Abstracts

Extrasynaptic GABA[sub]A[/sub] receptors (GABARs) produce small but long-lived tonic currents in response to the low levels of ambient GABA (up to [asymp]1 mM) that are present outside GABAergic synapses. GABARs may also be clustered near synapses (perisynaptic), where they respond to brief surges of extrasynaptic GABA resulting from synaptic overflow. The extrasynaptic GABARs are a mixed population, although [alpha]4[beta]x[delta] and [alpha]5[beta]3[gamma]2L receptors are particularly important in the thalamus and hippocampus. To function efficiently in the extrasynaptic milieu, it has been suggested that these receptors must be non-desensitizing and sensitive to low levels of GABA., We tested this hypothesis by applying GABA with a rapid drug application system to recombinant GABARs expressed in HEK 293T cells., We report that [alpha]4[beta]1[delta], [alpha]4[beta]2[delta], [alpha]4[beta]3[delta], and [alpha]5[beta]3[gamma]2L receptors responded to low levels of GABA, although the GABA EC[sub]50[/sub] varied considerably ([alpha]5[beta]3[gamma]2L [gt] [alpha]4[beta]3[delta] [gt] [alpha]4[beta]1[delta] [gt] [alpha]4[beta]2[delta]). Since extrasynaptic GABA is thought to change slowly, it is not surprising that all [alpha]4[beta]x[delta] combinations activated slowly, requiring nearly 20 ms to reach peak current in response to even saturating GABA concentrations. In contrast, [alpha]5[beta]3[gamma]2L receptors activated 10 fold more rapidly. All four GABAR isoforms desensitized extensively, with residual currents that were only 30% of peak current during a 4 sec application of 1 mM GABA. Interestingly, the residual currents, generally accepted as measures of tonic inhibition, were very similar at all but the lowest GABA concentrations. However, the GABA concentration producing this maximal [ldquo]tonic[rdquo] current varied from 3-20 [mu]M, depending on the receptor isoform. Thus, each GABAR responded to its own unique range of GABA concentrations. Finally, [alpha]5[beta]3[gamma]2L receptors expressed a small spontaneous current (10% of the response to 1 [mu]M GABA) that was blocked by picrotoxin. This current was not recorded in cells transfected with [alpha]1[beta]3[gamma]2L, [alpha]4[beta]3[gamma]2L, [alpha]1[beta]3[delta] or [alpha]4[beta]3[delta] receptors., In summary, we report that the kinetic properties of individual extrasynaptic GABARs are tuned for efficient response to extrasynaptic activation and suggest that the relatively high GABA EC[sub]50[/sub] and rapid activation of [alpha]5[beta]3[gamma]2L GABARs renders them ideally suited to function as perisynaptic receptors. Furthermore, a low level of spontaneous activity could allow [alpha]5[beta]3[gamma]2L GABARs to modulate neuronal activity, even in the absence of ambient GABA. In contrast, the highly sensitive but slowly activating [alpha]4[beta]x[delta] GABARs would function well as extrasynaptic receptors. Since ambient GABA levels may be altered during development as well as by drugs and disease states, these findings raise the exciting possibility that selective modulation of extrasynaptic GABAR subtypes may modify specific components of the tonic currents to treat illnesses such as epilepsy., (Supported by K08 NS 045122 to AHL, R01 NS33300 to RLM.)