Luteinizing hormone-releasing hormone (LHRH), oxytocin (OT) and vasopressin (VP) secreting cells in the rodent hypothalamus were selected for studying of the biophysical mechanisms involved in hormone secretion. Olfactory placodes from mouse embryos (day 16) were isolated and cultured on glass cover-slips for 2-3 weeks in defined medium. Hypothalamic slices of postnatal day 6 rat pups, which had been prelabelled with the vital fluorescent dye Fast Blue by intraperitoneal injection and retrograde transport were plated on glass cover-slips and cultured for 2-3 weeks. Both patch electrode voltage- and current-clamp techniques were applied. Electrical activities of putative LHRH-cells which emigrated from the olfactory placode displayed spontaneous spike discharges which were accompanied by transsynaptic activities. The soma region of these cells possessed initial transient sodium current (I-Na), potassium currents (I-R-V, I-R-A) and calcium current (I-Ca-t). Following electrophysiological analyses, the recorded cells were injected with Lucifer Yellow (L.Y.) for frequent identification by immunocytochemical staining for LHRH cells. Electrical activities of Fast Blue-stained cells in the supraoptic nucleus (SON) and paraventricular nucleus (PVN) also exhibited spontaneous regular and irregular discharges. Some cells displayed intermittent burst discharges. Some of these discharges were endogeneous, but transsynaptically driven discharges were also recorded. Following electrophysiological analyses, these LY-marked cells were also processed for their phenotypic identification (i.e., LHRH-, OT-, or VP-cells) by immunocytochemistry. An intracellular Ca2+-imaging and a high-time resolution single cell Ca2+-photometry system was constructed for the simultaneous measurement of electrophysiological activities and intracellular Ca2+ concentration changes in a single cell or a population of cells.
