COMPARATIVE-STUDIES ON BINDING OF 3 DIFFERENT LIGANDS TO THE N-METHYL-D-ASPARTATE RECOGNITION DOMAIN IN BRAIN SYNAPTIC-MEMBRANES TREATED WITH TRITON X-100
Z. Pingping et al., COMPARATIVE-STUDIES ON BINDING OF 3 DIFFERENT LIGANDS TO THE N-METHYL-D-ASPARTATE RECOGNITION DOMAIN IN BRAIN SYNAPTIC-MEMBRANES TREATED WITH TRITON X-100, Brain research, 609(1-2), 1993, pp. 253-261
Treatment with a low concentration of Triton X-100 almost tripled the
binding of 3]D,L-(E)-2-amino-4-propyl-5-phosphono-3-pentenoic acid (CG
P 39653), a novel competitive antagonist at an N-methyl-D-aspartate (N
MDA)-sensitive subclass of brain excitatory amino acid receptors, in s
ynaptic membranes of the rat brain. The binding linearly increased wit
h increasing protein concentrations of up to 0.4 mg/ml and also increa
sed in proportion to incubation time with a plateau within 60 min afte
r the initiation of incubation at 2-degrees-C in Triton-treated membra
nes. Elevation of incubation temperature from 2-degrees-C to 30-degree
s-C resulted in a marked decrease in the binding at equilibrium by 80%
, and a maximal level was obtained within 1 min after the initiation o
f incubation at 30-degrees-C with a gradual decline of up to 10 min. B
ound [H-3]CGP 3%53 was rapidly dissociated by the addition of excess u
nlabeled L-glutamic acid (Glu), and the time required to attain comple
te dissociation was 60 min at 2-degrees-C and 1 min at 30-degrees-C, r
espectively. Among several agonists and antagonists tested, Glu was th
e most potent displacer of [H-3]CGP 39653 binding with progressively l
ess potent displacement by D-2-amino-5-phosphonovaleric, /-)-3-(2-carb
oxypiperazin-4-yl)propyl-1-phosphonic (CPP), D-2-amino-7-phosphonohept
anoic, N-methyl-D-aspartic and N-methyl-L-aspartic acids. Agonists at
the glycine (Gly) domain on the NMDA receptor ionophore complex invari
ably inhibited the binding of [H-3]CGP 39653 without virtually affecti
ng the binding of [H-3]Cpp or [H-3]Glu in a manner that was sensitive
to antagonism by 4 different antagonists at the Gly domain. In contras
t, the Gly antagonist 1-hydroxy-3-aminopyrrolidone-2 markedly potentia
ted [H-3]CPP binding with the binding of both [H-3]CGP 39653 and [H-3]
Glu being unaltered. These results suggest that [H-3]CGP 39653 may pre
dominantly label an antagonist-preferring form of the NMDA domain in a
state different from that favorable to labeling by [H-3]CPP.