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 terminals 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.