Antibodies against the cardiac sodium/bicarbonate co-transporter (NBCe1) as pharmacological tools

Abstract

Background and purpose: Na⁺/HCO₃⁻ co-transport (NBC) regulates intracellular pH (pH_i) in the heart. We have studied the electrogenic NBC isoform NBCe1 by examining the effect of functional antibodies to this protein. Experimental approach: We generated two antibodies against putative extracellular loop domains 3 (a-L3) and 4 (a-L4) of NBCe1 which recognized NBCe1 on immunoblots and immunostaining experiments. pH_i was monitored using epi-fluorescence measurements in cat ventricular myocytes. Transport activity of total NBC and of NBCe1 in isolation were evaluated after an ammonium ion-induced acidosis (expressed as H⁺ flux, J_H, in mmol·L^-1 min^-1 at pH_i 6.8) and during membrane depolarization with high extracellular potassium (potassium pulse, expressed as ΔpH_i) respectively. Key results: The potassium pulse produced a pH_i increase of 0.18 ± 0.006 (n= 5), which was reduced by the a-L3 antibody (0.016 ± 0.019). The a-L-3 also decreased J_H by 50%. Surprisingly, during the potassium pulse, a-L4 induced a higher pH_i increase than control,(0.25 ± 0.018) whereas the recovery of pH_i from acidosis was faster (J_H was almost double the control value). In perforated-patch experiments, a-L3 prolonged and a-L4 shortened action potential duration, consistent with blockade and stimulation of NBCe1-carried anionic current respectively. Conclusions and implications: Both antibodies recognized NBCe1, but they had opposing effects on the function of this transporter, as the a-L3 was inhibitory and the a-L4 was excitatory. These antibodies could be valuable in studies on the pathophysiology of NBCe1 in cardiac tissue, opening a path for their potential clinical use.