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Pharmacological characterization of metabotropic glutamate receptor‐mediated high‐affinity GTPase activity in rat cerebral cortical membranes
Author(s) -
Nishi Nobuyuki,
Odagaki Yuji,
Koyama Tsukasa
Publication year - 2000
Publication title -
british journal of pharmacology
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 2.432
H-Index - 211
eISSN - 1476-5381
pISSN - 0007-1188
DOI - 10.1038/sj.bjp.0703464
Subject(s) - metabotropic glutamate receptor , gtpase , metabotropic glutamate receptor 2 , glycine , glutamate receptor , metabotropic receptor , agonist , metabotropic glutamate receptor 1 , biology , chemistry , biochemistry , receptor , amino acid
Activation of heterotrimeric guanine nucleotide‐binding regulatory proteins (G‐proteins) functionally coupled to metabotropic glutamate receptors (mGluRs) was assessed by agonist‐induced high‐affinity GTPase (EC3.6.1.‐) activity in rat cerebral cortical membranes.L ‐Glutamate (1 m M ) stimulated high‐affinity GTPase activity to the same extent throughout the incubation period up to 20 min, in a Mg 2+ ‐dependent manner. The addition of 1 m M L ‐glutamate augmented V max of the enzyme activity (1670 to 3850 pmol mg −1 protein 15 min −1 ) with slight increase in K M value (0.26 to 0.63 μ M ). The high‐affinity GTPase activity was stimulated by the following compounds with a rank order of potency of (2 S ,2′ R ,3′ R )‐2‐(2′,3′‐dicarboxycyclopropyl) glycine (DCG‐IV) > (2 S ,1′ S ,2′ S )‐2‐(carboxycyclopyropyl)glycine ( L ‐CCG‐I) > L ‐glutamate ≥ 2 R ,4 R ‐4‐aminopyrrolidine‐2,4‐dicarboxylate [(2 R ,4 R )‐APDC] > 1 S ,3 R ‐1‐aminocyclopentane‐1,3‐dicarboxylate [(1 S ,3 R )‐ACPD] > ( S )‐4‐carboxy‐3‐hydroxyphenylglycine [( S )‐4C3HPG] > ( S )‐3‐carboxy‐4‐hydroxyphenylglycine [( S )‐3C4HPG] > ibotenate, but not by L ‐(+)‐2‐amino‐4‐phosphonobutyrate ( L ‐AP4), ( RS )‐3,5‐dihydroxyphenylglycine [( RS )‐3,5‐DHPG], quisqualate, or L ‐serine‐ O ‐phosphate ( L ‐SOP), indicative of involvement of group II mGluRs, in particular mGluR2. (2 S )‐α‐Ethylglutamate (EGLU), a presumably selective antagonist against group II mGluRs, inhibited DCG‐IV‐stimulated high‐affinity GTPase activity in a competitive manner with an apparent K B of 220 μ M .L ‐Glutamate‐stimulated activity was eliminated by pretreatment of the membranes with sulfhydryl alkylating agent N ‐ethylmaleimide (NEM) at 30–50 μ M , indicating that G‐proteins of the G i family are involved. These results indicate that mGluR agonist‐induced high‐affinity GTPase activity in rat cerebral cortical membranes may be used to detect the functional interaction between group II mGluRs, in particular mGluR2, and NEM‐sensitive G i proteins.British Journal of Pharmacology (2000) 130 , 1664–1670; doi: 10.1038/sj.bjp.0703464