Rapid Release of Ca from Endoplasmic Reticulum Mediated by Na/Ca Exchange.

Phototransduction in is mediated by phospholipase C (PLC) and Ca-permeable TRP channels, but the function of endoplasmic reticulum (ER) Ca stores in this important model for Ca signaling remains obscure. We therefore expressed a low affinity Ca indicator (ER-GCaMP6-150) in the ER, and measured its fluorescence both in dissociated ommatidia and from intact flies of both sexes. Blue excitation light induced a rapid (tau ∼0.8 s), PLC-dependent decrease in fluorescence, representing depletion of ER Ca stores, followed by a slower decay, typically reaching ∼50% of initial dark-adapted levels, with significant depletion occurring under natural levels of illumination. The ER stores refilled in the dark within 100-200 s. Both rapid and slow store depletion were largely unaffected in InsP receptor mutants, but were much reduced in mutants. Strikingly, rapid (but not slow) depletion of ER stores was blocked by removing external Na and in mutants of the Na/Ca exchanger, CalX, which we immuno-localized to ER membranes in addition to its established localization in the plasma membrane. Conversely, overexpression of greatly enhanced rapid depletion. These results indicate that rapid store depletion is mediated by Na/Ca exchange across the ER membrane induced by Na influx via the light-sensitive channels. Although too slow to be involved in channel activation, this Na/Ca exchange-dependent release explains the decades-old observation of a light-induced rise in cytosolic Ca in photoreceptors exposed to Ca-free solutions. Phototransduction in is mediated by phospholipase C, which activates TRP cation channels by an unknown mechanism. Despite much speculation, it is unknown whether endoplasmic reticulum (ER) Ca stores play any role. We therefore engineered flies expressing a genetically encoded Ca indicator in the photoreceptor ER. Although NCX Na/Ca exchangers are classically believed to operate only at the plasma membrane, we demonstrate a rapid light-induced depletion of ER Ca stores mediated by Na/Ca exchange across the ER membrane. This NCX-dependent release was too slow to be involved in channel activation, but explains the decades-old observation of a light-induced rise in cytosolic Ca in photoreceptors bathed in Ca-free solutions.