This grant proposes to examine short term, frequency-dependent plasticity of excitatory and inhibitory synaptic function with the goal of contributing to an understanding of events which occur during the development of seizures in an epileptic focus and the spread of seizures from a focal area of abnormality to the entire cortex. The central hypothesis is that differential regualtion of inhibitory and excitatory synapses is responsible for the development and spread of seizure activity in the cortex. It is known that under most circumstances, repetitive activation causes inhibition to diminish and excitation to potential. In order to analyze such processes at the cellular level, a simplified preparation was developed to allow whole cell patch clamp recording of both neurons in a synaptic pair and the application of quantal analysis to help determine the mechanisms underlying synaptic plasticity. In the hippocampal cultures, synaptic inhibition profoundly decrements with repeated stimulation and this is due to presynaptic factors. Excitatory postsynaptic currents also decrement during repetitive action, also via a presynaptic mechanisms, whereas spontaneous transmitter released is paradoxically facilitated. Even under baseline, non-stimulated conditions, there are differences in the mechanisms underlying neurotransmitter release between inhibitory and excitatory neurons, and these may play a role in the differences in plasticity. All of the observed forms of synaptic plasticity are dependent on changes in Ca in the synaptic terminals and it is hypothesized that several Ca-dependent processes must be involved. The details of the Ca dependence will be characterized and the mechanisms by which changes in intraterminal Ca differential affect the release of neurotransmitter at excitatory and inhibitory terminals will be determined. Short term plasticity will also be studied in more mature tissue in hippocampal slice cultures. In this preparation, as the slice matures, excitatory synapse potentiate with repetitive activation. Using similar techniques to those employed in the cell culture system, the mechanisms underlying the potentiation and the development of the potentiating mechanisms will be analyzed. Finally, using a new technique which allows quantitative amplification of mRNAs from single cells, mRNA expression profiles of identified excitatory and inhibitory neurons, both in the cell culture ant the slice culture will be examined. The differences in the coordinated molecular expression in these two phenotypes will be analyzed, focusing on synaptic proteins, Ca channels, Ca-dependent enzymes which are part of the neurosecretory mechanisms and other Ca binding proteins. The molecular bases of the different forms of short term plasticity which the excitatory and inhibitory neurons exhibit will be determined. It is hypothesized that seizure develop in areas of abnormal excitability because of a differential regulation of excitatory and inhibitory neurons and that seizures spread throughout the brain by similar mechanisms. It is hoped that an increased understand of the nature of this plasticity in these two distinct cell types will lead to the development of strategies to suppress seizure development and spread.
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