A voltage-clamp study of the light response in solitary rods of the tiger salamander

Abstract

1. Single, isolated, rod photoreceptors were obtained by enzymatic dissociation of the tiger salamander (Ambystoma tigrinum) retina. These solitary cells retained the morphological features of rods of the intact retina and could be maintained in culture for several days. Solitary cells were penetrated with one or two micropipettes and their electrophysiology was studied by the voltage-clamp technique. 2. Intracellular recording with two micropipettes demonstrated that the inner segment of a solitary rod was effectively isopotential with the outer segment. 3. The time course of the voltage response to a flash resembled that of responses observed in rods in the intact retina. At low light intensities the response reached a peak in approximately 0.7 sec and then slowly declined. At high light intensities the time to peak response decreased and an initial transient arose as the response, after reaching the peak, quickly decreased to a less polarized plateau. 4. The normal voltage response could be compared with the current observed during a voltage clamp. At low light intensities the time course of the current response resembled the time course of the voltage response. When light intensity was increased the time course of the current response differed from the voltage response in that the time to peak amplitude remained relatively constant and an initial transient did not occur. It was possible to predict the current response produced by any intensity of light by using (i) an empirical equation which reproduced the time course of a dim response and (ii) the Michaelis-Menten equation. 5. The time course of the voltage-clamp current produced by a flash was the same at different values of maintained voltage. 6. The maximum amplitude of the voltage-clamp current produced by a flash or step of light was a non-linear function of membrane potential. It was relatively constant within the physiological range, decreased as the membrane potential was moved toward 0 mV, reversed polarity between 0 and 10 mV, and rapidly increased in magnitude as membrane potential was made more positive. Although this current was voltage dependent, no time dependence was evident (recording resolution greater than or equal to 5 msec). 7. Voltage-clamp experiments demonstrated an inward current which slowly developed after a hyperpolarizing voltage step. The effect of this voltage and time dependent current was to reduce, after a delay, the polarization initiated by light.