Na+- and cGMP-induced Ca2+ fluxes in frog rod photoreceptors.

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Abstract
We have examined the Ca2+ content and pathways of Ca2+ transport in frog rod outer segments using the Ca2+-indicating dye arsenazo III. The experiments employed suspensions of outer segments of truncated, but physiologically functional, frog rods (OS-IS), intact isolated outer segments (intact OS), and leaky outer segments (leaky OS with a plasma membrane leaky to small solutes, but with sealed disk membranes). We observed the following. (a) Intact OS or OS-IS isolated and purified in Percoll-Ringer''s solution contained an average of 2.2 mM total Ca2+, while leaky OS contained 2.0 mM total Ca2+. This suggests that most of the Ca2+ in OS-IS is contained inside OS disks. (b) Phosphodiesterase inhibitors increased the Ca2+ content to .apprx. 4.2 mM in intact OS or OS-IS, whereas the Ca2+ content of leaky OS was not altered. (c) Na-Ca exchange was the dominant pathway for Ca2+ efflux in both intact and leaky OS/OS-IS. The rate of Na-Ca exchange in intact OS/OS-IS was half-maximal between 30 and 50 mM Na+; at 50 mM Na+, this amounted to 5.8 .times. 107 Ca2+/OS .cntdot. s or 0.05 mM total Ca2+/S. This is much larger than the Ca2+ component of the dark current. (d) Other alkali cations could not replace Na+ in Na-Ca exchange in either OS-IS or leaky OS. They inhibited the rate of Na-Ca exchange (K .gtoreq. Rb > Cs .gtoreq. Li > TMA) and, as the inhibition became greater, a delay developed in the onset of Na-Ca exchange. The inhibition of Na-Ca exchange by alkali cations correlates with the prolonged duration of the photoresponse induced by these cations (Hodgkin, A. L., P. A. McNaughton, and B. J. Nunn. 1985. Journal of Physiology. 358: 447-468). (e) In addition to Na-Ca exchange, disk membranes in leaky OS showed a second pathway of Ca2+ transport activated by cyclic GMP (cGMP). The cGMP-activated pathway required the presence of alkali cations and had a maximal rate of 9.7 .times. 106 Ca2+/OS .cntdot. s. cGMP caused the release of only 30% of the total Ca2+ from leaky OS. The rate of Na-Ca exchange in leaky OS amounted to 1.9 .times. 10-7 Ca2+/OS .cntdot. s. (f) Even though the ratio of plasma to disk membrane surface area in frog OS is .apprx. 1/100, the measured Ca2+ flux across the plasma membrane would be expected to change the cytoplasmic Ca2+ levels three times faster than Ca2+ fluxes across the disk membrane.

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