1. Intracellular recordings were obtained from hippocampal pyramidal neurons maintained in vitro. Measurements were made of the conductance change induced by iontophoretically applied γ-aminobutyric acid (GABA) and, using voltage-clamp techniques, of inhibitory postsynaptic currents resulting from activation of inhibitory pathways. 2. Analysis of GABA iontophoretic charge-response curves indicated that there was considerable variation among neurons with respect to the slope of this relation. 3. The placement of the GABA-containing pipette did not appear to be responsible for the observed variation, since vertical repositioning of the pipette did not alter the slope of the charge-response relationship. 4. Steady iontophoresis of GABA from one barrel of a double-barreled pipette markedly affected the charge-response relation obtained when short pulses were applied to the other barrel. The curve was shifted to the left, and the slope was decreased. Concomitantly, the enhanced GABA-induced responses were prolonged. 5. Similar alterations in GABA responsiveness were observed when the uptake blocker, nipecotic acid, was iontophoretically applied. Furthermore, bath application of saline containing a reduced sodium concentration (25% of control) also produced a prolongation of GABA-mediated responses. 6. Under voltage clamp, inhibitory postsynaptic currents were observed to have biphasic decays. The initial, fast decay was prolonged by an average of 18% by nipecotic acid, whereas the later, slow phase was prolonged by 23%. 7. The results of these studies support the hypothesis that a saturable GABA uptake system is responsible for the observed variation in the charge-response curves and, in turn, underlies the apparent sensitizing effect of excess GABA application. The results also suggest that a reduction of transmitter uptake affects the time course of inhibitory postsynaptic currents in the hippocampus. © 1985 Plenum Publishing Corporation.