We tested an involvement of ryanodine receptors (RyRs), inositol 1,4,5-trisphosphate receptors (IP3₃Rs) and voltage-gated Ca²⁺ channels (VGCCs) in [Ca²⁺]_i mobilisation induced by stimulation of P₂X purinoceptors (P₂XRs) in smooth muscle cells (SMCs) from the guinea-pig small mesenteric arteries. Freshly isolated SMCs were loaded with the high affinity Ca²⁺ indicator fluo-3 and stimulated with 10 µM α,β-methyleneadenosine 5-triphosphate (α,β-meATP). Fast x-y confocal imaging revealed that activation of P₂XRs evoked an initial sub-plasmalemmal [Ca²⁺]_i upstroke (SPCU) at several restricted locations followed by propagating Ca²⁺ wave. The peak amplitude of α,β-meATP-induced SPCU was reduced: (1) by 37% (n=6) after inhibition of IP₃Rs (with 30 µM 2APB); (2) by 45% (n=6) after inhibition of RyRs (with 100 µM tetracaine) and (3) by 68% (n=6) after block of VGCCs (with 5 µM nicardipine). Although activation of P₂XRs does not engage metabotropic signalling pathway (involving activation of phospholipase C and intracellular mobilisation of IP₃), a contribution of IP₃Rs-mediated Ca²⁺ release to purinergic contractions was further demonstrated by isometric tension recording, in which smooth muscle rings from segments of second- or third-order mesenteric arteries were attached to isometric force transducer and stimulated with 10 µM α,β-meATP. Inhibition of either IP₃Rs or RyRs attenuated purinergic contraction. When VGCCs were inhibited with 5μM nicardipine, a relative contribution of RyRs-mediated Ca²⁺ release to purinergic contraction increased, thus suggesting that Ca²⁺ influx through VGCCs evokes predominantly IP₃Rs-mediated Ca²⁺ release. Immunostaining of RyRs and type 1 IP₃Rs in the isolated SMCs with identified SR and nucleus revealed that RyRs are predominantly located in the central/perinuclear SR elements, while sub-PM SR elements are enriched with IP₃Rs. These results suggest that not only Ca²⁺ entry through VGCCs but also RyRs- and IPP₃XRs-mediated Ca²⁺ release are involved in genesis of SPCU and contraction in response to P₂XR stimulation.