Demonstration of specific binding sites for pituitary adenylate cyclase activating polypeptide (PACAP) in rat astrocytes.
The high and low affinity binding sites for PACAP were identified in rat astrocytes using [125I]PACAP27 as the labeled ligand. Scatchard analysis of displacement of the bound tracer by unlabeled PACAP27 indicated the existence of two classes of binding sites, with the dissociation constant (Kd) = 1.22 +/- 0.4 nM, the binding maximal capacity (Bmax) = 821 +/- 218 fmols/mg protein for the high affinity binding site, and Kd = 0.59 +/- 0.06 microM, Bmax = 563 +/- 12 pmols/mg protein for the low affinity binding site, respectively. The specificity of [125I]PACAP27 binding was tested using PACAP38 and peptides structurally related to PACAP, such as VIP, GHRF, PHI, secretin and glucagon. PACAP38 completely displaced the binding of [125I]PACAP27 and Scatchard analysis also indicated the presence of two classes of binding sites with similar Kd and Bmax to those for PACAP27. VIP and GHRF competed with [125I]PACAP27, but to a much lesser extent than unlabeled PACAP27 in binding. Other peptides tested did not displace the binding of [125I]PACAP27 at 10(-6) M.
Tatsuno
I
I
US-Japan Biomedical Research Laboratories, Tulane University Hebert Center, Belle Chasse, LA 70037.
Gottschall
P E
PE
Köves
K
K
Arimura
A
A
D000704:Analysis of Variance; D000818:Animals; D001253:Astrocytes / Q000032:analysis / Q000378:metabolism*; D001665:Binding Sites; D001667:Binding, Competitive; D001921:Brain / Q000166:cytology; D002462:Cell Membrane / Q000032:analysis / Q000378:metabolism; D002478:Cells, Cultured; D005260:Female; D007700:Kinetics; D009479:Neuropeptides / Q000378:metabolism*; D051219:Pituitary Adenylate Cyclase-Activating Polypeptide; D011247:Pregnancy; D051381:Rats; D011919:Rats, Inbred Strains
C495403:Adcyap1 protein, rat; D009479:Neuropeptides; D051219:Pituitary Adenylate Cyclase-Activating Polypeptide