CHARACTERIZATION OF [H-3] CGS-19755 BINDING-SITES IN THE RAT SPINAL-CORD

[H-3] Cis-4-phosphonomethyl-2-piperidine carboxylic acid ([H-3]CGS 197 55) was used to investigate the pharmacology and characteristics of th e N-methyl-D-aspartate (NMDA) receptor recognition site from Triton X- 100-treated membranes of rat spinal cord and cerebral cortex. The asso ciation of [H-3]CGS 19755 was biphasic in both spinal cord and cerebra l cortical membranes reaching a maximum after 5 min of incubation then decreasing to a steady level after an additional 10 min, suggesting t hat a proportion of the binding is unstable. The dissociation of the s table binding component was biphasic with rate constants at 4 degrees C of 1.55 and 0.020 min(-1) for the spinal cord and 1.48 and 0.051 min (-1) for the cerebral cortex. These multiple sites could not be captur ed in the saturation studies which were best fitted to a one-site mode l using non-linear regression analysis. Depending on the time of incub ation with [H-3]CGS 19755, K-D and B-max values differed; 9.9-26.1 nM and 25-96 fmol/mg protein vs 14.0-26.5 nM and 449-900 fmol/mg protein for spinal cord and cerebral cortex, respectively. The rank order of p otency of inhibiting [H-3]CGS 19755 binding was similar in both tissue s: L-glutamate > CGS 19755 = CPP > NMDA. The specific NMDA agonist cis -2,4-methanoglutamate potently inhibited [H-3]CGS 19755 binding as did MDL 100,925, although the latter was one order of magnitude less pote nt in the spinal cord than in the brain. The Hill coefficients were si gnificantly lower than unity. In both tissues, AMPA, kainate and glyci ne competed poorly with [H-3]CGS 19755. However, MK-801 had a distinct inhibitory action (46% at 100 mu M) in the spinal cord but a negligib le action in the cerebral cortex. It is concluded that in Triton-treat ed membranes, both stable and unstable binding can be distinguished. F urthermore, spinal and cerebral cortical sites show differences in the ir sensitivities to MK-801 and MDL 100,925.