Isolation, characterization, and distribution of a novel neuropeptide, Rana RFamide (R‐RFa), in the brain of the European green frog Rana esculenta

Abstract

A novel neuropeptide of the RFamide peptide family was isolated in pure form from a frog (Rana esculenta) brain extract by using reversed-phase high performance liquid chromatography in combination with a radioimmunoassay for mammalian neuropeptide FF (NPFF). The primary structure of the peptide was established as Ser-Leu-Lys- Pro-Ala-Ala-Asn-Leu-Pro-Leu- Arg-Phe-NH(2). The sequence of this neuropeptide, designated Rana RFamide (R-RFa), exhibits substantial similarities with those of avian LPLRFamide, gonadotropin-inhibitory hormone, and human RFRP-1. The distribution of R-RFa was investigated in the frog central nervous system by using an antiserum directed against bovine NPFF. In the brain, immunoreactive cell bodies were primarily located in the hypothalamus, i.e., the anterior preoptic area, the suprachiasmatic nucleus, and the dorsal and ventral hypothalamic nuclei. The most abundant population of R-RFa-containing neurons was found in the periependymal region of the suprachiasmatic nucleus. R-RFa- containing fibers were widely distributed throughout the brain from the olfactory bulb to the brainstem, and were particularly abundant in the external layer of the median eminence. In the spinal cord, scattered immunoreactive neurons were found in the gray matter. R-RFa-positive processes were found in all regions of the spinal cord, but they were more abundant in the dorsal horn. This study provides the first characterization of a member of the RFamide peptide family in amphibians. The occurrence of this novel neuropeptide in the hypothalamus and median eminence and in the dorsal region of the spinal cord suggests that, in frog, R-RFa may exert neuroendocrine activities and/or may be involved in the transmission of nociceptive stimuli.

Identification of 26RFa, a hypothalamic neuropeptide of the RFamide peptide family with orexigenic activity

“…1A. Absorbance was measured at 215 and 280 nm, and eluting fractions were assayed for NPFF-like immunoreactivity (NPFF-LI) as described (2). Fraction 61 was selected for further purification and successively chromatographed on a 0.46 ϫ 25-cm Shodex C8P-50 4E C 8 column, a 0.46 ϫ 25-cm Vydac 218TP54 C 18 column, a 0.46 ϫ 25-cm Shodex C8P-50 4E C 8 column, and a 0.46 ϫ 25-cm Vydac 218TP54 C 18 column, at a flow rate of 1 ml͞min, by using the elution conditions indicated in Fig.…”

“…Since then, several FMRFamide-related peptides (RFRPs), which all possess the Arg-Phe-NH 2 motif at their C terminus, have been isolated in various groups of invertebrates (see ref. 2 for review), and it has been found that a single species may express multiple members of RFRPs. In particular, in Caenorhabditis elegans, 22 genes encoding 59 distinct RFRPs have been identified (3).…”

A novel G protein-coupled receptor for gonadotropin-inhibitory hormone in the Japanese quail (Coturnix japonica): identification, expression and binding activity

“…The isolated LPLRF-amide peptide from bullfrog hypothalami possessed growth hormone-releasing activity, and was designated as frog growth hormone-releasing peptide (fGRP) (Koda et al 2002). This fGRP is structurally identical to Rana RF-amide (R-RFa) purified by Chartrel et al (2002). The fGRP precursor also encoded one fGRP and three related peptides (fGRP-RP-1, -2, and -3; Sawada et al 2002a), which were identified as mature LPXRF-amide peptides (Ukena et al 2003b).…”

Role of LPXRFamide peptide in the neuroendocrine regulation of reproduction in fish

“…neuropeptide FF (NPFF), neuropeptide AF (NPAF) and neuropeptide SF (NPSF) [4][5][6][7][8]; (b) LPXRFamide (X = L or Q) peptide group, e.g. gonadotropin-inhibitory hormone (GnIH), mammalian RFamide-related peptides (RFRPs), frog growth hormone-releasing peptide (fGRP) and goldfish LPXRFamide peptide (gfLPXRFa) [9][10][11][12][13][14][15][16][17][18][19][20]; (c) prolactin-releasing peptide (PrRP) group, e.g. PrRP31, PrRP20, crucian carp RFamide (C-RFa), salmon RFa and tilapia PrRP [21][22][23][24]; (d) metastin group, e.g.…”