Sympathetic nervous system controls total peripheral vascular resistance and blood flow via regulation of the contraction of small arteries. One of the principal sympathetic neurotransmitters, ATP, acts on arterial smooth muscle cells (SMCs) via activation of P2X receptors (P2XRs) leading to an increase of [Ca2+]i and SMC contraction. We have recently demonstrated that phasic contractions of the guinea-pig small mesenteric arteries induced by P2XR stimulation are sensitive to inhibitors of voltage-gated Ca2+ channels (VGCCs), ryanodine receptors (RyRs) and inositol 1,4,5-trisphosphate receptors (IP3Rs) [1]. Here we analysed the respective role of RyRs and IP3Rs in elevation of [Ca2+]i induced by selective stimulation P2XRs in smooth muscle cells (SMCs) from the guinea-pig small mesenteric arteries. Data are presented as mean ± S.E.M. and compared using Student’s t-test. Freshly isolated fluo-3-loaded SMCs were stimulated with 10 µM α,β-methyleneadenosine 5-triphosphate (αβ-meATP). Fast x-y confocal imaging revealed that activation of P2XRs evoked an abrupt sub-plasmalemmal (sub-PM) [Ca2+]i upstroke (SPCU) followed by Ca2+ wave rapidly propagating towards SMC center. The effect of IP3R inhibition (30 µM 2-APB) was significantly (p<0.00001) stronger than that of RyR inhibition (100 µM tetracaine): reduction of the SPCU amplitude by 67±7% (n=16) and by 40±5% (n=26), respectively. The effect of simultaneous block of IP3Rs and RyRs was similar (p<0.001) to the effects of calcium store depletion with 10 µM CPA: attenuation of the response by 93±2% (n=7) and by 81±3% (n=24), respectively. The effect IP3R inhibition on SPCU was significantly (p<0.0001) attenuated by initial block of VGCCs (5 µM nicardipine): under these conditions 2-APB reduced SCPU amplitude only by 37±3% (n=9), thus, suggesting that IP3R-mediated Ca2+ release is facilitated by Ca2+ entry via VGCCs. Immunostaining of RyRs and IP3Rs in the SMCs with identified sarcoplasmic reticulum (SR) and nucleus revealed that sub-PM SR elements are enriched with type 1 IP3Rs while RyRs are located mainly in deeper SR. This structural peculiarity makes IP3Rs more accessible to Ca2+ entering the cell via VGCCs. As a result, IP3Rs may serve as a functional link (“intermediate amplifier”) between voltage-gated Ca2+ entry and RyR-mediated Ca2+ release. We conclude that depolarization of mesenteric artery SMCs following P2X receptor activation induces IP3R-mediated Ca2+ release from sub-PM SR, which is facilitated Ca2+ entry via VGCCs. This mechanism seems to be fundamental (as we have recently demonstrated this in renal microvascular SMCs [2]) and provides convergence of signalling pathways engaged in electromechanical and pharmacomechanical coupling [3] in vascular smooth muscle.