Serotonin (5HT) and its receptors are part of a major neurotransmitter system, whose malfunction is implicated in a variety of human diseases. To understand the malfunction, and possibly alleviate its consequences, we need to know more about the receptors on which 5HT acts. For this purpose a multidisciplinary study will be made of the synthesis, structure and function of 5HT receptors, using frog oocytes and recombinant DNA research. 5HT receptors will 'transplanted' from the brain into the membrane of Xenopus oocytes by isolating messenger RNA, from rat and human brains, and injecting it into the oocytes. Translation of the foreign mRNA by the oocyte causes it to acquire functional 5HT receptors. Electrophysiological techniques will be used to study the ionic membrane channels opened, or closed, by 5HT action and the involvement of cyclic nucleotides, phosphoinostides and calcium ions as channel operators will be investigated with biochemical and electrophysiological methods. Serotonin receptor types will be characterized through pharmacological studies, and a search will be made for selective drugs. This information will help develop compounds that may be used in the treatment of disorders of the 5HT system. Brain mRNA will be the subject of several studies. For example, the receptors and channels expressed by mRNA from the normal and diseased human brain will be investigated for their possible involvement in diseases such as Alzheimer, Cushing and Huntington; and the ontogenetic development of 5HT receptors will be studied by isolating mRNA from the developing brains and expressing it in oocytes. To determine the structure of 5HT receptors two approaches will be used: purification of the receptors and gene cloning. Production of monoclonal anti-receptor antibodies and of synthetic oligonucleotide probes will help greatly in cloning the genes. However, the receptor/channel cloning work will start even before these are produced. mRNA preparations will be fractionated to separate mRNAs coding for different receptors and channels. The partially purified mRNA fractions coding for 5HT receptors and chloride channels will be used to construct cDNA libraries that will be screened by various procedures, including functional assays in oocytes.
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