L-[3H]glutamate binding to a membrane preparation from crayfish muscle.
The binding of L-[3H]glutamate to an isolated membrane preparation from crayfish tail muscle has been studied. The muscle homogenate was osmotically shocked, frozen and thawed, and thoroughly washed before incubation with L-[3H]glutamate. The preparation showed high specific binding of L-glutamate with a KD of 0.12 microM and Bmax of 4.7 pmol/mg protein measured in Tris/HCl pH 7.3 and at 4 degrees C. Nonspecific binding was 5-10% of total binding. The glutamate binding was highly stereospecific [K0.5 (D-glutamate), 270 microM] and showed a high degree of discrimination between L-glutamate and L-aspartate [K0.5 (L-aspartate), 54 microM]. In mammalian CNS preparations potent agonists of L-glutamate such as kainate and N-methyl-D-aspartate had no effect at 1 mM, and quisqualate was a weak inhibitor of L-glutamate binding [K0.5 (quisqualate), 162 microM]. Ibotenate was the most potent inhibitor [K0.5 (ibotenate), 0.27 microM], and various esters of L-glutamate were of intermediate potency as displacers of L-[3H]glutamate binding (K0.5 values from 6 to 60 microM). The glutamate binding site from crayfish muscle is clearly different from any of the subclasses of glutamate receptors in mammalian CNS. A possible physiological function of the binding site is a postsynaptic receptor for glutamate, either an extra-junctional or a junctional receptor.
Syvertsen
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Apothekernes Laboratorium A.S., Oslo, Norway.
Fonnum
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D000818:Animals; D001224:Aspartic Acid / Q000031:analogs & derivatives / Q000378:metabolism / Q000494:pharmacology; D003400:Astacoidea / Q000378:metabolism*; D001667:Binding, Competitive; D002118:Calcium / Q000494:pharmacology; D002462:Cell Membrane / Q000378:metabolism; D005971:Glutamates / Q000378:metabolism*; D018698:Glutamic Acid; D007051:Ibotenic Acid / Q000494:pharmacology; D007608:Kainic Acid / Q000494:pharmacology; D009132:Muscles / Q000378:metabolism*; D016202:N-Methylaspartate; D010069:Oxadiazoles / Q000494:pharmacology; D016318:Quisqualic Acid; D017470:Receptors, Glutamate; D017981:Receptors, Neurotransmitter / Q000187:drug effects / Q000378:metabolism*; D013237:Stereoisomerism
D005971:Glutamates; D010069:Oxadiazoles; D017470:Receptors, Glutamate; D017981:Receptors, Neurotransmitter; D007051:Ibotenic Acid; D001224:Aspartic Acid; D018698:Glutamic Acid; D016202:N-Methylaspartate; D016318:Quisqualic Acid; D007608:Kainic Acid; D002118:Calcium