The tripeptide TRH has been identified in several regions of brain and the periphery and has been shown to serve important roles in regulating CNS function. The localization of TRH to specific neurons and circuits is a prerequisite to the study of its role in neurotransmission and in complex physiological functions such as arousal and sensation. We propose to use chemical, lesion and histochemical techniques to localize TRH and its related enzymes in the nervous system. On the basis of our previous experimental findings, we have selected 3 sites for detailed studies: I) the olfactory bulb, II) a hippocampus and III) the sensory papillae of the tongue. All three have high concentrations of TRH. synaptic organizations that are favorable for localization studies, and well documented physiological correlates. We have prepared antibodies against TRH, developed HPLC procedures for the separation and identification of TRH and its metabolites, and developed enzymatic assays for the two enzymes that metabolize TRH (deamidase and pyroglutamase). For bulb, hippocampus and tongue, lesions (e.g. kainic acid, 6-hydroxydopamine, radio-frequency) will be made to selectively remove various neuronal populations. Then the tissue will be assayed by chemical and immunocytochemical methods for TRH and its related enzymes (deamidase and pyroglutamase) to determine their localizations. Localization of TRH to specific neuronal populations and synapses will provide a basis for the study of its physiology. TRH may play important physiological roles in sensation and arousal, and correspondingly TRH, its analogues or its metabolites may have therapeutic importance in a number of clinical states such as narcolepsy, barbiturate overdosage and depression.
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