Cyclic AMP-dependent phosphorylation modifies the gating properties of L-type Ca2+ channels in bovine adrenal chromaffin cells.
We investigated the effects of cAMP-dependent phosphorylation on the voltage- and time-dependent gating properties of Ca2+ channel currents recorded from bovine adrenal chromaffin cells under whole-cell voltage clamp. Extracellular perfusion with the membrane-permeant activator of cAMP-dependent protein kinase, 8-bromo(8-Br)-cAMP (1 mM), caused a 49%, 29%, and 21% increase in Ca2+ current (ICa) amplitudes evoked by voltage steps to 0, +10, and +20 mV respectively (mean values from eight cells, p less than or equal to 0.05). Analysis of voltage-dependent steady-state activation (m infinity) curves revealed a 0.70 +/- 0.27 charge increase in the activation-gate valency (zm) following 8-Br-cAMP perfusion. Similar responses were observed when Ba2+ was the charge carrier, where zm was increased by 1.33 +/- 0.34 charges (n = 8). The membrane potential for half activation (V1/2) was also significantly shifted 6 mV more negative for IBa (mean, n = 8). The time course for IBa (and ICa) activation was well described by second-order m2 kinetics. The derived time constant for activation (tau m) was voltage-dependent, and the tau m/V relation shifted negatively after 8-Br-cAMP treatment. Ca2+ channel gating rates were derived from the tau m and m infinity 2 values according to a Hodgkin-Huxley type m2 activation process. The forward rate (alpha m) for channel activation was increased by 8-Br-cAMP at membrane potentials greater than or equal to 0 mV, and the backward rate (beta m) decreased at potentials less than or equal to + 10 mV. Time-dependent inactivation of ICa consisted of a slowly decaying component (tau h approximately 300 ms) and a "non-inactivating" steady-state component. The currents contributed by the two inactivation processes displayed different voltage dependences, the effects of 8-Br-cAMP being exclusively on the slowly inactivating L-type component.
Doupnik
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CA
Department of Physiology and Biophysics, University of Cincinnati College of Medicine, Ohio 45267-0576.
Pun
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D015124:8-Bromo Cyclic Adenosine Monophosphate / Q000494:pharmacology; D000311:Adrenal Glands / Q000166:cytology / Q000378:metabolism*; D000818:Animals; D015220:Calcium Channels / Q000378:metabolism*; D002417:Cattle; D002478:Cells, Cultured; D002838:Chromaffin System / Q000166:cytology / Q000378:metabolism*; D000242:Cyclic AMP / Q000502:physiology*; D004594:Electrophysiology; D006706:Homeostasis; D015640:Ion Channel Gating; D007700:Kinetics; D010766:Phosphorylation; D013997:Time Factors
D015220:Calcium Channels; D015124:8-Bromo Cyclic Adenosine Monophosphate; D000242:Cyclic AMP