Although thyroid hormones (THs) are strictly required during developing nervous system, the molecular bases of their action are poorly understood. The long-term goal of this project is to elucidate the molecular mechanism by which synaptic function is modulated by THs in embryonic tissue. The Neuromuscular junction (NMJ) of tadpole tail is an excellent system to study neuromodulation, because it dies by apoptosis in the presence of THs.
Specific Aim 1. Determine the distribution of cholinergic receptors and nerve terminals tadpole tails at several stages of development, The Characterization of the anatomical features of this synaptic system represents the first step towards understanding the non-genomic actions of THs. We will test the hypothesis that the AChRs responsible for synaptic transmission are preferential/y located within the extreme rostral of the muscular fiber.
Specific Aim 2. Determine the acute pre-synaptic action of THs on synaptic vesicle recycling and its relationship to intracellular calcium level. We will test the hypothesis that the enhanced mepc frequency observed in response to T3 results from augmented vesicle recycling. Two separated sub-aims will be considered: (a) to determine the role of THs in modulating vesicle recycling, and (b) to assess the role of intracellu/ar calcium level in mediating the actions of TH.
Specific Aim 3. Assess the acute postsynaptic action of thyroid hormones on the AChR. We will test the hypothesis that THs interact directly with the postsynaptic AC-receptor and secondly that the calcium mobilization by the AChR that is alter by T3 causes chanties in the AChR kinetic activity. The experiments will include three powerful techniques: Near Infrared (NIR) microscopy, Laser Scanning Confocal Microscopy (LSCM) and conventional electrophysiology. High and low affinity calcium dyes will be used to evaluate calcium levels in the pre- and post-synaptic regions. This study will provide information that is relevant to clinical applications including treatments in which T3 is being co-administrated and/or used. Importantly, neuronal apoptosis is now thought to play a significant role in the pathology of a number of degenerative diseases of the CNS (e.g. Parkinson's and Alzheimer's diseases) as well as in the periphery (for example, various neuropathies and retinal degeneration). The key role of apoptosis in these diverse physiological and pathological processes suggests that the pharmacology of apoptosis and the actions of THs will become important targets for biotechnology and for clinical applications.
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