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Journal of Neurophysiology, Vol 75, Issue 1 354-366, Copyright © 1996 by APS
ARTICLES |
J. P. Younger, S. T. McCarthy and W. G. Owen
Department of Molecular and Cell Biology, University of California, Berkeley 94720-3200.
1. Using Fura-2, we measured cytosolic free calcium concentrations in rod outer segments of the bullfrog (Rana catesbeiana) under a wide range of steady-state, adapting light intensities. We also measured rod circulating currents under the same adapting conditions. 2. Both the steady-state cytosolic free calcium concentration and the steady-state circulating current were halved by background lights that isomerized approximately 100 rhodopsin molecules per rod per second. A just-measurable reduction in calcium was evoked by a background 1/10 as bright. The steady-state calcium concentration and the steady-state circulating current were proportional to each other over the full range of intensities tested, the brightest of which suppresses the circulating current by 75% and reduces flash sensitivity by about two orders of magnitude relative to the rods' sensitivity in darkness. 3. Additional experiments, in which the Na+:Ca2+,K+ exchanger in the rod outer segment was inactivated while maintaining intracellular calcium at physiological levels, demonstrated that steady-state calcium concentrations in the outer segment are set only by the influx of calcium through the light-sensitive channels and its efflux via the Na+:Ca2+,K+ exchanger. We found no measurable light-induced release of calcium from internal stores and no evidence of any other calcium flux between the cytosol and intracellular compartments. 4. Taken together, these findings lead us to conclude that over the normal operating range of the rod, the selectivity of the cGMP-gated channels for calcium does not change as a result of background illumination. 5. Our data also suggest that a major role of calcium-dependent cGMP synthesis by guanylyl cyclase is to stabilize both the circulating current and the cytosolic free calcium concentration in darkness. This would minimize the dark noise of the rod and thereby increase the reliability with which dim stimuli can be detected in the dark-adapted state.
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