Silver ions induce a rapid Ca2+ release from isolated intact bovine rod outer segments by a cooperative mechanism

Abstract

Micromolar concentrations of silver ion activate large Ca2+ fluxes across the plasma membrane of intact rod outer segments isolated from bovine retinas (intact ROS). The rate of Ag+-induced Ca2+ efflux from intact ROS depended on the Ag+ concentration in a sigmoidal manner suggesting a cooperative mechanism with a Hill coefficient between 2 and 3. At a concentration of 50 microM Ag+ the rate of Ca2+ efflux was 7 x 10(6) Ca2+/outer segment/sec; this represents a change in total intracellular Ca2+ by 0.7 mM/outer segment/sec. Addition of the nonselective ionophore gramicidin in the absence of external alkali cations greatly reduced the Ag+-induced Ca2+ efflux from intact ROS, apparently by enabling internal alkali cations to leak out. Adding back alkali cations to the external medium restored Ag+-induced Ca2+ efflux when gramicidin was present. In the presence of gramicidin, Ag+-induced Ca2+ efflux from intact ROS was blocked by 50 microM tetracaine or L-cis diltiazem, whereas without gramicidin both blockers were ineffective. Both L-cis diltiazem and tetracaine are blockers of one kinetic component of cGMP-induced Ca2+ flux across ROS disk membranes. The ion selectivity of the Ag+-induced pathway proved to be broad with little discrimination between the alkali cations Li+, Na+, K+, and Cs+ or between Ca2+ and Mg2+. The properties of the Ag+-induced pathway(s) suggest that it may reflect the cGMP-dependent conductance opened in the absence of cGMP by silver ions.

Cation selectivity of and cation binding to the cGMP-dependent channel in bovine rod outer segment membranes.

“…Cation fluxes across ROS membrane vesicles were measured with the optical probe neutral red as described before (Schnetkamp et al, 1981;Schnetkamp, 1985a, b;Schnetkamp and Szerencsei, 1989). The physicochemical basis of this method is illustrated schematically in Fig.…”

“…1; second, the transmembrane potential in the case that the charged form of neutral red can cross the bilayer membrane. The binding of neutral red to ROS membranes is not affected by changes in membrane potential (Schnetkamp and Szerencsei, 1989), leaving the electrostatic surface potential as the dominant parameter controlling changes in binding of neutral red to ROS membranes. Thus, the binding of the positively charged protonated form of neutral red is a function of the electrostatic surface potential at the membrane/water interface, which in turn is a simple function of the concentration and valency of the cations in the aqueous solution (as illustrated in Fig.…”