CELLULAR MECHANISMS OF PRESYNAPTIC INHIBITION

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Modulation of transmitter release is integral to central nervous system (CNS) function. Understanding this phenomenon is, thus, important in understanding the vertebrate CNS, in which presynaptic terminals are mostly small and inaccessible. The goal of this project is to investigate amino acid receptor-mediated modulation of transmitter release from presynaptic terminals in the lamprey spinal cord. This preparation enables unequalled access to the terminal using imaging and electrophysiology. Properties of axonal voltage operated channels and presynaptic terminals will be investigated; particularly voltage operated Ca2+ channels. Activation of Ca2+ channels, while recording intracellular Ca2+ concentration by imaging Ca2+-sensitive fluorescent dye loaded into the axon through the recording electrode, will enable their localization. Voltage Step paradigms and pharmacological agents specific to.subtypes of Ca2+ channel will enable identification of channels at the presynaptic terminal. Concurrent to these experiments imaging of dye loaded paired postsynaptic somata will demonstrate spatial relationships between Ca2+ channels in the terminal and the site of synaptic contacts. Paired cell recordings of this form will be used to investigate the effects of amino acid application upon synaptic transmission. Application of reagents active at gamma aminobutyric acid (GABA) receptors will allow a detailed analysis of location and subtypes of GABA receptors involved in modulation of synaptic transmission. Observation of the effects of GABA receptor activation on voltage-activated Ca2+ entry to the presynaptic terminal and interference of 2nd messenger function by drug application to the interior of the terminal will be used to study the cellular mechanisms by which GABA receptor activation is transduced. Techniques of recording electrophysiologically and microfluorimetrically will also be pursued to investigate modes and sites of action of glutamate metabotropic receptors (mGluRs) in the pre- and post-synaptic elements of the lamprey spinal cord. Common mechanisms between mGluR and GABA receptor activation will be sought in the control of transmitter release, as well as mechanisms specific to each class of receptor. The roles of presynaptic GABA receptors and mGluRs in modulating motor output will be investigated. The effect of the activation or blockade of these receptors on spinal motor output will be tested. Additionally Ca2+ imaging of the axons will be used to investigate phase- and time-dependent changes in Ca2+ entry at different locations in the spinal cord. Thus it will be possible to relate cellular effects of presynaptic receptor activation to the control of the spinal motor system.