Strychnine-insensitive binding of [3H]glycine to synaptic membranes in rat brain, treated with Triton X-100.
Binding of radiolabelled glycine, a putative inhibitory neurotransmitter in mammalian lower central structures, was examined by using the synaptic membranes of the brain of rat, treated with Triton X-100. This treatment with Triton markedly potentiated the binding of [3H]glycine detected at 2 degrees C and 30 degrees C. However, this binding was not affected by three different convulsants, strychnine, picrotoxin and bicuculline. The binding was saturable at 2 degrees C, with increasing concentrations of [3H]glycine up to 1 microM. Scatchard analysis revealed that the binding sites consisted of a single component with a Kd of 202 nM and a Bmax of 1.74 pmol/mg protein. The binding was inhibited, not only by various amino acids structurally related to glycine, including D- and L-serine and D-, L- and beta-alanine, but was also eliminated by some peptides containing glycine, such as gamma-D- and gamma-L-glutamylglycine, glycine methylester and N-methyl-glycine. In addition, the strychnine-insensitive binding of [3H]glycine was significantly abolished by numerous quinoxaline antagonists for excitatory amino acid receptors in the brain. These results suggest that synaptic membranes of brain, treated with Triton X-100, are useful to detect the strychnine-insensitive binding of [3H]glycine and superior to untreated membranes in terms of the freedom from the confounding effects of some endogenous amino acids.
Ogita
K
K
Department of Pharmacology, Setsuman University, Osaka, Japan.
Suzuki
T
T
Yoneda
Y
Y
D000818:Animals; D001224:Aspartic Acid / Q000031:analogs & derivatives / Q000494:pharmacology; D001923:Brain Chemistry / Q000187:drug effects*; D005998:Glycine / Q000378:metabolism*; D066298:In Vitro Techniques; D007700:Kinetics; D016202:N-Methylaspartate; D017830:Octoxynol; D011092:Polyethylene Glycols / Q000494:pharmacology; D051381:Rats; D011919:Rats, Inbred Strains; D013331:Strychnine / Q000494:pharmacology*; D013570:Synaptic Membranes / Q000187:drug effects / Q000378:metabolism*
D001224:Aspartic Acid; D011092:Polyethylene Glycols; D016202:N-Methylaspartate; D017830:Octoxynol; D013331:Strychnine; D005998:Glycine