The thermal contribution to photoactivation in A2 visual pigments studied by temperature effects on spectral properties

Effects of temperature on the spectral properties of visual pigments were measured in the physiological range (5-28degreesC) in photoreceptor cells of bullfrog (Rana catesbeiana) and crucian carp (Carassius carassius). Absorbance spectra recorded by microspectrophotometry (MSP) in single cells and sensitivity spectra recorded by electroretinography (ERG) across the isolated retina were combined to yield accurate composite spectra from ca. 400 nm to 800 nm. The four photoreceptor types selected for study allowed three comparisons illuminating the properties of pigments using the dehydroretinal (A2) chromophore: (1) the two members of an A1/A2 pigment pair with the same opsin (porphyropsin vs. rhodopsin in bullfrog "red" rods); (2) two A2 pigments with similar spectra (porphyropsin rods of bullfrog and crucian carp); and (3) two A2 pigments with different spectra (rods vs. long-wavelength-sensitive (L-) cones of crucian carp). Qualitatively, the temperature effects on A2 pigments were similar to those described previously for the A1 pigment of toad "red" rods. Warming caused an increase in relative sensitivities at Very long wavelengths but additionally a small shift of lambda(max) toward shorter wavelengths. The former effect was used for estimating the minimum energy required for photoactivation (E-a) of the pigment. Bullfrog rod opsin with A2 chromophore had E-a = 44.2 +/- 0.9 kcal/mol, significantly lower (one-tailed P < 0.05) than the value E-a = 46.5 +/- 0.8 kcal/mol for the same opsin coupled to A1. The A2 rod pigment of crucian carp had E-a = 42.3 +/- 0.6 kcal/mol. which is significantly higher (one-tailed P < 0.01) than that of the L-cones in the same retina (E-a = 38.3 +/- 0.4 kcal/mol), whereas the difference compared with the bullfrog A2 rod pigment is not statistically significant (two-tailed P = 0.13). No strict connection between lambda(max) and E-a appears to exist among A2 pigments any more than among A1 pigments. Still, the A1 --> A2 chromophore substitution in bullfrog opsin causes three changes correlated as originally hypothesized by Barlow (1957): a red-shift of lambda(max), a decrease in E-a, and an increase in thermal noise.