Single-channel pharmacology of mibefradil in human native T-type and recombinant Ca(v)3.2 calcium channels.
To study the molecular pharmacology of low-voltage-activated calcium channels in biophysical detail, human medullary thyroid carcinoma (hMTC) cells were investigated using the single-channel technique. These cells had been reported to express T-type whole-cell currents and a Ca(v)3.2 (or alpha 1H) channel subunit. We observed two types of single-channel activity that were easily distinguished based on single-channel conductance, voltage dependence of activation, time course of inactivation, rapid gating kinetics, and the response to the calcium agonist (S)-Bay K 8644. Type II channels had biophysical properties (activation, inactivation, conductance) typical for high-voltage-activated calcium channels. They were markedly stimulated by 1 microM (S)-Bay K 8644, allowing to identify them as L-type channels. The channel termed type I is a low-voltage-activated, small-conductance (7.2 pS) channel that inactivates rapidly and is not modulated by (S)-Bay K 8644. Type I channels are therefore classified as T-type channels. They were strongly inhibited by 10 microM mibefradil. Mibefradil block was caused by changes in two gating parameters: a pronounced reduction in fraction of active sweeps and a slight shortening of the open-state duration. Single recombinant low-voltage-activated T-type calcium channels were studied in comparison, using human embryonic kidney 293 cells overexpressing the pore-forming Ca(v)3.2 subunit. Along all criteria examined (mechanisms of block, extent of block), recombinant Ca(v)3.2 interact with mibefradil in the same way as their native counterparts expressed in hMTC cells. In conclusion, the pharmacologic phenotype of these native human T-type channels--as probed by mibefradil--is similar to recombinant human Ca(v)3.2.
Michels
Guido
G
Department of Pharmacology, University of Cologne, Cologne, Germany.
Matthes
Jan
J
Handrock
Renate
R
Kuchinke
Ute
U
Groner
Ferdi
F
Cribbs
Leanne L
LL
Pereverzev
Alexey
A
Schneider
Toni
T
Perez-Reyes
Edward
E
Herzig
Stefan
S
D002121:Calcium Channel Blockers / Q000494:pharmacology*; D020746:Calcium Channels, L-Type / Q000187:drug effects / Q000378:metabolism; D020747:Calcium Channels, T-Type / Q000187:drug effects / Q000235:genetics / Q000378:metabolism*; D002478:Cells, Cultured; D004594:Electrophysiology; D006801:Humans; D020748:Mibefradil / Q000494:pharmacology*; D011994:Recombinant Proteins / Q000187:drug effects / Q000378:metabolism
D002121:Calcium Channel Blockers; D020746:Calcium Channels, L-Type; D020747:Calcium Channels, T-Type; D011994:Recombinant Proteins; D020748:Mibefradil