The glycine receptor deficiency of the mutant mouse spastic: evidence for normal glycine receptor structure and localization.
Homozygotes of the mutant mouse spastic exhibit reduced binding of 3H-strychnine to homogenates from various regions of the CNS compared with unaffected littermates (White and Heller, 1982). Here we report evidence that the spastic mutation coincides with a reduced concentration and an unaltered structure of the glycine receptor in spinal cord. Scatchard analysis of 3H-strychnine binding revealed a single binding site with a Bmax of 267 +/- 62 fmol/mg protein for spastic and of 864 +/- 220 fmol/mg protein for control mice; no difference was found for the corresponding KD values. Also Ki values of glycine for 3H-strychnine binding and displacement of 3H-strychnine by beta-alanine and taurine were indistinguishable for both preparations. Photoaffinity labeling of synaptic membranes with 3H-strychnine identified an Mr = 48,000 polypeptide in both control and spastic mouse membranes. Tryptic digestion of these membranes produced radiolabeled peptide fragments of identical molecular weights, suggesting that the proteolytic cleavage sites around the antagonist binding site are conserved in the mutant glycine receptor protein. Glycine receptors from both control and mutant mice were purified by affinity chromatography on aminostrychnine agarose. SDS/PAGE revealed three polypeptides of Mr = 48,000, 58,000, and 93,000 in both receptor preparations. Monoclonal antibodies directed against different subunits of the glycine receptor were applied to an enzyme-linked immunosorbent assay. The same pattern of immunoreactivity was obtained for glycine receptor from spinal cord of spastic homozygotes, control mice, and rats, suggesting conservation of the antigenic epitopes in the mutant receptor.(ABSTRACT TRUNCATED AT 250 WORDS)
D000345:Affinity Labels / Q000378:metabolism; D000818:Animals; D005455:Fluorescent Antibody Technique; D051379:Mice; D008818:Mice, Neurologic Mutants / Q000378:metabolism*; D008856:Microscopy, Fluorescence; D018009:Receptors, Glycine; D017981:Receptors, Neurotransmitter / Q000276:immunology / Q000302:isolation & purification / Q000378:metabolism* / Q000648:ultrastructure; D013331:Strychnine / Q000378:metabolism; D014018:Tissue Distribution; D014316:Tritium; D014466:Ultraviolet Rays
D000345:Affinity Labels; D018009:Receptors, Glycine; D017981:Receptors, Neurotransmitter; D014316:Tritium; D013331:Strychnine