Light Dependence of Calcium and Membrane Potential Measured in Blowfly Photoreceptors In Vivo

Light adaptation in insect photoreceptors is caused by an increase in the cytosolic Ca²⁺ concentration. To better understand this process, we measured the cytosolic Ca²⁺ concentration in vivo as a function of adapting light intensity in the white-eyed blowfly mutant chalky. We developed a technique to measure the cytosolic Ca²⁺ concentration under conditions as natural as possible. The calcium indicator dyes Oregon Green 1, 2, or 5N (Molecular Probes, Inc., Eugene, OR) were iontophoretically injected via an intracellular electrode into a photoreceptor cell in the intact eye; the same electrode was also used to measure the membrane potential. The blue-induced green fluorescence of these dyes could be monitored by making use of the optics of the facet lens and the rhabdomere waveguide. The use of the different Ca²⁺-sensitive dyes that possess different affinities for Ca²⁺ allowed the quantitative determination of the cytosolic Ca²⁺ concentration in the steady state. Determining the cytosolic Ca²⁺ concentration as a function of the adapting light intensity shows that the Ca²⁺ concentration is regulated in a graded fashion over the whole dynamic range where a photoreceptor cell can respond to light. When a photoreceptor is adapted to bright light, the cytosolic Ca²⁺ concentration reaches stable values higher than 10 μM. The data are consistent with the hypothesis that the logarithm of the increase in cytosolic Ca²⁺ concentration is linear with the logarithm of the light intensity. From the estimated values of the cytosolic Ca²⁺ concentration, we conclude that the Ca²⁺-buffering capacity is limited. The percentage of the Ca²⁺ influx that is buffered gradually decreases with increasing Ca²⁺ concentrations; at cytosolic Ca²⁺ concentration levels above 10 μM, buffering becomes minimal.