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Original Article

^a Department of Physiology, School of Medicine, University of California at San Francisco, San Francisco, California 94143
Dept. of Physiology, School of Medicine, Box 0444, University of California at San Francisco, San Francisco, CA 94143.415-476-4929

juan{at}itsa.ucsf.edu

We determined the Ca²⁺ dependence and time course of the modulation of ligand sensitivity in cGMP-gated currents of intact cone photoreceptors. In electro-permeabilized single cones isolated from striped bass, we measured outer segment current amplitude as a function of cGMP or 8Br-cGMP concentrations in the presence of various Ca²⁺ levels. The dependence of current amplitude on nucleotide concentration is well described by the Hill function with values of K_1/2, the ligand concentration that half-saturates current, that, in turn, depend on Ca²⁺. K_1/2 increases as Ca²⁺ rises, and this dependence is well described by a modified Michaelis-Menten function, indicating that modulation arises from the interaction of Ca²⁺ with a single site without apparent cooperativity. ^CaK_m, the Michaelis-Menten constant for Ca²⁺ concentration is 857 ± 68 nM for cGMP and 863 ± 51 for 8Br-cGMP. In single cones under whole-cell voltage clamp, we simultaneously measured changes in membrane current and outer segment free Ca²⁺ caused by sudden Ca²⁺ sequestration attained by uncaging diazo-2. In the presence of constant 8Br-cGMP, 15 µM, Ca²⁺ concentration decrease was complete within 50 ms and membrane conductance was enhanced 2.33 ± 0.95-fold with a mean time to peak of 1.25 ± 0.23 s. We developed a model that assumes channel modulation is a pseudo–first-order process kinetically limited by free Ca²⁺. Based on the experimentally measured changes in Ca²⁺ concentration, model simulations match experimental data well by assigning the pseudo-first-order time constant a mean value of 0.40 ± 0.14 s. Thus, Ca²⁺-dependent ligand modulation occurs over the concentration range of the normal, dark-adapted cone. Its time course suggests that its functional effects are important in the recovery of the cone photoresponse to a flash of light and during the response to steps of light, when cones adapt.

Key Words: retina • ligand-gated ion channel • phototransduction • adaptation • teleost fish

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This article has been cited by other articles:

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