title: Intracellular pH in mammalian stages of Trypanosoma cruzi is K+-dependent and regulated by H+-ATPases.
abstract: Regulation of intracellular pH (pHi) was investigated in Trypanosoma cruzi amastigotes and trypomastigotes using 2',7'-bis-(carboxyethyl)-5(and-6)-carboxyfluorescein (BCECF). pHi was determined to be 7.33 +/- 0.08 and 7.35 +/- 0.07 in amastigotes and trypomastigotes, respectively, and there were no significant differences in the regulation of pH, between the two stages. Steady-state pHi, recovery of pHi from acidification, and H+-efflux were all decreased markedly by the H+-ATPase inhibitors N,N'-dicyclohexylcarbodi-imide (DCCD), diethylstilbestrol (DES) and N-ethylmaleimide (NEM) supporting a significant role for a plasma membrane H+-ATPase in the regulation of pHi. pHi was maintained at neutrality over a range of external pH (pHe) from 5-8 in parasites suspended in a buffer containing Na+ and K+ (standard buffer) but was acidified at low pHe in the absence of these cations (choline buffer). The pHi of trypomastigotes decreased significantly when they transformed into amastigotes. The rate of recovery of pHi by acidified parasites was similar in Na+-free buffer and standard buffer but was slower in the absence of K+ (K+-free or choline buffer) and parasites suspended in choline buffer were acidic by 0.25 pH units as compared with controls. Ba2+ and Cs+ decreased the pHi of parasites suspended in standard but not choline buffer suggesting the presence of an inward directed K+ channel. The pHi of amastigotes and trypomastigotes suspended in Cl(-)-free buffer was decreased by 0.13 and 0.2 pH units, respectively, supporting the presence of a chloride conductive channel. No evidence of pH regulation via a Na+/H+ or Cl-/HCO3- exchanger was found. These results are consistent with the presence of a plasma membrane H+-ATPase that regulates pHi and is supported by K+ and Cl- channels.
authors.affiliation: Department of Pathobiology, College of Veterinary Medicine, University of Illinois at Urbana-Champaign, Urbana 61802, USA.
authors.forename: N
authors.initials: N
authors.lastname: Van Der Heyden
authors.forename: R
authors.initials: R
authors.lastname: Docampo
mesh_terms: D017878:4,4'-Diisothiocyanostilbene-2,2'-Disulfonic Acid / Q000031:analogs & derivatives / Q000494:pharmacology; D000255:Adenosine Triphosphate / Q000378:metabolism; D000584:Amiloride / Q000031:analogs & derivatives / Q000494:pharmacology; D000818:Animals; D001639:Bicarbonates / Q000737:chemistry / Q000494:pharmacology; D002021:Buffers; D002712:Chlorides / Q000737:chemistry / Q000494:pharmacology; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005786:Gene Expression Regulation; D017506:H(+)-K(+)-Exchanging ATPase / Q000378:metabolism*; D006706:Homeostasis; D006863:Hydrogen-Ion Concentration; D011188:Potassium / Q000378:metabolism*; D054328:Proton Pump Inhibitors; D011522:Protons; D012964:Sodium / Q000737:chemistry; D014349:Trypanosoma cruzi / Q000201:enzymology / Q000254:growth & development* / Q000378:metabolism*
chemical_list: D001639:Bicarbonates; D002021:Buffers; D002712:Chlorides; D004791:Enzyme Inhibitors; D054328:Proton Pump Inhibitors; D011522:Protons; C018564:dihydro-DIDS; D000584:Amiloride; D000255:Adenosine Triphosphate; D012964:Sodium; D017506:H(+)-K(+)-Exchanging ATPase; D017878:4,4'-Diisothiocyanostilbene-2,2'-Disulfonic Acid; D011188:Potassium; C039614:ethylisopropylamiloride