title: Mechanisms underlying the inhibitory effects of uroguanylin on NHE3 transport activity in renal proximal tubule.
abstract: We previously demonstrated that uroguanylin (UGN) significantly inhibits Na(+)/H(+) exchanger (NHE)3-mediated bicarbonate reabsorption. In the present study, we aimed to elucidate the molecular mechanisms underlying the action of UGN on NHE3 in rat renal proximal tubules and in a proximal tubule cell line (LLC-PK(1)). The in vivo studies were performed by the stationary microperfusion technique, in which we measured H(+) secretion in rat renal proximal segments, through a H(+)-sensitive microelectrode. UGN (1 μM) significantly inhibited the net of proximal bicarbonate reabsorption. The inhibitory effect of UGN was completely abolished by either the protein kinase G (PKG) inhibitor KT5823 or by the protein kinase A (PKA) inhibitor H-89. The effects of UGN in vitro were found to be similar to those obtained by microperfusion. Indeed, we observed that incubation of LLC-PK(1) cells with UGN induced an increase in the intracellular levels of cAMP and cGMP, as well as activation of both PKA and PKG. Furthermore, we found that UGN can increase the levels of NHE3 phosphorylation at the PKA consensus sites 552 and 605 in LLC-PK(1) cells. Finally, treatment of LLC-PK(1) cells with UGN reduced the amount of NHE3 at the cell surface. Overall, our data suggest that the inhibitory effect of UGN on NHE3 transport activity in proximal tubule is mediated by activation of both cGMP/PKG and cAMP/PKA signaling pathways which in turn leads to NHE3 phosphorylation and reduced NHE3 surface expression. Moreover, this study sheds light on mechanisms by which guanylin peptides are intricately involved in the maintenance of salt and water homeostasis.
authors.affiliation: Department of Physiology and Biophysics, Biomedical Sciences Institute, University of São Paulo, São Paulo/SP, Brazil. Lucilialessa17@gmail.com
authors.forename: Lucília M A
authors.initials: LM
authors.lastname: Lessa
authors.forename: Luciene R
authors.initials: LR
authors.lastname: Carraro-Lacroix
authors.forename: Renato O
authors.initials: RO
authors.lastname: Crajoinas
authors.forename: Camila N
authors.initials: CN
authors.lastname: Bezerra
authors.forename: Rafael
authors.initials: R
authors.lastname: Dariolli
authors.forename: Adriana C C
authors.initials: AC
authors.lastname: Girardi
authors.forename: Manassés C
authors.initials: MC
authors.lastname: Fonteles
authors.forename: Gerhard
authors.initials: G
authors.lastname: Malnic
mesh_terms: D000818:Animals; D001639:Bicarbonates / Q000378:metabolism*; D002227:Carbazoles / Q000494:pharmacology; D002460:Cell Line; D002478:Cells, Cultured; D000242:Cyclic AMP / Q000378:metabolism; D017868:Cyclic AMP-Dependent Protein Kinases / Q000037:antagonists & inhibitors; D006152:Cyclic GMP / Q000378:metabolism; D017869:Cyclic GMP-Dependent Protein Kinases / Q000037:antagonists & inhibitors; D007546:Isoquinolines / Q000494:pharmacology; D007687:Kidney Tubules, Proximal / Q000166:cytology / Q000187:drug effects* / Q000378:metabolism; D008297:Male; D045265:Natriuretic Peptides / Q000494:pharmacology*; D047428:Protein Kinase Inhibitors / Q000494:pharmacology; D051381:Rats; D017208:Rats, Wistar; D000074083:Sodium-Hydrogen Exchanger 3; D017923:Sodium-Hydrogen Exchangers / Q000378:metabolism*; D013449:Sulfonamides / Q000494:pharmacology
chemical_list: D001639:Bicarbonates; D002227:Carbazoles; D007546:Isoquinolines; D045265:Natriuretic Peptides; D047428:Protein Kinase Inhibitors; C000613387:Slc9a3 protein, rat; D000074083:Sodium-Hydrogen Exchanger 3; D017923:Sodium-Hydrogen Exchangers; D013449:Sulfonamides; C073601:KT 5823; C084362:uroguanylin; D000242:Cyclic AMP; D017868:Cyclic AMP-Dependent Protein Kinases; D017869:Cyclic GMP-Dependent Protein Kinases; D006152:Cyclic GMP; C063509:N-(2-(4-bromocinnamylamino)ethyl)-5-isoquinolinesulfonamide