Voltage-clamp analysis of membrane currents and excitation-contraction coupling in a crustacean muscle.
A single fibre preparation from the extensor muscle of a marine isopod crustacean is described which allows the analysis of membrane currents and simultaneously recorded contractions under two-electrode voltage-clamp conditions. We show that there are three main depolarisation-gated currents, two are outward and carried by K+, the third is an inward Ca2+ current, I(Ca). Normally, the K+ currents which can be isolated by using K+ channel blockers, mask I(Ca). I(Ca) activates at potentials more positive than -40 mV, is maximal around 0 mV, and shows strong inactivation at higher depolarisation. Inactivation depends on current rather than voltage. Ba2+, Sr2+ and Mg2+ can substitute for Ca2+. Ba2+ currents are about 80% larger than Ca2+ currents and inactivate little. The properties of I(Ca) characterise it as a high threshold L-type current. The outward current consists primarily of a fast, transient A current, I(K(A)) and a maintained, delayed rectifier current, I(K(V)). In some fibres, a small Ca2+-dependent K+ current is also present. I(K(A)) activates fast at depolarisation above -45 mV, shows pronounced inactivation and is almost completely inactivated at holding potentials more positive than -40 mV. I(K(A)) is half-maximally blocked by 70 microM 4-aminopyridine (4-AP), and 70 mM tetraethylammonium (TEA). I(K(V)) activates more slowly, at about -30 mV, and shows no inactivation. It is half-maximally blocked by 2 mM TEA but rather insensitive to 4-AP. Physiologically, the two K+ currents prevent all-or-nothing action potentials and determine the graded amplitude of active electrical responses and associated contractions. Tension development depends on and is correlated with depolarisation-induced Ca2+ influx mediated by I(Ca). The voltage dependence of peak tension corresponds directly to the voltage dependence of the integrated I(Ca). The threshold potential for contraction is at about -38 mV. Peak tension increases with increasing voltage steps, reaches maximum at around 0 mV, and declines with further depolarisation.
Weiss
T
T
Department of Biology, University of Konstanz, Germany.
Erxleben
C
C
Rathmayer
W
W
D000818:Animals; D002121:Calcium Channel Blockers / Q000494:pharmacology; D015220:Calcium Channels / Q000502:physiology; D002413:Cations, Divalent / Q000494:pharmacology; D002414:Cations, Monovalent / Q000494:pharmacology; D003035:Cobalt / Q000494:pharmacology; D003445:Crustacea / Q000502:physiology*; D004553:Electric Conductivity; D008297:Male; D008564:Membrane Potentials / Q000187:drug effects / Q000502:physiology; D009119:Muscle Contraction / Q000187:drug effects / Q000502:physiology*; D018482:Muscle, Skeletal / Q000187:drug effects / Q000502:physiology*; D018408:Patch-Clamp Techniques / Q000379:methods / Q000706:statistics & numerical data; D011188:Potassium / Q000502:physiology
D002121:Calcium Channel Blockers; D015220:Calcium Channels; D002413:Cations, Divalent; D002414:Cations, Monovalent; D003035:Cobalt; D011188:Potassium