G-PROTEIN-MEDIATED ACTIVATION OF A NONSPECIFIC CATION CURRENT IN CULTURED RAT RETINAL-PIGMENT EPITHELIAL-CELLS

We used whole-cell patch-clamp recording techniques to investigate G p rotein-activated currents in cultured rat retinal pigment epithelial ( RPE) cells. Using 140 mM KCl intracellular and 130 mM NaCl extracellul ar solutions, rat RPE cells possessed both inward and outward K+ curre nts. Upon addition of the nonhydrolyzable guanine triphosphate analogu e, guanosine-5'-O-(3-thiophosphate) (GTP gamma S, 0.1 mM), to the reco rding electrode, a nonspecific cation (NSC) current was elicited. The NSC current had a mean reversal potential of +5.7 mV in 130 mM extrace llular NaCl with Cs+-aspartate in the pipette, and was not affected by alterations in the extracellular Ca2+ or Cl- concentration. The GTP g amma S-activated current was found to be permeable to several monovale nt cations (K+, Na+, choline, TRIS, and NMDG). Addition of fluoroalumi nate, an activator of large molecular weight heterotrimeric GTP-bindin g proteins (G proteins), to the intracellular recording solution activ ated the NSC current. The G protein involved was pertussis toxin (PTX) -sensitive, since GTP gamma S failed to activate the NSC current in ce lls pretreated with PTX. Further investigation of second messenger mol ecules suggested that activation of the NSC current was not affected b y alterations in intracellular Ca2+ or ATP. From these results, we con clude that a G protein-regulated NSC current is present in rat RPE cel ls. Activation of the NSC current may sufficiently depolarize RPE cell s to activate outward K+ currents. This would provide a mechanism by w hich these cells could rid themselves of accumulated K+.