Facilitatory effect of Ins(1,4,5)P₃ on store-operated Ca²⁺-permeable cation channels in rabbit portal vein myocytes

In rabbit portal vein smooth muscle cells, store-operated Ca²⁺-permeable cation channels (SOCs) display multi-modal gating mechanisms. SOCs are activated by depletion of intracellular Ca²⁺ stores but also may be stimulated in a store-independent manner by noradrenaline acting on α-adrenoceptors and by diacylglycerol (DAG) via protein kinase C (PKC). In the present study we have investigated whether inositol 1,4,5-trisphosphate (Ins(1,4,5)P₃) modulates SOC activity in freshly dispersed rabbit portal vein myocytes with patch pipette recording techniques. Inclusion of 1 μM Ins(1,4,5)P₃ in the patch pipette solution increased whole-cell currents evoked by the Ca²⁺-ATPase inhibitor cyclopiazonic acid (CPA) by about 3-fold at -80 mV. In the cell-attached configuration the cell-permeable Ca²⁺ chelator BAPTA-AM stimulated SOC activity and after excision of an isolated inside-out patch bath application of 1 μM Ins(1,4,5)P₃ increased open channel probability (NP_o) by approximately 3-fold. Ins(1,4,5)P₃ also produced a similar increase in NP_o of SOCs stimulated by the phorbol ester, phorbol 12,13-dibutyrate (PDBu) in inside-out patches and these channel currents had a unitary conductance of about 2 pS. The equilibrium constant of Ins(1,4,5)P₃ on increasing PDBu-evoked SOC activity was about 0.4 μM. The facilitatory effect of Ins(1,4,5)P₃ was also manifest as markedly increasing the rate of activation of SOCs. The synergistic effect of Ins(1,4,5)P₃ was mimicked by the metabolically stable analogue 3-fluoro-Ins(1,4,5)P₃ and Ins(1,4)P₂, a metabolite of Ins(1,4,5)P₃, but was not inhibited by the classical Ins(1,4,5)P₃ receptor antagonist heparin. Finally Ins(1,4,5)P₃ also increased NP_o of SOCs activated by a PKC catalytic subunit. It is concluded that Ins(1,4,5)P₃ facilitates SOC opening via a heparin-insensitive mechanism at, or dose to, the channel protein.