Contribution of both Ca2+entry and Ca2+sensitization to the α1-adrenergic vasoconstriction of rat penile small arteries

Abstract

Sympathetic adrenergic nerves maintain the flaccid state of the penis through the tonic release of norepinephrine that contracts trabecular and arterial smooth muscle. Simultaneous measurements of intracellular Ca2+concentration ([Ca2+]i) and tension and experiments with α-toxin-permeabilized arteries were performed in branches of the rat dorsal penile artery to investigate the intracellular Ca2+signaling pathways underlying α1-adrenergic vasoconstriction. Phenylephrine increased both [Ca2+]iand tension, these increases being abolished by extracellular Ca2+removal and reduced by about 50% by the L-type Ca2+channel blocker nifedipine (0.3 μM). Non-L-type Ca2+entry through store-operated channels was studied by inhibiting the sarcoplasmic reticulum Ca2+-ATPase with cyclopiazonic acid (CPA). CPA (30 μM) induced variable phasic contractions that were abolished by extracellular Ca2+removal and by the store-operated channels antagonist 2-aminoethoxydiphenyl borate (2-APB, 50 μM) and largely inhibited by nifedipine (0.3 μM). CPA induced a sustained increase in [Ca2+]ithat was reduced in a Ca2+-free medium. Under conditions of L-type channels blockade, Ca2+readmission after store depletion with CPA evoked a sustained and marked elevation in [Ca2+]inot coupled to contraction. 2-APB (50 μM) inhibited the rise in [Ca2+]ievoked by CPA and the nifedipine-insensitive increases in both [Ca2+]iand contraction elicited by phenylephrine. In α-toxin-permeabilized penile arteries, activation of G proteins with guanosine 5′- O-(3-thiotriphosphate) and of the α1-adrenoceptor with phenylephrine both enhanced the myofilament sensitivity to Ca2+. This Ca2+sensitization was reduced by selective inhibitors of PKC, tyrosine kinase (TK), and Rho kinase (RhoK) by 43%, 67%, and 82%, respectively. As a whole, the present data suggest the α1-adrenergic vasoconstriction in penile small arteries involves Ca2+entry through both L-type and 2-APB-sensitive receptor-operated channels, as well as Ca2+sensitization mechanisms mediated by PKC, TK, and RhoK. A capacitative Ca2+entry coupled to noncontractile functions of the smooth muscle cell is also demonstrated.