Rapid coupling of calcium release to depolarization in Limulus polyphemus ventral photoreceptors as revealed by microphotolysis and confocal microscopy

Abstract

Microphotolysis and confocal microscopy were used to investigate the timing of calcium release and of the electrical response in Limulus polyphemus ventral photoreceptors. The fluorescent dyes Fluo-3 and Calcium Green-5N were used to monitor local Ca2+ elevations. Photolysis of caged inositol trisphosphate (InsP3) close to the plasma membrane of the light- sensitive rhabdomeral (R-) lobe resulted in Ca2+ elevation within 10-20 msec, 20-45 msec before the physiological response to light normally would be detected. Inward ionic current flow and depolarization followed InsP3- induced calcium release within 2.5 ± 3.3 msec. Voltage-clamping the cells and removal of extracellular Ca2+ did not affect the timing of the Ca2+ elevation that followed the photolysis of caged InsP3 or its relationship to the electrical response. In contrast to the physiological response to light, which only released calcium within the R-lobe, photolysis of InsP3 elevated Ca(i) in both lobes, although with much greater effect in the R-lobe, as compared with the bulk of the A-lobe, suggesting the presence of InsP3- sensitive calcium stores in both lobes. Photolysis of caged calcium [o- nitrophenyl EGTA (NPE)] at the edge of the R-lobe activated an inward ionic current within 1.8 ± 0.7 msec. This NPE-induced current reversed at a membrane potential of 10 ± 6 mV in the range typical of that of the light- activated current under physiological conditions. Calcium release, therefore, activates an inward current rapidly enough to contribute to the electrical response to light.