The down-regulation of alpha-adrenoceptor-mediated signaling casacade has been implicated in obesity but the underlying mechanism remains largely unknown. The present study investigated whether inositol 1,4,5-trisphosphate (IP3) receptor and protein kinase C (PKC) were involved in the reduction of alpha(1)-adrenoceptor agonist phenylephrine-evoked contraction in aortae of high fat diet-induced obese (DIO) mice. C57BL/6 mice were fed with a rodent diet containing 45 kcal% fat for 16 weeks to induce obesity. Isolated mouse aortae were suspended in myograph for isometric force measurement. Protein phosphorylations and expressions were determined by Western blotting. In C57BL/6 mouse aortae, phenylephrine-induced contraction was partially inhibited by either IP3 receptor antagonist heparin or PKC inhibitor GFX, and the combined treatment with heparin and GFX abolished the contraction. Phenylephrine-induced contraction was significantly less in the aortae of 010 mice than those of control mice; only GFX but not heparin attenuated the contraction, indicating a diminishing role of IP3 receptor in DIO mice. Western blotting showed the reduced expression and phosphorylation of IP3 receptor and the down-regulated expression of PKC, PKC beta, PKCS, and PKC( in DIO mouse aortae. Importantly, PKCS was more likely to maintain phenylephrine-mediated contraction in DIO mouse aortae because that (1) PKC delta inhibitor rottlerin but not PKC alpha and PKC beta inhibitor Go6976, PKC beta inhibitor hispidin, or PKC zeta pseudosubstrate inhibitor attenuated the contraction; and (2) PKCS phosphorylation was increased but phosphorylations of PKC alpha, PKC beta, and PKC zeta were reduced in DIO mouse aortae. The present study thus provides additional insights into the cellular mechanisms responsible for vascular dysfunction in obesity. (C) 2014 Elsevier Inc. All rights reserved.