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

The ubiquitous inositol 1,4,5-trisphosphate (InsP3) receptor (InsP3R) channel, localized primarily in the endoplasmic reticulum (ER) membrane, releases Ca2+ into the cytoplasm upon binding InsP3, 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 (iCa) passing through an open InsP3R 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, iCa for InsP3R channels in physiological ionic conditions has not been directly measured. Here, we report the first measurement of iCa through an InsP3R 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 InsP3R 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 iCa–[Ca2+]ER relation was linear, with no detectable dependence on [Mg2+]f. iCa was 0.15 ± 0.01 pA for a filled ER store with 500 µM [Ca2+]ER. The iCa–[Ca2+]ER relation suggests that Ca2+ released by an InsP3R channel raises [Ca2+]i near the open channel to ∼13–70 µM, depending on [Ca2+]ER. These measurements have implications for the activities of nearby InsP3-liganded InsP3R channels, and they confirm that Ca2+ released by an open InsP3R channel is sufficient to activate neighboring channels at appropriate distances away, promoting Ca2+-induced Ca2+ release.