External and internal actions in the response of salamander retinal rods to altered external calcium concentration.

1. The membrane current was recorded from retinal rods isolated from Ambystoma tigrinum using the suction pipette and whole-cell patch pipette techniques, while the concentration of calcium bathing the outer segment was rapidly reduced. 2. The increase in outer segment current induced by lowered external calcium in darkness could be resolved into two components, one as rapid as the time course of the solution change (as judged by the junction current) and the other somewhat slower. 3. Introduction of the calcium buffer BAPTA (1,2-bis(o-aminophenoxy)ethane-N ,N ,N' ,N'-tetraacetic acid) into the cell from a patch pipette led to a progressive slowing of the second component of current increase. 4. When several minutes had elapsed following rupture of the patch, to allow a substantial amount of BAPTA into the cell (ca. 10 mM in the patch pipette), the second component was slowed by a factor of about 20-fold, while the first component continued to have the same rapid time course as the solution change. 5. The rapid component is attributed to a direct effect of external calcium, Ca2+o, and the delayed component to an indirect effect mediated by a reduction in internal calcium, Ca2+i. 6. These results confirm that, in previous experiments in which BAPTA was introduced into photoreceptors, the internal calcium concentration was very significantly buffered. 7. When Ca2+o drops from 1 mM to less than 10(-8) M, the rapid external component corresponds to an increase in circulating current of 3- to 4-fold, and the internal component corresponds to an increase of at least 4- to 5-fold. However, the total current at late times is limited by electrical factors, so that the size of the internal effect is bound to be considerably greater.