This proposal represents the extension of my long-standing interest in neurotransmitter receptors and their regulation. The program I have described here will extend my expertise, up to now limited to pharmacological approaches to receptor function, to biochemical and eventually to gene regulation studies in the examination of serotonin 5HT-1 receptors and their regulation. Serotonergic function has been implicated as an important factor in several conditions of clinical interest including normal sleep, depression, narcolepsy and pain perception. Serotonin receptors have been divided into 5HT-1 and 5HT-2 subtypes based on their pharmacological specificities. Recent work in our laboratory has focused on serotonin 5HT-1 receptors and, in particular, the possibility of defective 5HT-1 receptor function in childhood autism. Our long-term goal is not only to gain a pharmacological and structural understanding of 5HT-1 receptors, but also an understanding of 5HT-1 receptor regulation. To this end, the program of training described here is divided into two parts. In Phase I of this program I will use my background in pharmacology to further characterize the 5HT-1 receptor, and will attempt further purification of the bovine 5HT-1 receptor previously solubilized in this laboratory employing affinity chromatography and high pressure liquid chromatography. Although purification of the receptor will allow many new approaches to the study of receptor structure and function, I will mainly utilize this in the preparation of anti-receptor monoclonal antibodies using in vitro immunization techniques. During Phase I, I will also acquire familiarity with some of the more straightforward molecular biology techniques. These will be applied in Phase II, in which monoclonal antibodies developed in Phase I will be used in an attempt to isolate 5HT-1 receptor mRNA for preparation of cDNA, to be used in turn in the study of 5HT-1 receptor regulation.
