Phosphodiesterase 5A inhibition decreases NHE-1 activity without altering steady state pH(i): role of phosphatases.
OBJECTIVE
This study aimed to identify the signaling pathway for the proposed link between phosphodiesterase-5A (PDE5A) inhibition and decreased cardiac Na(+)/H(+) exchanger (NHE-1) activity.


METHODS
NHE-1 activity was assessed in rat isolated papillary muscles by the Na(+)-dependent initial pH(i) recovery from a sustained acidosis (ammonium prepulse). ERK1/2, p90RSK and NHE-1 phosphorylation state during acidosis was determined.


RESULTS
PDE5A inhibition (1 μmol/L sildenafil, SIL) did not modify basal pH(i) but significantly blunted pH(i) recovery after sustained acidosis. Although preventing ERK1/2- p90RSK signaling pathway (10 μmol/L U0126) mimicked SIL effect, SIL did not blunt the acidosis-mediated increase in kinases activation. SIL+U0126 did not show additive effect on NHE-1 activity. Then, we hypothesized that SIL could be activating phophasatases (PP1 and/or PP2A) to directly dephosphorylate NHE-1 despite preserved ERK1/2-p90RSK activation. Non-specific phosphatases inhibition (1 μmol/L okadaic acid) canceled SIL effect on pH(i) recovery from acidosis. Same result was observed by inhibiting PP2A either with a lower dose of okadaic acid (1 nmol/L) or, more specifically, with 100 μmol/L endothall. Consistently, NHE-1 phosphorylation at Ser703 increased after acidosis, SIL prevented this effect and PP2A inhibition (endothall) reverted SIL effect.


CONCLUSIONS
We suggest that PDE5A inhibitors decrease NHE-1 phosphorylation and activity through a mechanism that involves PP2A activation.
Centro de Investigaciones Cardiovasculares, Facultad de Ciencias Médicas, Universidad Nacional de La Plata, La Plata, Argentina.
Romina G
RG
Díaz
Mariela B
MB
Nolly
Carolina
C
Massarutti
María J
MJ
Casarini
Carolina D
CD
Garciarena
Irene L
IL
Ennis
Horacio E
HE
Cingolani
Néstor G
NG
Pérez
D000138:Acidosis / Q000188:drug therapy; D000818:Animals; D002070:Butadienes / Q000494:pharmacology; D054706:Cyclic Nucleotide Phosphodiesterases, Type 5 / Q000378:metabolism*; D003998:Dicarboxylic Acids / Q000494:pharmacology; D006863:Hydrogen-Ion Concentration; D019950:Mitogen-Activated Protein Kinase 1 / Q000378:metabolism; D048052:Mitogen-Activated Protein Kinase 3 / Q000378:metabolism; D009570:Nitriles / Q000494:pharmacology; D019319:Okadaic Acid / Q000494:pharmacology; D010210:Papillary Muscles / Q000187:drug effects / Q000378:metabolism; D010726:Phosphodiesterase Inhibitors / Q000494:pharmacology*; D010766:Phosphorylation; D010879:Piperazines / Q000494:pharmacology; D054645:Protein Phosphatase 1 / Q000037:antagonists & inhibitors / Q000378:metabolism / Q000502:physiology*; D054648:Protein Phosphatase 2 / Q000037:antagonists & inhibitors / Q000378:metabolism / Q000502:physiology*; D011687:Purines / Q000494:pharmacology; D051381:Rats; D038744:Ribosomal Protein S6 Kinases, 90-kDa / Q000378:metabolism; D000068677:Sildenafil Citrate; D017923:Sodium-Hydrogen Exchangers / Q000378:metabolism*; D013450:Sulfones / Q000494:pharmacology
D002070:Butadienes; D003998:Dicarboxylic Acids; D009570:Nitriles; D010726:Phosphodiesterase Inhibitors; D010879:Piperazines; D011687:Purines; D017923:Sodium-Hydrogen Exchangers; D013450:Sulfones; C113580:U 0126; C069139:growth factor-activatable Na-H exchanger NHE-1; C100030:endothall; D019319:Okadaic Acid; D000068677:Sildenafil Citrate; D038744:Ribosomal Protein S6 Kinases, 90-kDa; D019950:Mitogen-Activated Protein Kinase 1; D048052:Mitogen-Activated Protein Kinase 3; D054645:Protein Phosphatase 1; D054648:Protein Phosphatase 2; D054706:Cyclic Nucleotide Phosphodiesterases, Type 5; C517384:Pde5a protein, rat