment (100 mg. kg(-1). d(-1), 3 weeks). The component of insulin-mediated vasorelaxation uncovered by endothelin receptor blockade was tetrahydrobiopterin-dependent because it was reversed by diaminohydroxyprimidine. These data demonstrate, for the first time, the functional interaction between insulin, endothelin-1, and tetrahydrobiopterin in modulating vascular tone in rat aortas in vitro and in vivo.