Image_1_Role of Phosphatidylinositol 3-Kinase (PI3K), Mitogen-Activated Protein Kinase (MAPK), and Protein Kinase C (PKC) in Calcium Signaling Pathways Linked to the α1-Adrenoceptor in Resistance Arteries.JPEG

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 Ca²⁺ 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 Ca²⁺ signaling pathways linked to adrenergic vasoconstriction in resistance arteries. Simultaneous measurements of intracellular calcium concentration ([Ca²⁺]_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 CaCl₂ were assessed in arteries activated with phenylephrine (PE) and kept in Ca²⁺-free solution, in the absence and presence of the selective antagonist of L-type Ca²⁺ channels nifedipine, cyclopiazonic acid (CPA) to block sarcoplasmic reticulum (SR) intracellular Ca²⁺ release or specific inhibitors of PI3K, ERK-MAPK, or PKC. Activation of α₁-adrenoceptors with PE stimulated both intracellular Ca²⁺ mobilization and Ca²⁺ entry along with contraction in resistance arteries. Both [Ca²⁺]_i and contractile responses were inhibited by nifedipine while CPA abolished intracellular Ca²⁺ mobilization and modestly reduced Ca²⁺ entry suggesting that α₁-adrenergic vasoconstriction is largely dependent Ca²⁺ influx through L-type Ca²⁺ channel and to a lesser extent through store-operated Ca²⁺ channels. Inhibition of ERK-MAPK did not alter intracellular Ca²⁺ mobilization but largely reduced L-type Ca²⁺ entry elicited by PE without altering vasoconstriction. The PI3K blocker LY-294002 moderately reduced intracellular Ca²⁺ release, Ca²⁺ entry and contraction induced by the α₁-adrenoceptor agonist, while PKC inhibition decreased PE-elicited Ca²⁺ entry and to a lesser extent contraction without affecting intracellular Ca²⁺ mobilization. Under conditions of ryanodine receptor (RyR) blockade to inhibit Ca²⁺-induced Ca²⁺-release (CICR), inhibitors of PI3K, ERK-MAPK, or PKC significantly reduced [Ca²⁺]_i increases but not contraction elicited by high K⁺ depolarization suggesting an activation of L-type Ca²⁺ entry in VSM independent of RyR. In summary, our results demonstrate that PI3K, ERK-MAPK, and PKC regulate Ca²⁺ handling coupled to the α₁-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 Ca²⁺ handling such as obesity, hypertension and diabetes mellitus, in which these signaling pathways are profoundly impaired.