title: Changes in the activity and expression of protein phosphatase-1 accompany the differential regulation of NHE3 before and after the onset of hypertension in spontaneously hypertensive rats.
abstract: OBJECTIVE
  The Na(+) /H(+) exchanger NHE3 activity decreases in the proximal tubule of spontaneously hypertensive rats (SHRs) as blood pressure increases, and this reduction is correlated with higher NHE3 phosphorylation levels at the PKA consensus site serine 552. This study tested the hypothesis that this lowered NHE3 activity is associated with an increase in PKA activity and expression, and/or a decrease in protein phosphatase-1 (PP1) activity and expression.
  
  
  METHODS
  Proximal tubule NHE3 activity was measured as the rate of bicarbonate reabsorption by stationary microperfusion. NHE3 phosphorylation and protein expression were determined by immunoblotting. PKA and PP1 activities were determined using specific substrates under optimal enzymatic conditions.
  
  
  RESULTS
  The PKA activator, 6-MB-cAMP, increased the phosphorylation levels of NHE3 at serine 552 in the renal cortex; this increase happens to a much greater extent in young pre-hypertensive SHRs (Y-SHRs) compared to adult SHRs with established hypertension (A-SHRs). Likewise, the inhibitory effect of 6-MB-cAMP on NHE3 transport activity was much more pronounced in the proximal tubules of Y-SHRs than in those of A-SHRs. Renal cortical activity of PKA was not significantly different between Y-SHRs and A-SHRs. On the other hand, Y-SHRs exhibited higher protein phosphatase 1 (PP1) activity, and their expression of the PP1 catalytic subunit PP1α in the renal cortex was also higher than in A-SHRs.
  
  
  CONCLUSIONS
  Collectively, these results support the idea that the lower NHE3 transport activity and higher phosphorylation occurring after the development of hypertension in SHRs are due, at least in part, to reduced PP1-mediated dephosphorylation of NHE3 at serine 552.
authors.affiliation: Laboratory of Genetics and Molecular Cardiology, Heart Institute (InCor), University of São Paulo Medical School, São Paulo, Brazil.
authors.forename: R O
authors.initials: RO
authors.lastname: Crajoinas
authors.forename: T D
authors.initials: TD
authors.lastname: Pessoa
authors.forename: M V
authors.initials: MV
authors.lastname: Rodrigues
authors.forename: G
authors.initials: G
authors.lastname: Malnic
authors.forename: A C C
authors.initials: AC
authors.lastname: Girardi
mesh_terms: D000818:Animals; D017868:Cyclic AMP-Dependent Protein Kinases / Q000378:metabolism; D006973:Hypertension / Q000378:metabolism*; D015151:Immunoblotting; D007687:Kidney Tubules, Proximal / Q000378:metabolism; D008297:Male; D010766:Phosphorylation; D054645:Protein Phosphatase 1 / Q000378:metabolism*; D051381:Rats; D011918:Rats, Inbred SHR; D011921:Rats, Inbred WKY; D060888:Real-Time Polymerase Chain Reaction; D000074083:Sodium-Hydrogen Exchanger 3; D017923:Sodium-Hydrogen Exchangers / Q000378:metabolism*
chemical_list: C000613387:Slc9a3 protein, rat; D000074083:Sodium-Hydrogen Exchanger 3; D017923:Sodium-Hydrogen Exchangers; D017868:Cyclic AMP-Dependent Protein Kinases; D054645:Protein Phosphatase 1
keywords: PKA; dephosphorylation; hypertension; renal proximal tubule; sodium transport