Renal basolateral membrane SO4/HCO3 transporter characterized by fluorescent acridine orange.
Acridine orange (AO), a pH-sensitive fluorescent indicator, was used to study the characteristics of SO4/HCO3 transport in basolateral membrane vesicles (BLMV) isolated from rabbit renal cortex. The BLMV preparation containing a low buffer concentration and preloaded with 25 mM HCO3 was mixed with buffer containing AO and SO4 in the absence of an initial pH gradient. SO4 influx tended to drive HCO3 efflux, causing intravesicular accumulation of AO and fluorescence quenching. There was no AO quenching in the absence of HCO3 with or without an external SO4 gradient. 100 microM 4,4'-dibenzamido-2,2'-disulfonic stilbene (DBDS) inhibited the fluorescence quenching completely. 25 and 100 mM external Cl did not cause AO quenching. There was no effect of pH (6.5-8.0) on SO4/HCO3 transport. The Kd for SO4 was 8.2 mM. A positive inwardly directed diffusion potential (K(in) = 5 mM, K(out) = 100 mM with valinomycin) did not exert any effect on the SO4/HCO3 transport, indicating that the transport process is insensitive to voltage. The Ki for DBDS inhibition of SO4/HCO3 transport was 2.3 microM.
Chen
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PY
Department of Internal Medicine, Taipei Medical College Hospital, Taiwan, R.O.C.
Tsai
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MC
D000165:Acridine Orange; D000818:Animals; D001639:Bicarbonates / Q000493:pharmacokinetics*; D001692:Biological Transport; D005260:Female; D005453:Fluorescence; D066298:In Vitro Techniques; D007672:Kidney Cortex / Q000378:metabolism*; D008566:Membranes / Q000378:metabolism; D011817:Rabbits; D013431:Sulfates / Q000493:pharmacokinetics*
D001639:Bicarbonates; D013431:Sulfates; D000165:Acridine Orange