Blue Light's Effects on Rhodopsin: Photoreversal of Bleaching in Living Rat Eyes

To determine whether blue light induces photoreversal of rhodopsin bleaching in vivo. Eyes of anesthetized albino rats were exposed to either green (550 nm) or deep blue (403 nm) light, and the time course of rhodopsin bleaching was determined. Rhodopsin was isolated from whole retinas by detergent extraction and measured photometrically. To inhibit photoreversal of bleaching, rats were perfused with 70 mM hydroxylamine (NH(2)OH), a known inhibitor of photoreversal. To determine whether blue-absorbing, photoreversible photoproducts were formed, rhodopsin was bleached to near completion with green light and then exposed to blue light. Finally, experimental results were simulated on a computer by means of a simple, three-component model involving a long-lived photoreversible photoproduct. Photoreversal of bleaching in blue light occurs in vivo as evidenced by the following: In the absence of NH(2)OH, bleaching of rhodopsin by blue light was slow and complex. In the presence of NH(2)OH, however, blue light bleached rhodopsin very fast with a simple, pseudo-first-order kinetic. A long-lived bleaching intermediate produced by green light exposure was photoreversed to rhodopsin by exposure to blue light. The three-component computer model, invoking a blue-absorbing, photoreversible, long-lived intermediate accurately described the data. Because of the instantaneous, nonmetabolic regeneration of rhodopsin by the process of photoreversal of bleaching, blue light exposure permits the absorption of large numbers of photons by rhodopsin and by a photoreversible intermediate of bleaching in vivo. These data may have an important impact on resolving mechanisms of blue light-mediated damage to the retina.