Inositol 1,4,5-tris-phosphate activation of inositol tris-phosphate receptor Ca2+ channel by ligand tuning of Ca2+ inhibition

Inositol 1,4,5-tris-phosphate (IP3) binding to its receptors (IP3R) in the endoplasmic reticulum (ER) activates Ca2+ release from the ER lumen to the cytoplasm, generating complex cytoplasmic Ca2+ concentration signals including temporal oscillations and propagating waves. IP3-mediated Ca2+ release is also controlled by cytoplasmic Ca2+ concentration with both positive and negative feedback. Single-channel properties of the IP3R in its native ER membrane were investigated by patch clamp electrophysiology of isolated Xenopus oocyte nuclei to determine the dependencies of IP3R on cytoplasmic Ca2+ and IP3 concentrations under rigorously defined conditions. Instead of the expected narrow bell-shaped cytoplasmic free Ca2+ concentration ([Ca2+](i)) response centered at approximate to 300 nM-1 mu M, the open probability remained elevated (approximate to 0.8) in the presence of saturating levels (10 mu M) of IP3, even as [Ca2+]i was raised to high concentrations, displaying two distinct types of functional Ca2+ binding sites: activating sites with half-maximal activating [Ca2+](i) (K-act) of 210 nM and Hill coefficient (H-act) approximate to 2; and inhibitory sites with half-maximal inhibitory [Ca2+](i) (K-inh) Of 54 mu M and Hill coefficient (H-inh) approximate to 4 Lowering IP3 concentration was without effect on Ca2+ activation parameters or H-inh, but decreased K-inh with a functional half-maximal activating IP3 concentration (K-IP3) of 50 nM and Hill coefficient (H-IP3) of 4 for IP3, These results demonstrate that Ca2+ is a true receptor agonist, whereas the sole function of IP3 is to relieve Ca2+ inhibition of IP3R. Allosteric tuning of Ca2+ inhibition by IP3 enables the individual IP3R Ca2+ channel to respond in a graded fashion, which has implications for localized and global cytoplasmic Ca2+ concentration signaling and quantal Ca2+ release.