Timing of Ca(2+) release from intracellular stores and the electrical response of Limulus ventral photoreceptors to dim flashes.

Abstract

Light-induced release of Ca(2+) from stores in Limulus ventral photoreceptors was studied using confocal fluorescence microscopy and the Ca(2+) indicator dyes, Oregon green-5N and fluo-4. Fluorescence was collected from a spot within 4 microm of the microvillar membrane. A dual-flash protocol was used to reconstruct transient elevations of intracellular free calcium ion concentration (Ca(i)) after flashes delivering between 10 and 5 x 10(5) effective photons. Peak Ca(i) increased with flash intensity to 138 +/- 76 microM after flashes delivering approximately 10(4) effective photons, while the latent period of the elevation of Ca(i) fell from approximately 140 to 21 ms. The onset of the light-induced elevation of Ca(i) was always highly correlated with that of the receptor potential. The time for Ca(i) to exceed 2 microM was approximately equal to that for the receptor potential to exceed 8 mV (mean difference; 2.2 +/- 6.4 ms). Ca(i) was also measured during steps of light delivering approximately 10(5) effective photons/s to photoreceptors that had been bleached with hydroxylamine so as to reduce their quantum efficiency. Elevations of Ca(i) were detected at the earliest times of the electrical response to the steps of light, when a significant receptor potential had yet to develop. Successive responses exhibited stochastic variation in their latency of up to 20 ms, but the elevation of Ca(i) and the receptor potential still rose at approximately the same time, indicating a shared process generating the latent period. Light-induced elevations of Ca(i) resulted from Ca(2+) release from intracellular stores, being abolished by cyclopiazonic acid (CPA), an inhibitor of endoplasmic reticulum Ca(2+) pumps, but not by removal of extracellular Ca(2+) ions. CPA also greatly diminished and slowed the receptor potential elicited by dim flashes. The results demonstrate a rapid release of Ca(2+) ions that appears necessary for a highly amplified electrical response to dim flashes.