Physical coupling between inositol 1,4,5-trisphosphate (IP3) receptors and transient receptor potential (Trp) channels has been demonstrated in both transfected and normal cells as a candidate mechanism for the activation of store-mediated Ca2+ entry (SMCE). We have investigated the properties of the coupling between the type II IP3 receptor and naturally expressed human Trp1 (hTrp1) in human platelets. Treatment with xestospongin C, an inhibitor of IP3 receptor function, abolished SMCE and coupling between the IP3 receptor and hTrp1. The coupling was activated by depletion of the intracellular Ca2+ stores, and was reversed by refilling of the stores. We have also examined the role of actin filaments in the activation and maintenance of the coupling. Stabilization of the cortical actin network with jasplakinolide prevented the coupling, indicating that, as with secretion, the actin filaments at the cell periphery act as a negative clamp which prevents constitutive coupling. In addition, the actin cytoskeleton plays a positive role, since disruption of the actin network inhibited the coupling when the Ca2+ stores were depleted. These results provide strong evidence for the activation of SMCE by a secretion-like coupling mechanism involving a reversible association between IP3 receptors and hTrp1 in normal human cells.

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