Multiple Cl(-)-independent binding sites for the excitatory amino acids: glutamate, aspartate and cysteine sulfinate in rat brain membranes.
As we have recently reported that Cl(-)-dependent glutamate (GLU) binding reflects GLU accumulation into membrane vesicles, the characteristics, kinetics and pharmacological specificities of L-[3H]glutamate (L-[3H]GLU) binding to crude rat brain synaptic membranes, were investigated in Cl(-)-free medium. L-[3H]GLU binding was systematically compared to that of L-[3H]cysteine sulfinate (L-[3H]CSA) and L-[3H]ASP), two other putative excitatory amino acids. A high affinity site was determined for each of these radioactive ligands (L-[3H]GLU: Kd = 0.14 microM, Bm = 3.4 pmol/mg protein; L-[3H]CSA: Kd = 0.07 microM, Bm = 2.2 pmol/mg protein; L-[3H]ASP: Kd = 5.8 microM, Bm = 31.2 pmol/mg protein). The pharmacological specificity of these Cl(-)-independent binding sites indicate the existence of at least 3 distinct high affinity sites, all different from the Cl(-)-dependent GLU binding 'site': one having a similar affinity for GLU and CSA, a second one preferring CSA, and a third one preferring ASP. Among the large quantity of structural analogs of the neuroexcitatory amino acids tested, only endogenous compounds (GLU, ASP and CSA) (except hydroxylamine-o-sulfate) were able to interact efficiently. No inhibition by classical agonists and antagonists (such as N-methyl-D-aspartate, quisqualate, kainate, 2-amino-4-phosphonobutyrate, or 2-amino-5-phosphonovalerate) was found. In addition to their high specificity, these Cl(-)-independent sites possess most other biochemical characteristics of receptor proteins.
D000818:Animals; D001224:Aspartic Acid / Q000378:metabolism*; D001921:Brain / Q000378:metabolism*; D002712:Chlorides / Q000502:physiology*; D003545:Cysteine / Q000031:analogs & derivatives* / Q000378:metabolism; D005971:Glutamates / Q000378:metabolism*; D018698:Glutamic Acid; D007700:Kinetics; D008297:Male; D008566:Membranes / Q000378:metabolism; D018377:Neurotransmitter Agents; D051381:Rats; D011919:Rats, Inbred Strains; D012964:Sodium / Q000502:physiology; D013570:Synaptic Membranes / Q000378:metabolism
D002712:Chlorides; D005971:Glutamates; D018377:Neurotransmitter Agents; D001224:Aspartic Acid; D018698:Glutamic Acid; D012964:Sodium; D003545:Cysteine; C013461:cysteine sulfinic acid