Control of cytosolic pH in two-cell mouse embryos: roles of H(+)-lactate cotransport and Na+/H+ exchange.
In this study we used imaging techniques with the fluorescent pH-sensitive dye 2',7'-bis(carboxyethyl)-5(6)-carboxyfluorescein to investigate the control of cytosolic pH (pHi) in two-cell mouse embryos in nominally HCO3(-)-free conditions. We found that the resting pHi of two-cell embryos (40-50 h after human chorionic gonadotropin) in HCO3(-)-free M2 was 7.31 +/- 0.01 (n = 172 embryos), which is significantly above the level predicted if H+ is at electrochemical equilibrium. We showed that two-cell embryos contain a H(+)-monocarboxylate cotransport system with apparent Michaelis constants for D-lactate, L-lactate, and pyruvate of 11.5, 3.7, and 3.5 mM, respectively. It is inhibited by p-chloromercuribenzoic acid (300 microM), p-chloromercuriphenylsulfonic acid (300 microM), and alpha-cyano-4-hydroxycinnamate (1 mM) and is insensitive to 4,4'-diisothiocyanodihydrostilbene-2,2'-disulfonic acid (500 microM). We also showed that the pHi response to the acid load produced by an NH4Cl pulse has two components, one due to H(+)-monocarboxylate cotransport and the other due to Na+/H+ exchange. We found no evidence that a H+ conductance was responsible in these cells for the recovery in pHi after an acid load.
Gibb
C A
CA
Department of Physiology, University of Sydney, New South Wales, Australia.
Poronnik
P
P
Day
M L
ML
Cook
D I
DI
D000643:Ammonium Chloride / Q000494:pharmacology; D000818:Animals; D001692:Biological Transport / Q000502:physiology*; D003600:Cytosol / Q000187:drug effects / Q000378:metabolism*; D004622:Embryo, Mammalian / Q000166:cytology / Q000378:metabolism*; D005260:Female; D006859:Hydrogen / Q000378:metabolism*; D006863:Hydrogen-Ion Concentration / Q000187:drug effects; D051379:Mice / Q000196:embryology; D008815:Mice, Inbred Strains; D012964:Sodium / Q000494:pharmacology; D017923:Sodium-Hydrogen Exchangers / Q000378:metabolism*
D017923:Sodium-Hydrogen Exchangers; D000643:Ammonium Chloride; D006859:Hydrogen; D012964:Sodium