THE NEURO-2A NEUROBLASTOMA CELL-LINE EXPRESSES [MET]-ENKEPHALIN AND VASOPRESSIN MESSENGER-RNA AND PEPTIDE

Mouse neuroblastoma Neuro-2a cells were examined for the expression of pro-enkephalin mRNA, protein, and Met-enkephalin ([Met]-Enk) peptide. Reverse transcriptase/polymerase chain reaction (RT/PCR) and in situ hybridization demonstrated the presence of pro-enkephalin mRNA in thes e cells. Immunocytochemistry using an antibody which recognizes pro-en kephalin and high pressure liquid chromatography (HPLC) followed by ra dioimmunoassay indicated that pro-enkephalin was synthesized in these cells and processed to yield the bioactive pentapeptide, [Met]-Enk. Fu rthermore, release studies showed that the [Met]-Enk was secreted from these cells with high K+ stimulation. Using double labeling, in situ hybridization combined with immunocytochemistry, we demonstrated that prohormone convertase 2 (PC2) mRNA is colocalized with pro-enkephalin in the same Neuro-2a cells, suggesting that this enzyme may be respons ible for processing this precursor. We also showed the presence of vas opressin mRNA and arginine-vasopressin peptide in these cells using in situ hybridization and immunocytochemistry, respectively. Thus, the N euro-2a cells are a multiple neuropeptide-producing cell line and an e xcellent model for studying the mechanisms involved in the synthesis, intracellular targeting and processing of endogenous pro-enkephalin an d pro-vasopressin, as well as other transfected neuropeptide precursor s.