Effects of intracellular Ca2+ chelation on the light response in Drosophila photoreceptors

In order to test the hypothesis that excitation in Drosophila photoreceptors is mediated by Ca2+ released from internal stores, the Ca2+ buffers EGTA, BAPTA and di-bromo-BAPTA (DBB) were introduced into dissociated photoreceptors via whole-cell recording pipettes. All buffers were preloaded with Ca2+ to provide the same free Ca2+ concentration (250 nM). EGTA (up to 18 mM free buffer) had only weak effects upon voltage-clamped flash responses in normal Ringer's solution (1.5 mM Ca (2 0+)), and no effect in Ca2+ -free solution. The maximum BAPTA concentration tested (14.4 mM free BAPTA) reduced the initial rate of rise by ca. 5000-fold in normal Ringer's solution; by ca. 500-fold in Ca2+ free solution; and only ca. 60-fold in the absence of Mg2+, which preferentially blocks one component of the light-sensitive current. Although BAPTA delayed the time-to-peak in normal Ringer's solution, responses in Ca2+ free Ringer's solution were accelerated. These results support the role of Ca2+ influx in regulating sensitivity and response kinetics; however, in view of the high concentrations required to attenuate responses in Ca2+ free Ringer's solution, the role of Ca2+ release in excitation remains unclear. DBB was ca. 2-3 fold more potent than BAPTA, and at concentrations > 5 mM had a qualitatively different action, greatly delaying the time-to-peak. This suggests DBB may have distinct pharmacological actions or access to compartments inaccessible to BAPTA. The only current activated by introducing 5-500 microM Ca2+ (buffered with nitrilo-triacetic acid) was electrogenic Na+/Ca2+ exchange. When this was blocked by removing Nao0+, a novel cationic conductance was activated. However, its properties did not resemble those the light-activated conductance, and thus do not support the hypothesis that Ca2+ is sufficient for excitation.