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Am J Physiol Cell Physiol 278: C1200-C1211, 2000;
0363-6143/00 $5.00
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Vol. 278, Issue 6, C1200-C1211, June 2000

An electrogenic Na+-HCOminus 3 cotransporter (NBC) with a novel COOH-terminus, cloned from rat brain

Mark O. Bevensee, Bernhard M. Schmitt, Inyeong Choi, Michael F. Romero, and Walter F. Boron

Department of Cellular and Molecular Physiology, Yale University School of Medicine, New Haven, Connecticut 06520

We screened rat brain cDNA libraries and used 5' rapid amplification of cDNA ends to clone two electrogenic Na+-HCO-3 cotransporter (NBC) isoforms from rat brain (rb1NBC and rb2NBC). At the amino acid level, one clone (rb1NBC) is 96% identical to human pancreas NBC. The other clone (rb2NBC) is identical to rb1NBC except for 61 unique COOH-terminal amino acids, the result of a 97-bp deletion near the 3' end of the open-reading frame. Using RT-PCR, we confirmed that mRNA from rat brain contains this 97-bp deletion. Furthermore, we generated rabbit polyclonal antibodies that distinguish between the unique COOH-termini of rb1NBC (alpha rb1NBC) and rb2NBC (alpha rb2NBC). alpha rb1NBC labels an ~130-kDa protein predominantly from kidney, and alpha rb2NBC labels an ~130-kDa protein predominantly from brain. alpha rb2NBC labels a protein that is more highly expressed in cortical neurons than astrocytes cultured from rat brain; alpha rb1NBC exhibits the opposite pattern. In expression studies, applying 1.5% CO2/10 mM HCO-3 to Xenopus oocytes injected with rb2NBC cRNA causes 1) pHi to recover from the initial CO2-induced acidification and 2) the cell to hyperpolarize. Subsequently, removing external Na+ reverses the pHi increase and elicits a rapid depolarization. In the presence of 450 µM DIDS, removing external Na+ has no effect on pHi and elicits a small hyperpolarization. The rate of the pHi decrease elicited by removing Na+ is insensitive to removing external Cl-. Thus rb2NBC is a DIDS-sensitive, electrogenic NBC that is predominantly expressed in brain of at least rat.

intracellular pH; pH regulation; alternative splice variant; bicarbonate; sodium


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