These experiments will test the hypothesis that changing the driving force for the synaptic currents gated by activation of the inhibitory GABAA receptor is an effective way to increase the efficacy of these currents and thereby increase neuronal inhibition. The rationale for this hypothesis is based on Ohm's law: the amount of current that flows through inhibitory GABAA channels can be altered either by changing the GABAA receptor-mediated conductance, or by changing the driving force for the current. Although many available drugs increase the GABAA receptor-mediated conductance, the driving force has not been manipulated. This is because the processes that establish the anionic transmembrane equilibrium, and thus the driving force for GABAA currents, are just now being understood at a molecular level. One determinant of the neuronal anionic transmembrane equilibrium is CIC-2, a cloned inward rectifier that stabilizes the Cl- equilibrium potential near the resting membrane potential. To test our hypothesis, adenovirus vectors will be used to express CIC-2 in cultured sensory neurons and in granule cell neurons of the hippocampal dentate gyms. These neurons maintain high intracellular Cl- levels, are depolarized rather than hyperpolarized by GABAA receptor activation, and do not naturally express ClC-2. The impact of CIC-2 on Cl- homeostasis, the efficacy of GABAergic inhibition, and potentiation of inhibition by anticonvulsants will then be tested at the cellular level using whole-cell recordings, and at the network level by measuring input-output responses of the dentate gyrus. These experiments will determine whether the effects of anticonvulsant drugs that increase the conductance through which inhibitory synaptic currents flow can be complimented by increasing the driving force for those currents. This new approach to the modulation of neuronal inhibition may provide a foundation for the development of alternate strategies for the treatment of intractable epilepsy.
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