Role of Phosphatidylinositol 3-Kinase (PI3K), Mitogen-Activated Protein Kinase (MAPK), and Protein Kinase C (PKC) in Calcium Signaling Pathways Linked to the α-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 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 signaling pathways linked to adrenergic vasoconstriction in resistance arteries. Simultaneous measurements of intracellular calcium concentration ([Ca]) 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 were assessed in arteries activated with phenylephrine (PE) and kept in Ca-free solution, in the absence and presence of the selective antagonist of L-type Ca channels nifedipine, cyclopiazonic acid (CPA) to block sarcoplasmic reticulum (SR) intracellular Ca release or specific inhibitors of PI3K, ERK-MAPK, or PKC. Activation of α-adrenoceptors with PE stimulated both intracellular Ca mobilization and Ca entry along with contraction in resistance arteries. Both [Ca] and contractile responses were inhibited by nifedipine while CPA abolished intracellular Ca mobilization and modestly reduced Ca entry suggesting that α-adrenergic vasoconstriction is largely dependent Ca influx through L-type Ca channel and to a lesser extent through store-operated Ca channels. Inhibition of ERK-MAPK did not alter intracellular Ca mobilization but largely reduced L-type Ca entry elicited by PE without altering vasoconstriction. The PI3K blocker LY-294002 moderately reduced intracellular Ca release, Ca entry and contraction induced by the α-adrenoceptor agonist, while PKC inhibition decreased PE-elicited Ca entry and to a lesser extent contraction without affecting intracellular Ca mobilization. Under conditions of ryanodine receptor (RyR) blockade to inhibit Ca-induced Ca-release (CICR), inhibitors of PI3K, ERK-MAPK, or PKC significantly reduced [Ca] increases but not contraction elicited by high K depolarization suggesting an activation of L-type Ca entry in VSM independent of RyR. In summary, our results demonstrate that PI3K, ERK-MAPK, and PKC regulate Ca handling coupled to the α-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 handling such as obesity, hypertension and diabetes mellitus, in which these signaling pathways are profoundly impaired.