Role of Phosphatidylinositol 3-Kinase (PI3K), Mitogen-Activated Protein Kinase (MAPK), and Protein Kinase C (PKC) in Calcium Signaling Pathways Linked to the α(1)-Adrenoceptor in Resistance Arteries

Insulin resistance plays a key role in the pathogenesis of type 2 diabetes and is also related to other health problems like obesity, hypertension, and metabolic syndrome. Imbalance between insulin vascular actions via the phosphatidylinositol 3-Kinase (PI3K) and the mitogen activated protein kinase (MAPK) signaling pathways during insulin resistant states results in impaired endothelial PI3K/eNOS- and augmented MAPK/endothelin 1 pathways leading to endothelial dysfunction and abnormal vasoconstriction. The role of PI3K, MAPK, and protein kinase C (PKC) in Ca(2+) handling of resistance arteries involved in blood pressure regulation is poorly understood. Therefore, we assessed here whether PI3K, MAPK, and PKC play a role in the Ca(2+) signaling pathways linked to adrenergic vasoconstriction in resistance arteries. Simultaneous measurements of intracellular calcium concentration ([Ca(2+)](i)) in vascular smooth muscle (VSM) and tension were performed in endothelium-denuded branches of mesenteric arteries from Wistar rats mounted in a microvascular myographs. Responses to CaCl(2) were assessed in arteries activated with phenylephrine (PE) and kept in Ca(2+)-free solution, in the absence and presence of the selective antagonist of L-type Ca(2+) channels nifedipine, cyclopiazonic acid (CPA) to block sarcoplasmic reticulum (SR) intracellular Ca(2+) release or specific inhibitors of PI3K, ERK-MAPK, or PKC. Activation of α(1)-adrenoceptors with PE stimulated both intracellular Ca(2+) mobilization and Ca(2+) entry along with contraction in resistance arteries. Both [Ca(2+)](i) and contractile responses were inhibited by nifedipine while CPA abolished intracellular Ca(2+) mobilization and modestly reduced Ca(2+) entry suggesting that α(1)-adrenergic vasoconstriction is largely dependent Ca(2+) influx through L-type Ca(2+) channel and to a lesser extent through store-operated Ca(2+) channels. Inhibition of ERK-MAPK did not alter intracellular Ca(2+) mobilization but largely reduced L-type Ca(2+) entry elicited by PE without altering vasoconstriction. The PI3K blocker LY-294002 moderately reduced intracellular Ca(2+) release, Ca(2+) entry and contraction induced by the α(1)-adrenoceptor agonist, while PKC inhibition decreased PE-elicited Ca(2+) entry and to a lesser extent contraction without affecting intracellular Ca(2+) mobilization. Under conditions of ryanodine receptor (RyR) blockade to inhibit Ca(2+)-induced Ca(2+)-release (CICR), inhibitors of PI3K, ERK-MAPK, or PKC significantly reduced [Ca(2+)](i) increases but not contraction elicited by high K(+) depolarization suggesting an activation of L-type Ca(2+) entry in VSM independent of RyR. In summary, our results demonstrate that PI3K, ERK-MAPK, and PKC regulate Ca(2+) handling coupled to the α(1)-adrenoceptor in VSM of resistance arteries and related to both contractile and non-contractile functions. These kinases represent potential pharmacological targets in pathologies associated to vascular dysfunction and abnormal Ca(2+) handling such as obesity, hypertension and diabetes mellitus, in which these signaling pathways are profoundly impaired.