Fraction of the Dark Current Carried by Ca²⁺ through Cgmp-Gated Ion Channels of Intact Rod and Cone Photoreceptors

The selectivity for Ca²⁺ over Na⁺, PCa/PNa, is higher in cGMP-gated (CNG) ion channels of retinal cone photoreceptors than in those of rods. To ascertain the physiological significance of this fact, we determined the fraction of the cyclic nucleotide–gated current specifically carried by Ca²⁺ in intact rods and cones. We activated CNG channels by suddenly (<5 ms) increasing free 8Br-cGMP in the cytoplasm of rods or cones loaded with a caged ester of the cyclic nucleotide. Simultaneous with the uncaging flash, we measured the cyclic nucleotide–dependent changes in membrane current and fluorescence of the Ca²⁺-binding dye, Fura-2, also loaded into the cells. The ratio of changes in fura-2 fluorescence and the integral of the membrane current, under a restricted set of experimental conditions, is a direct measure of the fractional Ca²⁺ flux. Under normal physiological salt concentrations, the fractional Ca²⁺ flux is higher in CNG channels of cones than in those of rods, but it differs little among cones (or rods) of different species. Under normal physiological conditions and for membrane currents ≤200 pA, the Ca²⁺ fractional flux in single cones of striped bass was 33 ± 2%, and 34 ± 6% in catfish cones. Under comparable conditions, the Ca²⁺ fractional flux in rod outer segments of tiger salamander was 21 ± 1%, and 14 ± 1% in catfish rods. Fractional Ca²⁺ flux increases as extracellular Ca²⁺ rises, with a dependence well described by the Michaelis-Menten equation. KCa, the concentration at which Ca²⁺ fractional flux is 50% was 1.98 mM in bass cones and 4.96 mM in tiger salamander rods. Because Ca²⁺ fractional flux is higher in cones than in rods, light flashes that generate equal photocurrents will cause a larger change in cytoplasmic Ca²⁺ in cones than in rods.