Effects of injections of calcium and EGTA into the outer segments of retinal rods of <i>Bufo marinus</i>

1. Intracellular recordings were made from the outer segments of rods in the isolated, superfused retina of Bufo marinus. Cells were impaled under observation with a compound microscope fitted with an infra-red image converter. Changes of membrane voltage and some concomitant changes of input resistance were measured in response to light, membrane polarization and iontophoretic injections.&lt;p&gt;&lt;/p&gt; 2. By means of a double barrel micropipette, charge was passed into a rod from a micropipette barrel that contained Ca2+ while no net current crossed the plasma membrane. In about half the cells, immediately after the injection, a hyperpolarization was observed that decayed with a time course similar to the decay of the receptor potential.&lt;p&gt;&lt;/p&gt; 3. Membrane hyperpolarization also occurred after a depolarizing current stopped flowing into a rod through a single barrel pipette that contained only K-acetate. This hyperpolarization was accompanied by an increase of membrane conductance. The reversal potential for the conductance-increase was between the voltage in the dark and the voltage in the absence of [Na+]out. A larger hyperpolarization became evident after an equal depolarizing current stopped flowing into a rod through a pipette that also contained Ca2+; this larger after-hyperpolarization was due to both the cessation of depolarizing current and the injection of Ca2+.&lt;p&gt;&lt;/p&gt; 4. A depolarization of 10-20 mV that lasted 2-60 sec became evident after hyperpolarizing current stopped flowing into a rod through a single-barrel pipette filled with K-EGTA. During the after-depolarization, the responses to small, dim spots of light were attenuated. No depolarization was observed after passing hyperpolarizing currents into rods through pipettes that contained either acetate-, SO2-4 or MOPS-.&lt;p&gt;&lt;/p&gt; 5. These results show that sequestration of [Ca2+]in depolarizes the plasma membrane and that an increase in [Ca2+]in hyperpolarizes the membrane mimicking the later part of the receptor potential. These findings support the hypothesis of Yoshikami &#38; Hagins (1971) that Ca2+ is an intracellular messenger for excitation in vertebrate rods