INSULIN INHIBITS VASCULAR SMOOTH-MUSCLE CONTRACTION AT A SITE DISTAL TO INTRACELLULAR CA2+ CONCENTRATION

Several hypertensive states are associated with resistance to insulin- induced glucose disposal and insulin-induced vasodilation. Insulin can inhibit vascular smooth muscle (VSM) contraction at the level of the VSM cell, and resistance to insulin's inhibition of VSM cell contracti on may be of pathophysiological importance. To understand the VSM cell ular mechanisms by which insulin resistance leads to increased VSM con traction, we sought to determine how insulin inhibits contraction of n ormal VSM. It has been shown that insulin lowers the contractile agoni st-stimulated intracellular Ca2+ (Ca-i(2+)) transient in VSM cells. In this study, our goal was to see whether insulin inhibits VSM cell con traction at steps distal to Ca-i(2+) and, if so, to determine whether the mechanism is dependent on nitric oxide synthase (NOS) and cGMP Pri mary cultured VSM cells from canine femoral artery were bathed in a ph ysiological concentration of extracellular Ca2+ and permeabilized to C a2+ with a Ca2+ ionophore, either ionomycin or A-23187. The resultant increase in Ca-i(2+) contracted individual cells, as measured by photo microscopy. Preincubating cells with 1 nM insulin for 30 min did not a ffect basal Ca-i(2+) or the ionomycin-induced increase in Ca-i(2+), as determined by fura 2 fluorescence measurements, but it did inhibit io nomycin-and A-23187-induced contractions by 47 and 51%, respectively ( both P < 0.05). In the presence of 1.0 mu M ionized C-a2+ ionomycin-in duced contractions were inhibited by insulin in a dose-dependent manne r. In the presence of ionomycin, insulin increased cGMP production by 43% (P < 0.05). 1H-[1,2,4]oxadiazolo[4,3-a]quinoxalin-1-one (10 mu M), a selective inhibitor of guanylate cyclase that blocked cGMP producti on in these cells, completely blocked the inhibition by insulin of ion omycin-induced contraction. It was found that the cells expressed the inducible isoform of NOS. N-G-monomethyl-L-arginine or N-G-nitro-L-arg inine methyl ester (0.1 mM), inhibitors of NOS, did not affect ionomyc in-induced contraction but prevented insulin from inhibiting contracti on. We conclude that insulin stimulates cGMP production and inhibits V SM contraction in the presence of elevated Ca-i(2+) This inhibition by insulin of VSM contraction at sites where Ca-i(2+) could not be rate limiting is dependent on NOS and cGMP.