Properties of AMPA receptors expressed in rat cerebellar granule cell cultures: Ca2+ influx studies.
Cultured cerebellar granule cells become vulnerable to excitatory amino acids, especially to NMDA and kainate, by 9 days in vitro. In the same time, the sensitivity of cells to (RS)-alpha-amino-3-hydroxy-5-methyl-isoxazole-4-propionate (AMPA), in terms of AMPA-induced toxicity or 45Ca2+ uptake, was very low. The low AMPA responsiveness was due to receptor desensitization, because agents known to block desensitization, cyclothiazide and the lectins concanavalin A and wheat germ agglutinin, rendered granule cells vulnerable to AMPA and produced a pronounced stimulation of 45Ca2+ accumulation. 45Ca2+ influx was induced specifically by AMPA-receptor stimulation, because it was blocked virtually completely by 2,3-dihydroxy-6-nitro-7-sulfamoylbenzoquinoxaline (NBQX) and the benzodiazepine GYKI 52466 (selective non-NMDA receptor antagonists). Nevertheless, indirect routes activated by cellular responses to AMPA-receptor stimulation contributed significantly to the overall 45Ca2+ influx. These included Ca2+ uptake through NMDA-receptor channels, voltage-sensitive Ca2+ channels, and via Na+/Ca2+ exchange. However, nearly one-fifth of the total 45Ca2+ influx remained unaccounted for and this estimate was similar to 45Ca2+ influx observed under Na(+)-free conditions. This observation suggested that a significant proportion of the Ca2+ flux passes through the AMPA-receptor channel proper, a view supported by Co2+ uptake into nearly all granule cells on exposure to AMPA in the presence of cyclothiazide. Results are discussed in light of the reported AMPA receptor-subunit composition of cerebellar granule cells in vitro.
Hack
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N
Graduate School of Neurosciences, Netherlands Institute for Brain Research, Amsterdam.
Balázs
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R
D000818:Animals; D014151:Anti-Anxiety Agents; D001569:Benzodiazepines / Q000494:pharmacology; D001581:Benzothiadiazines / Q000494:pharmacology; D002118:Calcium / Q000378:metabolism*; D002132:Calcium Radioisotopes; D002470:Cell Survival / Q000187:drug effects; D002478:Cells, Cultured; D002531:Cerebellum / Q000187:drug effects / Q000378:metabolism*; D003035:Cobalt / Q000378:metabolism; D003208:Concanavalin A / Q000494:pharmacology; D018846:Excitatory Amino Acids / Q000037:antagonists & inhibitors; D007608:Kainic Acid / Q000494:pharmacology; D016202:N-Methylaspartate / Q000494:pharmacology; D011810:Quinoxalines / Q000494:pharmacology; D051381:Rats; D018091:Receptors, AMPA / Q000037:antagonists & inhibitors / Q000378:metabolism*; D014909:Wheat Germ Agglutinins / Q000494:pharmacology; D018350:alpha-Amino-3-hydroxy-5-methyl-4-isoxazolepropionic Acid / Q000494:pharmacology
D014151:Anti-Anxiety Agents; D001581:Benzothiadiazines; D002132:Calcium Radioisotopes; D018846:Excitatory Amino Acids; D011810:Quinoxalines; D018091:Receptors, AMPA; D014909:Wheat Germ Agglutinins; C061950:GYKI 52466; D003208:Concanavalin A; C062865:2,3-dioxo-6-nitro-7-sulfamoylbenzo(f)quinoxaline; D001569:Benzodiazepines; D003035:Cobalt; D016202:N-Methylaspartate; D018350:alpha-Amino-3-hydroxy-5-methyl-4-isoxazolepropionic Acid; C004639:cyclothiazide; D007608:Kainic Acid; D002118:Calcium