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Extracellular Hco3 − Dependence of Electrogenic Na/Hco3 Cotransporters Cloned from Salamander and Rat Kidney

I. I. Grichtchenko,M. Romero,W. Boron

Published 2000 in The Journal of General Physiology

ABSTRACT

We studied the extracellular [HCOabstract 3 −] dependence of two renal clones of the electrogenic Na/HCO3 cotransporter (NBC) heterologously expressed in Xenopus oocytes. We used microelectrodes to measure the change in membrane potential (ΔVm) elicited by the NBC cloned from the kidney of the salamander Ambystoma tigrinum (akNBC) and by the NBC cloned from the kidney of rat (rkNBC). We used a two-electrode voltage clamp to measure the change in current (ΔI) elicited by rkNBC. Briefly exposing an NBC-expressing oocyte to HCOabstract 3 −/CO2 (0.33–99 mM HCOabstract 3 −, pHo 7.5) elicited an immediate, DIDS (4,4-diisothiocyanatostilbene-2,2-disulfonic acid)-sensitive and Na+-dependent hyperpolarization (or outward current). In ΔVm experiments, the apparent K m for HCOabstract 3 − of akNBC (10.6 mM) and rkNBC (10.8 mM) were similar. However, under voltage-clamp conditions, the apparent K m for HCOabstract 3 − of rkNBC was less (6.5 mM). Because it has been reported that SOabstract 3 =/HSO abstract 3 − stimulates Na/HCO3 cotransport in renal membrane vesicles (a result that supports the existence of a COabstract 3 = binding site with which SOabstract 3 = interacts), we examined the effect of SOabstract 3 =/HSO abstract 3 − on rkNBC. In voltage-clamp studies, we found that neither 33 mM SOabstract 4 = nor 33 mM SOabstract 3 =/HSOabstract 3 − substantially affects the apparent K m for HCO abstract 3 −. We also used microelectrodes to monitor intracellular pH (pHi) while exposing rkNBC-expressing oocytes to 3.3 mM HCOabstract 3 −/0.5% CO2. We found that SO abstract 3 =/HSOabstract 3 − did not significantly affect the DIDS-sensitive component of the pHi recovery from the initial CO2 -induced acidification. We also monitored the rkNBC current while simultaneously varying [CO2]o, pHo, and [COabstract 3 =]o at a fixed [HCOabstract 3 −]o of 33 mM. A Michaelis-Menten equation poorly fitted the data expressed as current versus [COabstract 3 =]o . However, a pH titration curve nicely fitted the data expressed as current versus pHo. Thus, rkNBC expressed in Xenopus oocytes does not appear to interact with SOabstract 3 =, HSOabstract 3 −, or COabstract 3 =.

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