title: Dopamine D1-like receptor-mediated inhibition of Cl/HCO3- exchanger activity in rat intestinal epithelial IEC-6 cells is regulated by G protein-coupled receptor kinase 6 (GRK 6).
abstract: The present study investigated the effect of dopamine D1-like receptor stimulation on the Cl-/HCO3- exchange activity in rat intestinal epithelial IEC-6 cells. The Cl-/HCO3- exchange activity was found to be a chloride-dependent, DIDS-sensitive and niflumate-insensitive process. The presence of the SLC26A6 anion exchanger was detected by both RT-PCR and immunoblotting analysis in IEC-6 cells, in which three different small interfering RNAs (siRNAs) targeting SLC26A6 markedly inhibited Cl-/HCO3- exchange. Activation of dopamine D1-like receptors with SKF 38393 inhibited Cl-/HCO3- exchanger activity, this being antagonized by the D1 selective antagonist SKF 83566. However, effects of SKF 38393 were maximal at 5 min of exposure to the agonist and rapidly diminished with no effect at 15 min, suggestive of agonist-induced desensitization of D1-like receptors. Pretreatment of cells with heparin, a non-selective inhibitor of G protein-coupled receptor kinases (GRKs), prevented the observed attenuation of SKF 38393-induced inhibition of Cl-/HCO3- exchange. Overnight pretreatment with anti-GRK6A and anti-GRK6B, but not with anti-GRK4 antibodies, prevented the loss of SKF 38393-mediated effects. Both PKA and PKC signaling pathways participate in SKF 38393-mediated inhibition of Cl-/HCO3- exchange. These findings suggest that SLC26A6 is at least one of the anion exchanger's family members responsible for Cl-/HCO3- exchange in IEC-6 cells. Dopamine D1 receptors in IEC-6 rapidly desensitize to D1-like agonist stimulation and GRK 6, but not GRK 4, appear to be involved in agonist-mediated responsiveness and desensitization.
authors.affiliation: Institute of Pharmacology and Therapeutics, Faculty of Medicine, Porto, Portugal.
authors.forename: Sónia
authors.initials: S
authors.lastname: Fraga
authors.forename: Yingjin
authors.initials: Y
authors.lastname: Luo
authors.forename: Pedro
authors.initials: P
authors.lastname: Jose
authors.forename: Kambiz
authors.initials: K
authors.lastname: Zandi-Nejad
authors.forename: David B
authors.initials: DB
authors.lastname: Mount
authors.forename: Patrício
authors.initials: P
authors.lastname: Soares-da-Silva
mesh_terms: D015647:2,3,4,5-Tetrahydro-7,8-dihydroxy-1-phenyl-1H-3-benzazepine / Q000031:analogs & derivatives / Q000494:pharmacology; D000818:Animals; D002478:Cells, Cultured; D027963:Chloride-Bicarbonate Antiporters / Q000037:antagonists & inhibitors / Q000235:genetics / Q000378:metabolism* / Q000502:physiology*; D018491:Dopamine Agonists / Q000494:pharmacology; D018492:Dopamine Antagonists / Q000494:pharmacology; D054771:G-Protein-Coupled Receptor Kinase 4; D054768:G-Protein-Coupled Receptor Kinases; D015870:Gene Expression; D006493:Heparin / Q000494:pharmacology; D006494:Heparin Antagonists / Q000494:pharmacology; D007150:Immunohistochemistry; D007413:Intestinal Mucosa / Q000187:drug effects / Q000201:enzymology / Q000378:metabolism*; D007422:Intestines / Q000737:chemistry / Q000166:cytology; D017346:Protein Serine-Threonine Kinases / Q000032:analysis / Q000037:antagonists & inhibitors / Q000378:metabolism*; D012333:RNA, Messenger / Q000032:analysis; D051381:Rats; D017447:Receptors, Dopamine D1 / Q000819:agonists* / Q000378:metabolism
chemical_list: D015647:2,3,4,5-Tetrahydro-7,8-dihydroxy-1-phenyl-1H-3-benzazepine; D027963:Chloride-Bicarbonate Antiporters; D018491:Dopamine Agonists; D018492:Dopamine Antagonists; D054771:G-Protein-Coupled Receptor Kinase 4; D054768:G-Protein-Coupled Receptor Kinases; D006493:Heparin; D006494:Heparin Antagonists; D017346:Protein Serine-Threonine Kinases; D012333:RNA, Messenger; D017447:Receptors, Dopamine D1; C044846:SKF83566; C089176:G-protein-coupled receptor kinase 6; C517854:Grk4 protein, rat