title: GABAB1 and GABAB2 receptor subunits co-expressed in cultured human RPE cells regulate intracellular Ca2+ via Gi/o-protein and phospholipase C pathways.
abstract: GABAB receptors associate with Gi/o-proteins that regulate voltage-gated Ca(2+) channels and thus the intracellular Ca(2+) concentration ([Ca(2+)]i), there is also reported cross-regulation of phospholipase C. These associations have been studied extensively in the brain and also shown to occur in non-neural cells (e.g. human airway smooth muscle). More recently GABAB receptors have been observed in chick retinal pigment epithelium (RPE). The aims were to investigate whether the GABAB receptor subunits, GABAB1 and GABAB2, are co-expressed in cultured human RPE cells, and then determine if the GABAB receptor similarly regulates the [Ca(2+)]i of RPE cells and if phospholipase C is involved. Human RPE cells were cultured from five donor eye cups. Evidence for GABAB1 and GABAB2 mRNAs and proteins in the RPE cell cultures was investigated using real time polymerase chain reaction, western blots and immunofluorescence. The effects of the GABAB receptor agonist baclofen, antagonist CGP46381, a Gi/o-protein inhibitor pertussis toxin, and the phospholipase C inhibitor U73122 on [Ca(2+)]i in cultured human RPE were demonstrated using Fluo-3. Both GABAB1 and GABAB2 mRNA and protein were identified in cell cultures of human RPE; antibody staining was co-localized to the cell membrane and cytoplasm. One-hundred micromolars of baclofen caused a transient increase in the [Ca(2+)]i of RPE cells regardless of whether Ca(2+) was added to the buffer. Baclofen-induced increases in the [Ca(2+)]i were attenuated by pre-treatment with CGP46381, pertussis toxin, and U73122. GABAB1 and GABAB2 are co-expressed in cell cultures of human RPE. GABAB receptors in RPE regulate the [Ca(2+)]i via a Gi/o-protein and phospholipase C pathway.
authors.affiliation: Department of Ophthalmology, Qilu Hospital, Shandong University, Jinan, Shandong 250012, China. Electronic address: zycheng2008@hotmail.com.
authors.forename: Z-Y
authors.initials: ZY
authors.lastname: Cheng
authors.affiliation: The Key Laboratory of Cardiovascular Remodeling and Function Research, Chinese Ministry of Education and Chinese Ministry of Health, Qilu Hospital, Shandong University, Jinan, Shandong 250012, China.
authors.forename: X-P
authors.initials: XP
authors.lastname: Wang
authors.affiliation: School of Optometry and Vision Science, Faculty of Health, and Institute of Health and Biomedical Innovation, Queensland University of Technology, Brisbane, QLD 4059, Australia.
authors.forename: K L
authors.initials: KL
authors.lastname: Schmid
authors.affiliation: Department of Ophthalmology, The Second Hospital, Jinan, Shandong 250001, China.
authors.forename: X-G
authors.initials: XG
authors.lastname: Han
mesh_terms: D001418:Baclofen / Q000494:pharmacology; D002118:Calcium / Q000378:metabolism*; D002478:Cells, Cultured; D004791:Enzyme Inhibitors / Q000494:pharmacology; D004963:Estrenes / Q000494:pharmacology; D058786:GABA-B Receptor Agonists / Q000494:pharmacology; D058788:GABA-B Receptor Antagonists / Q000494:pharmacology; D019206:GTP-Binding Protein alpha Subunits, Gi-Go / Q000037:antagonists & inhibitors / Q000378:metabolism*; D006801:Humans; D042541:Intracellular Space / Q000187:drug effects / Q000378:metabolism*; D037342:Pertussis Toxin / Q000494:pharmacology; D010721:Phosphinic Acids / Q000494:pharmacology; D011760:Pyrrolidinones / Q000494:pharmacology; D012333:RNA, Messenger / Q000378:metabolism; D011963:Receptors, GABA-A / Q000235:genetics / Q000378:metabolism*; D055213:Retinal Pigment Epithelium / Q000187:drug effects / Q000378:metabolism; D010738:Type C Phospholipases / Q000037:antagonists & inhibitors / Q000378:metabolism*
chemical_list: D004791:Enzyme Inhibitors; D004963:Estrenes; D058786:GABA-B Receptor Agonists; D058788:GABA-B Receptor Antagonists; C546264:GABRB1 protein, human; C495710:GABRB2 protein, human; D010721:Phosphinic Acids; D011760:Pyrrolidinones; D012333:RNA, Messenger; D011963:Receptors, GABA-A; C060229:1-(6-((3-methoxyestra-1,3,5(10)-trien-17-yl)amino)hexyl)-1H-pyrrole-2,5-dione; C086109:3-aminopropyl-cyclohexylmethylphosphinic acid; D037342:Pertussis Toxin; D010738:Type C Phospholipases; D019206:GTP-Binding Protein alpha Subunits, Gi-Go; D001418:Baclofen; D002118:Calcium
keywords: G(i/o)-protein; GABA(B) receptor; calcium; phospholipase C; retinal pigment epithelium; γ-aminobutyric acid