GLUTAMATE-RECEPTOR (GAMMA-AMINO-3-HYDROXY-5-METHYL-4-ISOXAZOLE PROPIONATE AND KAINATE SUBTYPE) ACTIVITY ENHANCED BY DIZOCILPINE (MK801) IN RAT HIPPOCAMPUS - RAPID CHEMICAL KINETIC MEASUREMENTS OF SODIUM FLUX AND RECEPTOR DESENSITIZATION WITH NATIVE MEMBRANES
P. Serfozo et Dj. Cash, GLUTAMATE-RECEPTOR (GAMMA-AMINO-3-HYDROXY-5-METHYL-4-ISOXAZOLE PROPIONATE AND KAINATE SUBTYPE) ACTIVITY ENHANCED BY DIZOCILPINE (MK801) IN RAT HIPPOCAMPUS - RAPID CHEMICAL KINETIC MEASUREMENTS OF SODIUM FLUX AND RECEPTOR DESENSITIZATION WITH NATIVE MEMBRANES, European journal of biochemistry, 228(2), 1995, pp. 498-505
L-Glutamate mediated transmembrane influx of Na-22(+) into a native me
mbrane vesicle preparation from rat hippocampus was resolved into comp
onents due to glutamate receptor and Na+-glutamate symport, by kinetic
analysis and pharmacological specificity. Measurements made with a qu
ench-flow technique showed that receptor-mediated cation exchange proc
eeded in two phases, the faster phase progressively attenuated by a de
sensitization process with a half time of approximately 90 ms. The inf
lux of Na-22(+) continued in the second phase with a half time of 6 s.
Receptor-mediated Na-22(+) influx was replicated with the glutamate m
imetics, alpha-amino-3-hydroxy-5-methyl-4-isoxazole propionate and kai
nate but not with N-methyl-D-aspartate and was inhibited by 6- cyano-7
-nitroquinoxaline-2,3-dione but not by 2-amino-5-phosphonovaleric acid
. Receptor-mediated Na-22(+) influx did not require glycine and was pH
de pendent; influx was inhibited al pH values less than pH 5.0. Thus,
the receptor-mediated activity was of the gamma-amino-3-hydroxy-5-met
hyl-4-isoxazole propionate-kainate subtype. The N-methyl-D-aspartate r
eceptor inhibitor 0,11-dihydro-5H-dibenzo[a,d]cyclohepten-5,10-imine (
dizocilpine or MK801), increased receptor-mediated transmembrane Na-22
(+) flux in both phases, but did not accelerate receptor desensitizati
on. Receptor-mediated radiotracer exchange was the major contribution
to Na-22(+) influx in times less than 5 s and the receptor-mediated re
sponses could be measured with a small correction for sodium-glutamate
symport or in the presence of a glutamate-uptake inhibitor. A third p
hase of Na-22(+) influx accessed a different internal volume and proce
eded with the same first-order rate constant as [H-3]glutamate uptake
when measured simultaneously. This was shown to be mediated by the glu
tamate uptake mechanism, with a half time of 20-40 s varying with the
preparation. The Na+-glutamate symport became the largest cause of Na-
22(+) influx after at least 5 s, the final magnitude being, on average
, twofold larger than the receptor-mediated Na-22(+) influx.