The human neuroblastoma cell line, IMR-32 possesses a GABAA receptor lacking the benzodiazepine modulatory site.
GABAA receptors were identified in IMR-32 cell membranes by the binding of [35S]t-butyl-bicyclophosphorothionate ([35S]TBPS) to the chloride channel. GABA (IC50 2.2 microM), muscimol (IC50 0.8 microM), picrotoxin (IC50 1.7 microM), pentobarbitone (IC50 108 microM), etomidate (IC50 53 microM), chlormethiazole (IC50 98 microM) and Ro 5-3663 (IC50 280 microM) all inhibited [35S]TBPS binding. The potency of these drugs at the [35S]TBPS binding site in IMR-32 cell membranes did not correlate with their potency on [35S]TBPS binding to rat cortical membranes (linear correlation of pIC50 values, r = 0.75, NS). No specific binding of the benzodiazepine ligands [3H]flunitrazepam or [3H]Ro 15-4513 to IMR-32 cell membranes was observed. Chloride efflux from IMR-32 cells was studied using the fluorescent dye 6-methoxy-N-(3-sulphopropyl) quinolinium. Chloride efflux was stimulated by GABA and muscimol (0.1-100 microM) but not by the GABAB agonist baclofen (100 microM). In the absence of exogenous GABA chloride efflux was stimulated by chlormethiazole (1-100 microM) in a picrotoxin-sensitive manner. Flurazepam (1-100 microM) both alone and in the presence of GABA had no effect on chloride efflux. It is concluded that IMR-32 cells contain a functional GABAA receptor which differs from that in rat cortex both in its general pharmacology and specifically in the absence of the allosteric modulatory site sensitive to benzodiazepines.
Astra Neuroscience Research Unit, London, U.K.
S M
SM
Anderson
R J
RJ
De Souza
A J
AJ
Cross
D000818:Animals; D001386:Azides / Q000378:metabolism; D001569:Benzodiazepines / Q000378:metabolism; D001643:Bridged Bicyclo Compounds / Q000378:metabolism; D019086:Bridged Bicyclo Compounds, Heterocyclic; D002462:Cell Membrane / Q000378:metabolism; D018118:Chloride Channels; D005445:Flunitrazepam / Q000378:metabolism; D006801:Humans; D007700:Kinetics; D008565:Membrane Proteins / Q000378:metabolism; D009447:Neuroblastoma / Q000378:metabolism*; D011869:Radioligand Assay; D051381:Rats; D011963:Receptors, GABA-A / Q000378:metabolism*; D014407:Tumor Cells, Cultured
D001386:Azides; D001643:Bridged Bicyclo Compounds; D019086:Bridged Bicyclo Compounds, Heterocyclic; D018118:Chloride Channels; D008565:Membrane Proteins; D011963:Receptors, GABA-A; D001569:Benzodiazepines; D005445:Flunitrazepam; C037476:tert-butylbicyclophosphorothionate; C042957:Ro 15-4513