Involvement of AE3 isoform of Na+-independent Cl-/HCO-3 exchanger in myocardial pHi recovery from intracellular alkalization

Myocardial pH(i) recovery from intracellular alkalization results in part from the acid load (-J(H+)) carried by Cl-/HCO3- anion-exchangers (AE). Three AE isoforms, AE1, AE2 and AE3, have been identified in cardiac membranes, but the function of each isoform on pH(i) homeostasis is still under investigation. This work explored, by means of specific antibodies, the role of AE3 isoform in myocardial pH(i) regulation. We developed rabbit polyclonal antibodies against the extracellular "loops": one connecting the fifth to sixth and the other one the seventh to eighth transmembrane domains (loops 3 and 4, respectively) of AE3, and their effect on pH(i) regulation was studied in rat papillary muscles. The anti-AE3 loop 3 antibody decreased -J(H+) in response to myocardial alkalization (from a mean control value of 1.06 +/- 0.26 to 0.32 +/- 0.13 mmol/L/min, n = 7, P < 0.05) without affecting the baseline pH(i) (7.22 +/- 0.03 vs. 7.21 +/- 0.04). The anti-AE3 loop 4 antibody did not modify either pH(i) recovery or baseline pH(i). Under control conditions, endothelin-1 (ET-1) increased -J(H+) in response to myocardial alkalization from 1.30 +/- 0.18 to 2.01 +/- 0.33 mmol/L/min (n=5, P < 0.05). This effect of ET-1 on -J(H+), was abolished by anti-AE3 loop 3 antibody. In addition, the MgATP-induced stimulation of AE activity was reduced by the anti-AE3 loop 3 antibody. These data support the key role of the AE3 isoform in myocardial pH(i) recovery from alkaline loads and also in the stimulatory effect of ET-1 on AE activity. To a lesser extent, it may also contribute to the effect of MgATP on pH(i). (c) 2006 Elsevier Inc. All rights reserved.