Specific binding of 125I-hippocampal cholinergic neurostimulating peptide (HCNP) to rat brain membranes: characterization and regional distribution.
An undecapeptide-hippocampal cholinergic neurostimulating peptide (HCNP), originally purified from young rat hippocampus, enhances cholinergic phenotype development in the medial septal nucleus in vitro. To survey and characterize the HCNP receptor within the central nervous system, we used iodinated HCNP as a labeled ligand. In preliminary experiments, [125I]HCNP binding was highest in the crude P2 membrane fraction, so all subsequent experiments were performed using this fraction. The binding was saturable and reversible, and unlabeled ligand inhibited it. Scatchard analysis of the concentration-dependent saturation of binding indicated a single population of non-interacting sites with K(d) 4.0+/-0.7 nM and B(max) 10.7+/-3.8 pmol/mg protein. Dissociation experiments revealed a dissociation constant (K(-1)) of 0.07 min(-1). Inhibition experiments using HCNP and its shorter peptide fragments suggested that the active binding site resided close to the peptide's C-terminal sequence. Since [125I]HCNP binding was found in crude P2 membrane fractions from animals at all ages examined, HCNP may also perform important functional roles in the adult brain. Further, the predominant distribution of the receptor in the P2 membrane fraction, and the similarity in distribution patterns between the binding site and HCNP-precursor protein mRNA expression suggest that the peptide exerts its functions in the vicinity of the dendrites of the neurons that produce it.
Morishita
Manabu
M
Second Department of Internal Medicine, Medical School, Nagoya City University, Mizuho-ku, Nagoya 467-8601, Japan.
Otsuka
Yasushi
Y
Matsukawa
Noriyuki
N
Suzuki
Hiko
H
Nakazawa
Hideka
H
Maki
Mina
M
Katou
Hideki
H
Ueda
Ryuzou
R
Ojika
Kosei
K
D000818:Animals; D001921:Brain / Q000166:cytology / Q000378:metabolism*; D007457:Iodine Radioisotopes / Q000378:metabolism; D009479:Neuropeptides / Q000378:metabolism*; D011485:Protein Binding / Q000502:physiology; D051381:Rats; D017208:Rats, Wistar; D013347:Subcellular Fractions / Q000378:metabolism
D007457:Iodine Radioisotopes; D009479:Neuropeptides; C075274:hippocampal cholinergic neurostimulating peptide