Optogenetic regulation of PIP2 alters the voltage-dependence of activation and fast inactivation of skeletal muscle Nav1.4 channels.

Abstract

Since the late 1970's phosphatidylinositol-4,5-bisphosphate PI(4,5)P2 has garnered much attention for its role as a substrate that initiates the production of both inositol-1,4,5-triphosphate (IP3) and 1,2-diacylglycerol (DAG). These secondary messengers regulate the cellular function of many ion channels including voltage-gated sodium channels (NaV). In addition to this, PI(4,5)P2 can recruit and modulate the activity of many ion channels. However, to date, it is yet to be elucidated whether PI(4,5)P2 modulates the activity of NaV channels. NaV channels are critical in the initiation and propagation of action potentials in excitable cells. The voltage-dependent activation and instantaneous fast inactivation gating are tightly regulated as they set the refractory period in excitable cells. Changes in this regulation lead to various sodium channelopathies. With simultaneous use of both patch-clamp and optogenetic techniques, here, we report the effect of dephosphorylation of PI(4,5)P2 by specific phosphoinositide (PI)- phosphatases on Nav1.4 gating. In this study, we utilized the 4’ phosphatase Sac2 (CRY2-Sac2) or the 5’ phosphatase CRY2-5’PtaseOCRL (CRY2-5’POCRL) to show that dephosphorylating PI(4,5)P2 left-shifts the voltage-dependent gating of NaV1.4, the skeletal muscle channel, to more hyperpolarized membrane potentials. This data provides evidence that PI(4,5)P2 is important for the gating of NaV1.4 channels.