Speed of Ca2+ Channel Modulation by Neurotransmitters in Rat Sympathetic
Neurons.
Jiuying Zhou, Mark S. Shapiro and Bertil Hille.
Dept. of Physiology and Biophysics, Univ. of Washington, Box 357290,
Seattle, WA 98195-7290.
APStracts 4:0005N, 1997.
ABSTRACT
We have measured the onset and recovery speed of inhibition of N-type Ca2+
channels in adult rat superior cervical ganglion neurons by somatostatin (SS),
norepinephrine (NE) and oxotremorine-M (Oxo-M, a muscarinic agonist), using
the whole-cell configuration of the patch-clamp method with 5 mM external
Ca2+. With a local perfusion pipette system that changed the solution
surrounding the cell within 50 ms, we applied agonists at various times before
a brief depolarization from -80 mV that elicited ICa. At concentrations that
produced maximal inhibition, the onset time constants for membrane-delimited
inhibition by SS (0.5 mM), NE (10 mM) and oxo-M (20 mM) were 2.1 s, 0.7 s and
1.0 s, respectively. The time constants for NE inhibition depended only weakly
on the concentration, ranging from 1.2 s to 0.4 s in the concentration range
from 0.5 to 100 mM. Inhibition by oxo-M (20 mM) through a different G protein
pathway that uses a diffusible cytoplasmic messenger had a time constant near
9 s. The recovery rate constant from membrane-delimited inhibition was between
0.09-0.18 s-1, significantly higher than the intrinsic GTPase rate of purified
G protein Go, suggesting that Ca2+ channels or other proteins in the plasma
membrane act as GTPase activating proteins. We also measured the rate of
channel reinhibition after relief by strong depolarizing prepulses, which
should reflect the kinetics of final steps in the inhibition process. In the
presence of different concentrations of NE, reinhibition was 4 to 7 times
faster than the onset of inhibition, indicating that the slowest step of
inhibition must precede the binding of G protein to the channel. We propose a
kinetic model for the membrane-delimited NE inhibition of Ca2+ channels. It
postulates two populations of receptors with different affinities for NE, a
single population of G proteins and a single population of Ca2+ channels. This
model closely simulated the time courses of onset and recovery of inhibition
and reinhibition, as well as the dose-response curve for inhibition of Ca2+
channels by NE.
Received 19 December 1996; accepted in final form 19 December 1996.
APS Manuscript Number J782-6.
Article publication pending J. Neurophysiol.
ISSN 1080-4757 Copyright 1997 The American Physiological Society.
Published in APStracts on 21 January 1997