Combined electrophysiological, immunocytochemical and peptide release measurements in the hypothalamic slice

Sagittal hypothalamic slices were prepared from cycling female guinea pigs and incubated for 10-12 h during which time intracellular recordings were made from 274 arcuate and cell-poor zone (ARC-CPZ) neurons. Many of the cells exhibited spontaneous activity and a small percentage could be driven antidromically via median eminence stimulation and orthodromically via stria terminalis stimulation. During the long-term recording, the effluent medium was collected in 30 of the experiments and radioimmunoassayed using a high-titer, conformational antisera for the peptide luteinizing hormone-releasing hormone (LH-RH). The findings revealed that LH-RH is released in detectable amounts from slices prepared from a single hypothalamus, that the amount of LH-RH released is dependent on the stage of the cycle, and that it can be stimulated by known secretagogues like prostaglandin E2. Following the electrophysiological experiments, the slices were fixed and processed for immunocytochemistry using a specific antisera (EL-14) for LH-RH. LH-RH neurons were stained throughout the ARC-CPZ area with intense fiber staining in the median eminence. These findings demonstrate that the hypothalamic slice preparation can be utilized for combined electrophysiological, immunocytochemical and peptide release measurements.