Using suction electrodes, photocurrent responses to 100-ms saturating flashes were recorded from isolated retinal rods of the larval-stage tiger salamander (Ambystoma tigrinum). The delay period (7" c ) that preceded recovery of the dark current by a criterion amount (3 pA) was analyzed in relation to the flash intensity (If), and to the corresponding fractional bleach (R* 0 /R lol ) of the visual pigment; Rl/R lol was compared with R*/R lol , the fractional bleach at which the peak level of activated transducin approaches saturation. Over an approximately 8 In unit range of If that included the predicted value of R*/R lol , T c increased linearly with In If. Within the linear range, the slope of the function yielded an apparent exponential time constant (T C ) of 1.7 ± 0.2 s (mean ± S.D.). Background light reduced the value of T c measured at a given flash intensity but preserved a range over which T c increased linearly with In If-, the linear-range slope was similar to that measured in the absence of background light. The intensity dependence of T c resembles that of a delay (T d ) seen in light-scattering experiments on bovine retinas, which describes the period of essentially complete activation of transducin following a bright flash; the slope of the function relating T d and In flash intensity is thought to reflect the lifetime of photoactivated visual pigment (/?*) (Pepperberg et al., 1988; Kahlert et al., 1990). The present data suggest that the electrophysiological delay has a similar basis in the deactivation kinetics of / ? ' , and that T C represents T R >, the lifetime of R* in the phototransduction process. The results furthermore suggest a preservation of the "dark-adapted" value of T R * within the investigated range of background intensity.

Photoactivation of rhodopsin initiates both excitation and adaptation in vertebrate rod photoreceptors. Bleaching of rhodopsin to free opsin and all-trans-retinal in isolated rods produces a stable desensitization (bleaching adaptation) that is much larger than expected from pigment depletion alone. In our experiments, a 93% bleach produced a 500-fold increase in the light intensity required for saturation of the light response. This component of adaptation was 32-fold larger than the 16-fold increase expected from pigment depletion alone.11-cis-Retinal, when delivered to isolated rods from liposomes, combines with free opsin to form a bleachable photopigment that fully restores sensitivity. 11-cis-Locked analogues of retinal combine with opsin to form unbleachable pigments in isolated bleached rods from the tiger salamander. They restore sensitivity to a substantial (16-to 25-fold) but incomplete extent. The analogues apparently relieve a stable component of adaptation when they interact with opsin. Because these analogues do not detectably excite rods, the structural requirements of both retinal and opsin for the relief of adaptation are different from those of excitation. The biochemical basis of light adaptation resulting from pigment bleaching and the minimum structural requirements of retinal for its relief remain to be determined.

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