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

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 (αrb1NBC) and rb2NBC (αrb2NBC). αrb1NBC labels an ∼130-kDa protein predominantly from kidney, and αrb2NBC labels an ∼130-kDa protein predominantly from brain. αrb2NBC labels a protein that is more highly expressed in cortical neurons than astrocytes cultured from rat brain; αrb1NBC exhibits the opposite pattern. In expression studies, applying 1.5% CO₂/10 mM ${HCO}_3^{-}$ toXenopus oocytes injected with rb2NBC cRNA causes 1) pH_i to recover from the initial CO₂-induced acidification and 2) the cell to hyperpolarize. Subsequently, removing external Na⁺ reverses the pH_i increase and elicits a rapid depolarization. In the presence of 450 μM DIDS, removing external Na⁺ has no effect on pH_i and elicits a small hyperpolarization. The rate of the pH_idecrease 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.