V-type H+-ATPase in the human eccrine sweat duct: immunolocalization and functional demonstration.
We investigated for the presence of a vacuolar-type H+-ATPase (V-ATPase) in the human eccrine sweat duct (SD). With the use of immunocytochemistry, an anti-V- ATPase antibody showed a strong staining at the apical membrane and a weaker one in the cytoplasm. Cold preservation followed by rewarming did not alter this staining pattern. With the use of the pH-sensitive dye 2',7'-bis(2-carboxyethyl)-5(6)-carboxyfluorescein on isolated and perfused straight SD under HCO-free conditions and in the absence of Na+, proton extrusion was determined from the recovery rate of intracellular pH (dpH(i)/dt) following an acid load. Oligomycin (25 microM), an inhibitor of F-type ATPases, decreased dpH(i)/dt by 88 +/- 6%, suggesting a role for an ATP-dependent process involved in pH(i) recovery. Moreover, dpH(i)/dt was inhibited at 95 +/- 3% by 100 nM luminal concanamycin A, a specific inhibitor of V-ATPases, whereas 10 microM bafilomycin A1, another specific inhibitor of V-ATPases, was required to decrease dpH(i)/dt by 73%. These results strongly suggest that a V-ATPase is involved in proton secretion in the human eccrine SD.
Granger
D
D
Groupe de Recherche en Transport Membranaire, Université de Montréal, Montreal, Quebec, Canada H3C 3J7.
Marsolais
M
M
Burry
J
J
Laprade
R
R
D000143:Acids / Q000494:pharmacology; D000900:Anti-Bacterial Agents / Q000494:pharmacology; D001692:Biological Transport / Q000187:drug effects / Q000502:physiology; D004439:Eccrine Glands / Q000166:cytology / Q000201:enzymology*; D004791:Enzyme Inhibitors / Q000494:pharmacology; D005260:Female; D006801:Humans; D006863:Hydrogen-Ion Concentration / Q000187:drug effects; D007150:Immunohistochemistry; D066298:In Vitro Techniques; D007424:Intracellular Fluid / Q000187:drug effects / Q000378:metabolism; D018942:Macrolides; D009840:Oligomycins / Q000494:pharmacology; D010477:Perfusion; D017494:Proton Pumps / Q000187:drug effects / Q000378:metabolism; D013696:Temperature; D025262:Vacuolar Proton-Translocating ATPases / Q000037:antagonists & inhibitors / Q000378:metabolism*
D000143:Acids; D000900:Anti-Bacterial Agents; D004791:Enzyme Inhibitors; D018942:Macrolides; D009840:Oligomycins; D017494:Proton Pumps; C036978:concanamycin A; C040929:bafilomycin A1; D025262:Vacuolar Proton-Translocating ATPases