Azide- and vanadate-sensitive M-phase alkalinity and cytosolic acidification of Chang liver cells.
Flow cytometric cell-by-cell evaluation of NH4Cl acidification of human Chang cells showed that at steady state, 3% of the cells remained alkalinized (> pHi 7) over an extended period (up to 80 min) despite the absence of extracellular Na+ and HCO3-. In fluorescence microscopy, the acidification-resistant cells were characteristically rounded M-phase cells. Both mean cytosolic pH and M-phase alkalinity were however sensitive to (a) azide and oligomycin, inhibitors of F-ATPase (ATP synthase), and to (b) vanadium ions, the phosphate analogue of P-ATPase (ATP-hydrolyzing), in dose-dependent and time-dependent manners. Dead cell indices were constant at approximately 10%. Thiocyanate chaotrophic anions, which cleave the V-ATPase structure, had no effect. Since ATP synthesizing F-ATPase (ATP synthase) is coupled to ATP-hydrolyzing P-ATPase as 'master-&-slave', azide- and oligomycin-sensitivity corroborated with vanadate-sensitivity in suggesting energized proton pumping modulating (a) M-phase alkalinity and (b) cytosolic pH, against acidification.
Sit
K H
KH
Department of Anatomy, Faculty of Medicine, National University of Singapore, Kent Ridge.
Paramanantham
R
R
Bay
B H
BH
Wong
K P
KP
D000251:Adenosine Triphosphatases / Q000037:antagonists & inhibitors; D001386:Azides / Q000494:pharmacology*; D002478:Cells, Cultured; D003600:Cytosol / Q000187:drug effects / Q000201:enzymology / Q000378:metabolism*; D005434:Flow Cytometry; D006801:Humans; D006863:Hydrogen-Ion Concentration; D008099:Liver / Q000166:cytology / Q000201:enzymology / Q000378:metabolism*; D008938:Mitosis / Q000502:physiology*; D014638:Vanadates / Q000494:pharmacology*
D001386:Azides; D014638:Vanadates; D000251:Adenosine Triphosphatases