[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.