Prolongation of actions of Ca2+ early in photo transduction by 9-demethylretinal

During adaptation Ca2+ acts on a step early in phototransduction, Which is normally available for only a brief period after excitation. To investigate the identity of this step, we studied the effect of the light-induced decl ine in intracellular Ca2+ concentration on the response to a bright Hash in normal rods, and in rods bleached and regenerated with 11-cis 9-demethylre tinal, which forms a photopigment with a prolonged photoactivated lifetime. Changes in cytoplasmic Ca2+ were opposed by rapid superfusion of the outer segment with a 0Na+/0Ca(2+) solution designed to minimize Ca2+ fluxes acro ss the surface membrane. After regeneration of a bleached rod with 9-demeth lyretinal, the response in Ringer's to a 440-nm bright flash was prolonged in comparison with the unbleached control, and the response remained in sat uration for 10-15s. If the dynamic fall in Ca-i(2+) induced by the flash wa s delayed by stepping the outer segment to 0Na(+)/0Ca(2+) solution just bef ore the flash and returning it to Ringer's shortly before recovery, then th e response saturation was prolonged further, increasing linearly by 0.41 +/ -0.01 of the time spent in this solution. In contrast, even long exposures to 0Na+/0Ca(2+) solution of rods containing native photopigment evoked only a modest response prolongation on the return to Ringer's. Furthermore, if the rod was preexposed to steady subsaturating light, thereby reducing the cytoplasmic calcium concentration, then the prolongation of the bright Hash response evoked by 0Na+/0Ca(2+) solution was reduced in a graded manner wi th increasing background intensity. These results indicate that altering th e chromophore of rhodopsin prolongs the time Course of the Ca2+-dependent s tep early in the transduction cascade so that it dominates response recover y, and suggest that it is associated with photopigment quenching by phospho rylation.