Regulation of Cx45 hemichannels mediated by extracellular and intracellular calcium

Abstract

Connexin45 (Cx45) hemichannels (HCs) open in the absence of Ca²⁺ and close in its presence. To elucidate the underlying mechanisms, we examined the role of extra- and intracellular Ca²⁺ on the electrical properties of HCs. Experiments were performed on HeLa cells expressing Cx45 using electrical (voltage clamp) and optical (Ca²⁺ imaging) methods. HCs exhibit a time- and voltage-dependent current (I _hc), activating with depolarization and inactivating with hyperpolarization. Elevation of [Ca²⁺]_o from 20 nM to 2 μM reversibly decreases I _hc, decelerates its rate of activation, and accelerates its deactivation. Our data suggest that [Ca²⁺]_o modifies the channel properties by adhering to anionic sites in the channel lumen and/or its outer vestibule. In this way, it blocks the channel pore and reversibly lowers I _hc and modifies its kinetics. Rapid lowering of [Ca²⁺]_o from 2 mM to 20 nM, achieved early during a depolarizing pulse, led to an outward I _hc that developed with virtually no delay and grew exponentially in time paralleled by unaffected [Ca²⁺]_i. A step increase of [Ca²⁺]_i evoked by photorelease of Ca²⁺ early during a depolarizing pulse led to a transient decrease of I _hc superimposed on a growing outward I _hc; a step decrease of [Ca²⁺]_i elicited by photoactivation of a Ca²⁺ scavenger provoked a transient increase in I _hc. Hence, it is tempting to assume that Ca²⁺ exerts a direct effect on Cx45 hemichannels.