The process of synaptic transnuission. appears to be highly conserved across species, allowing functional questions about the vertebrate central nervous system to be addressed in more tractable systems. For example, over 30 genes implicated in synaptic transmission have been identified in the nematode, Caenorhabditis elegans, and homologs of these genes have subsequently been identified in vertebrates. Defects in neurotransmission due to neurodegeneration can lead to severe deficits in nervous system function evident in disorders such as Parkinson's and Alzeihmer's diseases. One of the most powerful approaches to study the roles of synaptic proteins is to examine synaptic physiology in mutant animals lacking these proteins. A preparation suitable for detailed electrophysiological analysis of synaptic function in C.elegans has not previously been available, primarily due to the technical difficulties of dissecting such a small organism. Preliminary data provided in this proposal demonstrate the feasibility of whole-cell, voltage-clamp recording synaptic activity in a newly developed C. elegans body wall neuromuscular preparation. In order to use this preparation to probe synaptic transmission in mutant animals, a baseline characterization of the wildtype synapse is required. The C. elegans neuromuscular junction is innervated by both GABAergic and cholinergic motor neurons. In order to characterize release at this synapse it will be necessary to study these two inputs separately.
Aim I will identify pharmacological tools to separate the two synaptic inputs.
Aim 2 will use these pharmacological tools to characterize spontaneous GABAergic and cholinergic synaptic activity.
Aim 3 will develop methods to evoke GABA and ACh release at the neuromuscular junction.
In aim 4, the Ca 2+-dependence of evoked release will be quantified.
Aim 5 will test the synaptic plasticity of the neuromuscular synapse using protocols to elicit paired-pulse and short-term facilitation, augmentation and post-tetanic potentiation. This preparation is expected to provide a valuable new tool in which to combine the strengths of C. elegans genetics and molecular biology with electrophysiological analyses. Completion of this project is expected to lay the foundation for future evaluation of over 30 proteins implicated in synaptic transmission.
|Broadie, Kendal S; Richmond, Janet E (2002) Establishing and sculpting the synapse in Drosophila and C. elegans. Curr Opin Neurobiol 12:491-8|
|Richmond, Janet E; Broadie, Kendal S (2002) The synaptic vesicle cycle: exocytosis and endocytosis in Drosophila and C. elegans. Curr Opin Neurobiol 12:499-507|
|Koushika, S P; Richmond, J E; Hadwiger, G et al. (2001) A post-docking role for active zone protein Rim. Nat Neurosci 4:997-1005|
|Richmond, J E; Weimer, R M; Jorgensen, E M (2001) An open form of syntaxin bypasses the requirement for UNC-13 in vesicle priming. Nature 412:338-41|
|Richmond, J E; Jorgensen, E M (1999) One GABA and two acetylcholine receptors function at the C. elegans neuromuscular junction. Nat Neurosci 2:791-7|
|Richmond, J E; Davis, W S; Jorgensen, E M (1999) UNC-13 is required for synaptic vesicle fusion in C. elegans. Nat Neurosci 2:959-64|