title: Expression levels of the alpha4 subunit of the GABA(A) receptor in differentiated neuroblastoma cells are correlated with GABA-gated current.
abstract: The alpha4 subunit of the GABA(A) receptor (GABAR) is capable of rapid plasticity, increased by chronic exposure to positive GABA modulators, such as the neurosteroid 3alpha-OH-5alpha[beta]-pregnan-20-one (THP). Here, we show that 48 h exposure of differentiated neuroblastoma cells (IMR-32) to 100 nM THP increases alpha4 expression, without changing the current density or the concentration-response curve. Increased expression of alpha4-containing GABAR was verified by a relative insensitivity of GABA (EC(20))-gated current to modulation by the benzodiazepine (BZ) lorazepam (0.01-100 microM), and potentiation of current by flumazenil and RO15-4513, characteristic of alpha4betagamma2 pharmacology. In contrast to THP, compounds which decrease GABA-gated current, such as the BZ inverse agonist DMCM, the GABAR antagonist gabazine and the open channel blocker penicillin, decreased alpha4 expression after a 48 h exposure, without changing BZ responsiveness. However, pentobarbital, another positive GABA modulator, increased alpha4 expression, while the BZ antagonist flumazenil had no effect. In order to test whether changes in current were responsible for increased alpha4 expression, decreases in the Cl(-) driving force were produced by chronic exposure to the NKCC1 blocker bumetanide (10 microM). When applied under these conditions of reduced GABA-gated current, THP failed to increase alpha4 expression. The results of this study suggest that alpha4 expression is correlated with changes in GABA-gated current, rather than simply through ligand-receptor interactions. These findings have relevance for GABAR subunit plasticity produced by fluctuations in endogenous steroids across the menstrual cycle, when altered BZ sensitivity is reported.
authors.affiliation: Department of Physiology and Pharmacology, SUNY Downstate Medical Center, 450 Clarkson Avenue, Brooklyn, NY 11203, USA.
authors.forename: Xiangping
authors.initials: X
authors.lastname: Zhou
authors.forename: Sheryl S
authors.initials: SS
authors.lastname: Smith
mesh_terms: D001569:Benzodiazepines / Q000494:pharmacology; D002034:Bumetanide / Q000494:pharmacology; D002243:Carbolines / Q000494:pharmacology; D002454:Cell Differentiation; D045744:Cell Line, Tumor; D003513:Cycloheximide / Q000494:pharmacology; D054314:Drug Inverse Agonism; D018757:GABA Modulators / Q000494:pharmacology; D058785:GABA-A Receptor Agonists; D058787:GABA-A Receptor Antagonists; D006801:Humans; D015640:Ion Channel Gating; D008024:Ligands; D009447:Neuroblastoma; D010424:Pentobarbital / Q000494:pharmacology; D010852:Picrotoxin / Q000494:pharmacology; D011280:Pregnanolone / Q000494:pharmacology; D021122:Protein Subunits / Q000096:biosynthesis; D011500:Protein Synthesis Inhibitors / Q000494:pharmacology; D011963:Receptors, GABA-A / Q000096:biosynthesis*; D049994:Sodium Potassium Chloride Symporter Inhibitors; D064506:Solute Carrier Family 12, Member 2; D005680:gamma-Aminobutyric Acid / Q000494:pharmacology*
chemical_list: D001569:Benzodiazepines; D002034:Bumetanide; D002243:Carbolines; D003513:Cycloheximide; D018757:GABA Modulators; D058785:GABA-A Receptor Agonists; D058787:GABA-A Receptor Antagonists; D008024:Ligands; D010424:Pentobarbital; D010852:Picrotoxin; D011280:Pregnanolone; D021122:Protein Subunits; D011500:Protein Synthesis Inhibitors; D011963:Receptors, GABA-A; D049994:Sodium Potassium Chloride Symporter Inhibitors; D064506:Solute Carrier Family 12, Member 2; D005680:gamma-Aminobutyric Acid; C546260:GABRA4 protein, human; C578877:SLC12A2 protein, human; C034818:methyl 6,7-dimethoxy-4-ethyl-beta-carboline-3-carboxylate