Characterization of basolateral Na/H exchange (Na/H-1) in MDCK cells

Sebastiano Vilella1, Lorenzo Guerra1, Corinna Helmle Kolb1, Heini Murer1
1Department of Physiology, University of Zürich, Zürich, Switzerland

Tóm tắt

MDCK cells were grown to confluent monolayers on permeant filter supports; pH was analysed by using the pH-sensitive fluorescent probe 2′7′-biscarboxyethyl-5,6-carboxyfluorescein and a routine spectrofluorometer equipped with a perfusion cuvette [Krayer-Pawlowska et al. (1990) J Membr Biol 120:173–183]. Superfusion of the basolateral (but not apical) cell surface with Na+-containing solutions led to immediate recovery of pHi from an acid load (NH4 prepulse). This pHi recovery was reversibly inhibited by ethylisopropylamiloride indicating Na/H exchange activity. Na/H exchange activity showed an apparent K m for Na+ of about 25 mM Na+ and an apparent K i for inhibition by dimethylamiloride of around 0.2 μM; inhibition by dimethylamiloride was competitive with Na+ interaction. Lowering pHi prior to analysis of Na/H exchange leads to sharp activation of Na/H exchange; the apparent V max for Na/H exchange is increased more than tenfold by lowering the pHi from 7.0 to 6.7 without an effect on apparent K m values for Na+ interaction. It is concluded that MDCK cells (strain I) grown on a permeant support contain only basolateral Na/H exchange activity, most likely Na/H-1 [for nomenclature see Igarashi et al. (1991) Kidney Int 40:S84–S89].

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Tài liệu tham khảo

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