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GABA_A Receptor Kinetics in the Cerebellar Nuclei: Evidence for Detection of Transmitter from Distant Release Sites

Jason R. Pugh ^* and Indira M. Raman ^* 

^* Institute for Neuroscience and Department of Neurobiology and Physiology, Northwestern University, Evanston, Illinois

Correspondence: Address reprint requests to Indira M. Raman, Dept. of Neurobiology and Physiology, 2205 Tech Drive, Northwestern University, Evanston, IL 60208. Tel.: 847-467-7912; Fax: 847-491-5211; E-mail: i-raman{at}northwestern.edu.

Neurons of the cerebellar nuclei receive GABAergic input from Purkinje cells. Purkinje boutons have several closely spaced presynaptic densities without GABA transporters, raising the possibility that neurotransmitter released by one presynaptic site diffuses to multiple postsynaptic sites. To test whether such local spillover may contribute to transmission, we studied gating of GABA_A receptors at 31–33°C in cerebellar nuclear neurons acutely dissociated from mice. Currents were evoked by rapid application of long steps, brief pulses, and high-frequency trains of GABA to outside-out patches. Receptors desensitized and deactivated rapidly, and dose-response measurements estimated an EC₅₀ of 30 µM. From these data, a kinetic scheme was developed that replicated the recorded currents. Next, we simulated diffusion of GABA in the synaptic cleft, constrained by previous electron microscopic data, and drove the kinetic GABA_A receptor model with modeled concentration transients. Simulations predicted receptor occupancies of 100% directly opposite the release site and 50% at distant postsynaptic densities, such that receptors up to 700 nm from a release site opened on the timescale of the inhibitory postsynaptic currents before desensitizing. Further simulations of probabilistic release from multiple-site boutons suggested that local spillover-mediated transmission slows the onset and limits the extent of depression during high-frequency signaling.

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