CaMKII Contributes to Intracellular pH Recovery from an Acid Load via the Activation of the Na+/H+ Exchanger

MUNDIÑA-WEILENMANN C; VILA PETROFF M; MATTIAZZI A

New London, USA

Conferencia; Gordon Research Conferences. Cardiac Regulatory Mechanism; 2008

Gordon Research Conferences.

Na+/H+ exchanger (NHE) activation plays a key role in pHi recovery from acidosis. This activation has been attributed to two main factors: H+ interaction with an allosteric site and phosphorylation of the exchanger by the ERK1/2 cascade, when acidosis was sustained. Recent evidence has linked the spontaneous mechanical recovery from acidosis with the activation of Ca2+/calmodulin-dependent protein kinase II (CaMKII). However, the impact of this kinase on NHE activity and pHi recovery is not well defined. We tested the hypothesis that CaMKII activates NHE and contributes to pHi recovery from acidosis. For this purpose we subjected adult rat cardiac myocytes, loaded with the pHi indicator SNARF-1/AM, to a sustained intracellular acidosis, generated by NH4Cl pulse, followed by cero Na+ and its washout and to three different protocols of acute acidosis: 1) NH4Cl pulse and its washout; 2) Substitution of HEPES buffer with NaHCO3 buffer equilibrated with 5% CO2-95% O2 (Bic) at the same pHo (7.4) and 3) Hypercapnia at constant pHo induced by switching from a Bic solution to a modified Bic solution equilibrated with 20% CO2-80% O2, and with higher [NaHCO3] to keep pHo constant at 7.4. In all cases, the rate of pHi recovery (dpHi/dt), measured by epifluorescence, was used as an index of NHE activity. Regardless of the approach used, pretreatment with two structurally different CaMKII inhibitors KN-93 (1µM) or AIP (2.5 µM), significantly decreased the rate of pHi recovery from acidosis whereas pretreatment of cells with the inactive analog KN-92 did not affect pHi recovery. The effect of CaMKII was independent of the ERK pathway since inhibition by KN-93 did not abolish the acidosis-induced ERK1/2 phosphorylation. The results demonstrate that CaMKII regulates NHE activity contributing to pHi recovery from acidosis. This ERK1/2 independent mechanism could thus play an important role in the mechanical recovery from acidosis.