A Na+/Ca2+ exchange mechanism in apical membrane vesicles of the retinal pigment epithelium.

The retinal pigment epithelium (RPE) lying between the neural retina and the choroid, performs as a transport organ for solutes and water between the choriocapillaries and the subretinal space. It also has the function to maintain the microenvironment of photoreceptors including the regulation of calcium ions during light or dark adaptation. In order to further elucidate the transport functions of the RPE, apical membranes were isolated from RPE by differential precipitation with divalent ions. In this work bovine tissues were used as well as elasmobranch tissues. For the latter, we have already purified and characterized membrane vesicles in a previous paper. Na(+)-K(+)-ATPase, alkaline phosphatase, and 5'-nucleotidase, which are marker enzymes of the apical membrane, were highly enriched in the final membrane fraction. The majority of the fraction consists of right side out vesicles. The fluorescent indicator for sodium, SBFI, or the calcium specific indicator, Fura-2, were pre-loaded into the apical membrane vesicles of RPE of either dogfish eyes or bovine eyes. When an outwardly-directed Ca2+ gradient was formed across the vesicular membranes, the Ca2+ influx was also enhanced by 136% for dogfish RPE and 167% for bovine RPE. This Na+ gradient dependent Ca2+ influx was blocked by bepridil, an antiarrhythmic agent which is a Na+/Ca2+ exchanger inhibitor. These results indicate that a Na+/Ca2+ exchanger is present in the apical membrane of bovine and dogfish RPE.