Mice 114 mg/kg i.p. injection

Xenopus oocytes expressed with human GABAA receptor alpha-1, beta-1 and gamma-2L subunits were used for activity evaluation. Oocytes were placed in a 250-ml recording chamber (Warner Instruments, Hamden, CT), and drug-induced currents were measured using standard two-electrode voltage clamp techniques with a Dagan TEV-201 amplifier. Oocytes were clamped at 270 mV and perfused with Barth’s solution at 1 ml/min by gravity flow. Oocytes were maintained at room temperature (21–23°C) during the electrophysiological experiments. Chloralose were delivered to the oocytes by gravity flow using an Autobank 8 perfusion system (AutoMate Scientific, Oakland, CA) at 2.4 ml/min. Base-line currents were recorded for 10 sec followed by a 15-sec drug application. The entire drug response was recorded for 1 min. Oocytes were given a 5- to 10-min recovery period between drug applications. For most experiments, GABAA receptors were activated using 30 mM GABA. Reversal potentials for GABA and a-chloralose were determined by measuring the current-voltage relationship of the responses. Current- voltage curves were measured by initially clamping oocytes at 2120 mV and ramping the membrane potential from 2120 mV to 170 mV over 750 msec. The ramp episode was repeated three times within 10 sec and the responses averaged. Before determination of the reversal potential of drug-induced responses, the base-line current was measured for the ramp protocol.