We have previously demonstrated that uroguanylin (UGN) significantly inhibits NHE3-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-PK1). 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 PKG inhibitor KT5823 or by the PKA inhibitor H-89. The effects of UGN in vitro were found to be similar to those obtained by microperfusion. Indeed, we have observed that incubation of LLC-PK1 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-PK1 cells. Finally, treatment of LLC-PK1 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 upon mechanisms by which guanylin peptides are intricately involved in the maintenance of salt and water homeostasis.
- Renal Microperfusion
- Bicarbonate Reabsorption
- Copyright © 2011, American Journal of Physiology - Renal Physiology