Neuropeptide FF (FLFQPQRFamide) was originally isolated from the bovine brain and found to have opiate modulating activity. To gain a better understanding of the mechanisms involved in the opioid modulatory role of NPFF, a study was initiated to isolate proNPFF specific cDNA. Initially, degenerate 27-mer oligonucleotides deduced from NPFF was used to screen rat hypothalamic cDNA library and was found to be difficult. To facilitate the cloning, it may be desirable to have multiple oligonucleotide probes (deduced from different parts of NPFF precursor protein). Thus in this study, NPFF-like peptides which are distinct from NPFF were isolated from the rat CNS and the purified peptides were then characterized. One of the purified peptide was characterized as SLAAPQRFamide. The peptide sequence was determined by sequencing and the specificity of the NPFF antiserum. The peptide sequence was further confirmed by comparing the endogenous peptide with the syntesized SLAAPQRFamide in a reverse phase HPLC. The synthetic peptide SLAAPQRFamide was found to bind to the NPFF receptor with a high affinity indicating that NPFF and SLAAPQRFamide may have the same precursor. We were able to partially sequence one other peptide and this peptide was found to have Pro-Ser-Leu-Ala at its n-terminal end. These additional peptide sequences will be used to prepare additional oligonucleotide probes to aid in the isolation of proNPFF specific cDNA. NPFF is highly localized within discrete central autonomic regions including nucleus of the solitary tract (NTS) and the lateral parabrachial nucleus. The possible role of NPFF in the autonomic regulation of cardiovascular responses in these regions was studied by examining the activation of NTS NPFF neurons, using Fos immunoreactivity. The results indicated that NPFF neurons in the NTS respond to specific cardiovascular stimulation produced by hypotension, but not to other stressful stimuli such as centrally-generated opiate withdrawal.
