ISSN:
1432-2013
Keywords:
Patch clamp
;
Ca2+ channel
;
Cardiac ventricular muscle
Source:
Springer Online Journal Archives 1860-2000
Topics:
Medicine
Notes:
Abstract Elementary Ca2+ and Ba2+ currents were recorded from cell-attached membrane patches of ventricular myocytes from adult guinea pig hearts using the improved patch-clamp technique (Hamill et al. 1981). High concentrations of Ba2+ or Ca2+ (50 or 90 mM) were used in the pipettes to increase the signal-to-noise ratio. All data were derived from elementary current analyses in patches containing only one channel. 1) In response to voltage steps, channel openings occurred singly or in bursts of closely spaced unitary current pulses separated by wider shut intervals. During depolarizations of small amplitude from the resting potential, channel openings occurred almost randomly, whereas during larger depolarizations the events were grouped preferentially at the beginning. 2) Channel openings became more probable with increased depolarization; simultaneously, unitary current amplitudes declined in an ohmic manner. Elementary current amplitudes were slightly larger, when 50 mM Ba2+ replaced 50 mM Ca2+ in the pipettes (slope conductances 9 and 10 pS, respectively), but more than doubled, when Ba2+ was increased to 90 mM (slope conductance 18 pS). Clear outward currents through Ca2+ channels were not observed under these conditions. 3) Peak amplitudes of reconstructed mean currents doubled when 50 mM Ba2+ replaced 50 mM Ca2+ and were larger still when 90 mM Ba2+ was used in the pipettes. The current-volrage relations of the reconstructed mean currents showed a positive shift along the voltage axis as Ba2+ was increased or substituted equimolarly by Ca2+. Correspondingly, the open state probability-voltage relations (activation curves) showed a parallel shift as Ba2+ was increased, which was less pronounced when Ba2+ was replaced equimolarly by Ca2+. 4) Determination of Ca2+ channel inactivation using 90 mM Ba2+ in the pipettes indicated an overlap with channel activation in a limited voltage range, resulting in a steadystate “window” current. Inactivation can occur without divalent cation influx. 5) Formation of an inside-out patch resulted in a fast rundown of elementary Ca2+ channel currents. 6) Channel openings were often grouped in bursts. The lifetimes of the open state, the bursts, and the closed states were estimated for Ba2+ and Ca2+ as permeating ions. At least two exponentials were needed to fit the histogram of the lifetimes of all closed states. The lifetimes of the individual openings and bursts were mono-exponentially distributed. The kinetics of the Ca+ channel depended on the voltage and the permeating ion. During +30 mV depolarizations, no significant effect of the permeating ion on channel gating could be detected. The significant increase in burst length (t b) during +50 mV depolarizations, however, seemed to be only due to an increase in the lifetime of the open state (t o) for Ba2+, whereas for Ca2+,t o was only moderately prolonged but simultaneously, the number of openings per burst increased. 7) A three-state sequential scheme is peoposed to model the activation pathway of Ca2+ channels. The latency-to-first-event histogram is also consistent with a process in which multiple closed states precede the open state.
Type of Medium:
Electronic Resource
URL:
http://dx.doi.org/10.1007/BF00657238
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