title: Detection of different adenosine triphosphatases in human placental brush border membranes.
abstract: The microvillous membrane of human placental syncytiotrophoblast cells contains a high ATPase activity. The purpose of this study was to characterize this activity and to investigate the presence of vacuolar type H+ ATPase in this membrane. Intact brush border membrane vesicles strongly hydrolyzed ATP, reflecting the presence of ATPase on the external side of the membrane. The ATPase activity was entirely Mg2+ dependent and increased with pH. At pH 7.5, Vmax was 31.0 +/- 1.7 mumol/mg/20 min and Km 0.18 +/- 0.03 mM ATP. Hydrolysis of ATP was not influenced by the presence of bicarbonate or alkaline phosphatase inhibitors, but at pH 8 it decreased by half following addition of 100 microM dicyclohexylcarbodiimide (DCCD). At pH 7.5, 1 mM N-ethylmaleimide (NEM) depressed this activity by less than 5%. Opening the membrane vesicles with 0.1% desoxycholate (DOC) or Triton-X neither revealed any additional ATPase activity nor altered the low sensitivity to NEM. Treatment of these membranes with 1% cholate decreased the ATPase activity by more than 70% and did not enhance the sensitivity of ATP hydrolysis to NEM. 10(-7) M Bafilomycin, which reduced by 56 +/- 9% the ATPase activity in dog kidney brush border membranes treated with 0.1% DOC, had no effect on placental brush border membranes subjected to the same procedure. Finally, neither immunocytochemical staining using monoclonal antibody to the M(r) 31000 subunit of V-type H+ ATPase, nor electron microscopic examination detected the presence of H(+) ATPase in placental membranes. In conclusion, the placental brush border membrane is the site of a strong "ecto" ATPase activity which is partially DCCD sensitive.(ABSTRACT TRUNCATED AT 250 WORDS)
authors.affiliation: Maisonneuve-Rosemont Hospital, Research Center, Montreal, Canada.
authors.forename: M G
authors.initials: MG
authors.lastname: Brunette
authors.forename: B
authors.initials: B
authors.lastname: Bastani
authors.forename: M
authors.initials: M
authors.lastname: Leclerc
authors.forename: R
authors.initials: R
authors.lastname: Narbaitz
mesh_terms: D000251:Adenosine Triphosphatases / Q000145:classification / Q000302:isolation & purification*; D000255:Adenosine Triphosphate / Q000378:metabolism; D000469:Alkaline Phosphatase / Q000378:metabolism; D000818:Animals; D000900:Anti-Bacterial Agents / Q000494:pharmacology; D019826:Cholic Acid; D002793:Cholic Acids / Q000494:pharmacology; D003840:Deoxycholic Acid / Q000494:pharmacology; D004024:Dicyclohexylcarbodiimide / Q000494:pharmacology; D004285:Dogs; D005033:Ethylmaleimide / Q000494:pharmacology; D005260:Female; D006801:Humans; D006863:Hydrogen-Ion Concentration; D006868:Hydrolysis; D066298:In Vitro Techniques; D007700:Kinetics; D018942:Macrolides; D008565:Membrane Proteins / Q000378:metabolism; D008871:Microvilli / Q000201:enzymology*; D010920:Placenta / Q000201:enzymology*; D011247:Pregnancy; D006180:Proton-Translocating ATPases / Q000302:isolation & purification; D051381:Rats; D014638:Vanadates / Q000494:pharmacology
chemical_list: D000900:Anti-Bacterial Agents; D002793:Cholic Acids; D018942:Macrolides; D008565:Membrane Proteins; D003840:Deoxycholic Acid; D014638:Vanadates; D004024:Dicyclohexylcarbodiimide; D000255:Adenosine Triphosphate; D000469:Alkaline Phosphatase; D000251:Adenosine Triphosphatases; D006180:Proton-Translocating ATPases; D019826:Cholic Acid; D005033:Ethylmaleimide