Unitary Ca2+ current through recombinant type 3 InsP3 receptor channels under physiological ionic conditions

The ubiquitous inositol 1,4,5-trisphosphate (InsP(3)) receptor (InsP(3)R) channel, localized primarily in the endoplasmic reticulum (ER) membrane, releases Ca2+ into the cytoplasm upon binding InsP(3), generating and modulating intracellular Ca2+ signals that regulate numerous physiological processes. Together with the number of channels activated and the open probability of the active channels, the size of the unitary Ca2+ current (i(Ca)) passing through an open InsP(3)R channel determines the amount of Ca2+ released from the ER store, and thus the amplitude and the spatial and temporal nature of Ca2+ signals generated in response to extracellular stimuli. Despite its significance, i(Ca) for InsP(3)R channels in physiological ionic conditions has not been directly measured. Here, we report the first measurement of i(Ca) through an InsP(3)R channel in its native membrane environment under physiological ionic conditions. Nuclear patch clamp electrophysiology with rapid perfusion solution exchanges was used to study the conductance properties of recombinant homotetrameric rat type 3 InsP(3)R channels. Within physiological ranges of free Ca2+ concentrations in the ER lumen ([Ca2+](ER)), free cytoplasmic [Ca2+] ([Ca2+](i)), and symmetric free [Mg2+] ([Mg2+](f)), the i(Ca)-[Ca2+](ER) relation was linear, with no detectable dependence on [Mg2+](f). i(Ca) was 0.15 +/- 0.01 pA for a filled ER store with 500 mu M [Ca2+](ER). The i(Ca)-[Ca2+](ER) relation suggests that Ca2+ released by an InsP(3)R channel raises [Ca2+](i) near the open channel to similar to 13-70 mu M, depending on [Ca2+](ER). These measurements have implications for the activities of nearby InsP(3)-liganded InsP(3)R channels, and they confirm that Ca2+ released by an open InsP(3)R channel is sufficient to activate neighboring channels at appropriate distances away, promoting Ca2+-induced Ca2+ release.