The ability to reproduce is dependent upon the periodic release by the brain of the neuropeptide luteinizing hormone releasing hormone (LHRH). The long-term objective underlying the proposed research is to achieve an understanding of how the brain generates, synchronizes and modulates the periodic output of LHRH. The hypothesis that LHRH directly alters specific conductances of neurons that participate in the neural circuits controlling periodic LHRH release will be tested. It is known that LHRH can affect neuronal activity. The neurons of the arcuate nucleus (ARC) are known to affect LHRH output. The ARC is also the location of the only beta- endorphin-containing cell bodies in the forebrain and beta-endorphin is an important modulator of LHRH release. Thus, the effects of LHRH on ARC neurons, including identified beta-endorphin neurons, will be examined. In addition, we will determine if LHRH has direct effects on LHRH cells. The experiments in this proposal are intended to 1) investigate the nature of the responses to LHRH by neurons that mediate LHRH output, 2) identify the cellular mechanisms of action of LHRH that cause these responses and 3) determine how identified beta-endorphin neurons in the ARC and identified LHRH neurons in the preoptic/septal regions are affected electrophysiologically by LHRH. Electrical recordings of these neurons will be obtained using in vitro slices of guinea pig brain. The first specific aim is to determine some of the characteristics of the effects of LHRH. That is, the percent of ARC cells that alter their spontaneous activity in response to the presence of extracellular LHRH will be established, as will the direction, the duration, the time to onset and the repeatability of that response. Dose-response relationships and the effects of antagonists and fragments of LHRH on neuronal activity also will be determined. The second specific aim is to determine the nature of the membrane response to LHRH and the specific ionic conductances that underly the observed changes in total membrane conductance. These conductances will be characterized using intracellular recording and current- and voltage-clamp techniques. Four potassium conductances, for which there are a priori reasons for hypothesizing that they play a role in the effects induced by LHRH, will be specifically examined. The third specific aim is to characterize the response of individual beta-endorphin neurons to LHRH administration. This will be accomplished by immunocytochemical staining of the ARC neurons that were recorded from intracellularly and tested with LHRH. Finally, the proposal aims to characterize the electrical responses of LHRH-containing neurons to LHRH administration by using a combination of electrophysiology and immunocytochemistry similar to that used in the ARC but on neurons in the preoptic/septal areas.
