The Electrogenic Sodium Bicarbonate Cotransporter NBCe1 as a Conduit for CO2

Abstract

The established role of NBCe1‐A (SLC4A4) is to cotransport Na+ and HCO3− (or more likely, CO3=). We present evidence that NBCe1‐A also conducts CO2. We record the ionic current carried by NBCe1‐A (INBC)—a direct measure of cotransport activity, while simultaneously assessing the CO3= and CO2 traffic through NBCe1‐A from maximal rates of intracellular pH (pHi) changes, (dpHi/dt)max. We employ a continuous‐flow, rapid‐mixing approach for generating out‐of‐equilibrium (OOE) CO2/HCO3− solutions to control [CO2], [HCO3−], and extracellular pH (pHo) individually in the bath. Exposing Xenopus oocytes expressing NBCe1‐A to a pure‐HCO3− (0% CO2/33 mM HCO3−) OOE solution at pHo 7.5 causes pHi to rise +(dpHi/dt)max, moderately fast due to electrogenic Na+/CO3= uptake. Exposing NBCe1‐A expressing oocytes to a pure‐CO2 (5% CO2/0 mM HCO3−) OOE solution causes pHi to fall, −(dpHi/dt)max, rapidly as CO2 enters. Computer simulations predict that in a pHo 7.5‐equilibrated 5% CO2/33 mM HCO3− solution (EQ), the pHi in NBCe1‐A oocytes should fall at a rate approximately halfway between that of the pure‐HCO3− and pure‐CO2 OOE conditions. However, experiments show that the pHi of EQ‐exposed NBCe1‐A oocytes falls more rapidly than predicted in the simulations. Importantly, the INBC associated with the 5% CO2/33 mM HCO3− condition at pHo 7.5 is virtually identical to the INBC during the pure‐HCO3− condition. To increase [CO3=]o by ~30‐fold and accelerate NBCe1‐A, we raised pHo to 9.0. In pure‐HCO3−/pHo 9.0, pHi rises twice as fast as recorded at pHo 7.5. In pure‐CO2/pHo 9.0, pHi falls, though not as rapidly as it rises in pure‐HCO3−/pHo 9.0. Thus, we expected pHi to rise in a 5% CO2/33 mM HCO3−/pHo 9.0 OOE solution (i.e., follow midpoint between pure‐HCO3− and pure‐CO2). However, our data show that pHi falls with a magnitude almost identical to that for pure‐CO2/pHo 9.0, even though the magnitude of the concurrent INBC in both pure‐HCO3− and 5% CO2/33 mM HCO3−/pHo 9.0 is twice those at pHo 7.5. In contrast, experiments performed on oocytes expressing AE1 show that (dpHi/dt)max values in the EQ solution approximately equal the sum of (dpHi/dt)max rates in pure‐HCO3− and pure‐CO2 OOE conditions. A trivial explanation for our data would be carbonic anhydrase (CA) activity caused by expression of NBCe1‐A. However, a stopped‐flow assay for CA activity developed in our Lab shows that this is not the case. We propose that a simple explanation for our data is that when NBCe1‐A is transporting Na+ + CO3=, it also acts as a new class of CO2 channel—the first that does not have a permanent pore. We speculate that conformational changes integral to Na+ + CO3= transport, create transient pathways within the NBCe1 molecule that allow CO2 to pass from the extra‐ to the intracellular fluid.Support or Funding InformationOffice of Naval Research (N00014‐15‐1‐2060 & N00014‐16‐1‐2535), NIH (R01‐DK113197, U01‐GM111251, K01‐DK107787)This abstract is from the Experimental Biology 2019 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.