Functional Expression of Human Rod and Cone Na/Ca–K Exchangers in Xenopus laevis Rods

Abstract

Abstract: : Purpose: Calcium extrusion from the outer segments of rods and cones via cell–specific Na/Ca–K exchangers is crucial for their ability to adapt to ambient light. In order to study how the rod and cone exchanger properties affect the physiology of photoreceptors, we produced transgenic Xenopus expressing human rod or cone Na/Ca–K exchanger in the rods. Methods: We made transgenic Xenopus frogs expressing human rod or cone Na/Ca–K exchanger under the control of the CMV promoter. GFP under the same promoter was co–introduced for initial screening of the animals. Using a suction electrode, membrane current was recorded from single rod photoreceptors from transgenic and control animals. Results: Rods expressing human cone Na/Ca–K exchanger showed reduced sensitivity, lower dark current and faster response kinetics in darkness compared to control rods expressing just the GFP marker. The decay time constant of the exchange current in bright light was three times shorter in transgenic rods than in control cells, indicating faster extrusion of calcium from the outer segment of transgenic rods. Equal background lights induced smaller changes in sensitivity, dark current, and response kinetics in transgenic rods than in control rods. Interestingly, in preliminary experiments, similar results were obtained from transgenic rods expressing the human rod Na/Ca–K exchanger. Conclusions: We have demonstrated for the first time the functional expression of human rod and cone Na/Ca–K exchangers in amphibian rods. The over–expression of either rod or cone exchanger accelerates the calcium extrusion, reduces the sensitivity, and accelerates the response kinetics of transgenic rods. Thus, the presence of transgenic Na/Ca–K exchanger in rods mimics the effects of background light by shifting their functional dynamic range.

Keywords: photoreceptors • electrophysiology: non–clinical • calcium