Pentobarbital inhibition of human recombinant alpha1A P/Q-type voltage-gated calcium channels involves slow, open channel block.
OBJECTIVE
Pre-synaptic neurotransmitter release is largely dependent on Ca(2+) entry through P/Q-type (Ca(V)2.1) voltage-gated Ca(2+) channels (PQCCs) at most mammalian, central, fast synapses. Barbiturates are clinical depressants and inhibit pre-synaptic Ca(2+) entry. PQCC barbiturate pharmacology is generally unclear, specifically in man. The pharmacology of the barbiturate pentobarbital (PB) in human recombinant alpha(1A) PQCCs has been characterized.


METHODS
PB effects on macroscopic Ca(2+)(I(Ca)) and Ba(2+)(I(Ba)) currents were studied using whole-cell patch clamp recording in HEK-293 cells heterologously expressing (alpha(1A))(human)(beta(2a)alpha(2)delta-1)(rabbit) PQCCs.


RESULTS
PB reversibly depressed peak current (I(peak)) and enhanced apparent inactivation (fractional current at 800 ms, r(800)) in a concentration-dependent fashion irrespective of charge carrier (50% inhibitory concentration: I(peak), 656 microM; r(800), 104 microM). Rate of mono-exponential I(Ba) decay was linearly dependent on PB concentration. PB reduced channel availability by deepening non-steady-state inactivation curves without altering voltage dependence, slowed recovery from activity-induced unavailable states and produced use-dependent block. PB (100 microM) induced use-dependent block during physiological, high frequency pulse trains and overall depressed PQCC activity by two-fold.


CONCLUSIONS
The results support a PB pharmacological mechanism involving a modulated receptor with preferential slow, bimolecular, open channel block (K(d)= 15 microM). Clinical PB concentrations (<200 microM) inhibit PQCC during high frequency activation that reduces computed neurotransmitter release by 16-fold and is comparable to the magnitude of Ca(2+)-dependent facilitation, G-protein modulation and intrinsic inactivation that play critical roles in PQCC modulation underlying synaptic plasticity. The results are consistent with the hypothesis that PB inhibition of PQCCs contributes to central nervous system depression underlying anticonvulsant therapy and general anaesthesia.
Schober
A
A
The Department of Anesthesiology, New York University Langone Medical Center, New York, NY 10016, USA.
Sokolova
E
E
Gingrich
K J
KJ
D002121:Calcium Channel Blockers / Q000494:pharmacology*; D015220:Calcium Channels / Q000378:metabolism*; D018929:Cell Culture Techniques; D002460:Cell Line; D004305:Dose-Response Relationship, Drug; D055724:Electrophysiological Phenomena; D006801:Humans; D015640:Ion Channel Gating / Q000187:drug effects*; D010424:Pentobarbital / Q000494:pharmacology*; D011994:Recombinant Proteins / Q000378:metabolism; D013997:Time Factors; D014162:Transfection; D020864:Calcium Channels, N-Type
D002121:Calcium Channel Blockers; D015220:Calcium Channels; D010424:Pentobarbital; D011994:Recombinant Proteins; C105656:CACNA1A protein, human; D020864:Calcium Channels, N-Type