Pentobarbital produces activation and block of {alpha}1{beta}2{gamma}2S GABAA receptors in rapidly perfused whole cells and membrane patches: divergent results can be explained by pharmacokinetics.

Millimolar concentrations of the barbiturate pentobarbital (PB) activate gamma-aminobutyric acid (GABA) type A receptors (GABARs) and cause blockade reported by a paradoxical current increase or "tail" upon washout. To explore the mechanism of blockade, we investigated PB-triggered currents of recombinant alpha(1)beta(2)gamma(2S) GABARs in whole cells and outside-out membrane patches using rapid perfusion. Whole cell currents showed characteristic bell-shaped concentration dependence where high concentrations triggered tail currents with peak amplitudes similar to those during PB application. Tail current time courses could not be described by multi-exponential functions at high concentrations (> or =3,000 microM). Deactivation time course decayed over seconds and was slowed by increasing PB concentration and application time. In contrast, macropatch tail currents manifested eightfold greater relative amplitude, were described by multi-exponential functions, and had millisecond rise times; deactivation occurred over fractions of seconds and was insensitive to PB concentration and application time. A parsimonious gating model was constructed that accounts for macropatch results ("patch" model). Lipophilic drug molecules migrate slowly through cells due to avid partitioning into lipophilic subcellular compartments. Inclusion of such a pharmacokinetic compartment into the patch model introduced a slow kinetic component in the extracellular exchange time course, thereby providing recapitulation of divergent whole cell results. GABA co-application potentiated PB blockade. Overall, the results indicate that block is produced by PB concentrations sixfold lower than for activation involving at least three inhibitory PB binding sites, suggest a role of blocked channels in GABA-triggered activity at therapeutic PB concentrations, and raise an important technical question regarding the effective rate of exchange during rapid perfusion of whole cells with PB.