Journal
PROCEEDINGS OF THE NATIONAL ACADEMY OF SCIENCES OF THE UNITED STATES OF AMERICA
Volume 98, Issue 26, Pages 15354-15359Publisher
NATL ACAD SCIENCES
DOI: 10.1073/pnas.261253998
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Funding
- NIAID NIH HHS [R01 AI030464, R01 AI45098, R01 AI30464, R56 AI030464, R01 AI045098, R01 AI22444] Funding Source: Medline
- NIMH NIH HHS [R01 MH60261] Funding Source: Medline
- NINDS NIH HHS [R01 NS39103] Funding Source: Medline
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The midgut of mosquito larvae maintains a specific lumen alkalinization profile with large longitudinal gradients (pH approximate to 3 units.mm(-1)) in which an extremely alkaline (pH approximate to 11) anterior midgut lies between near-neutral posterior midgut and gastric cecum (pH 7-8). A plasma membrane HI V-ATPase energizes this alkalinization but the ion carriers involved are unknown. Capillary zone electrophoresis of body samples with outlet conductivity detection showed a specific transepithelial distribution of chloride and bicarbonate/carbonate ions, with high concentrations of both anions in the midgut tissue: 68.3 +/- 5.64 and 50.8 +/- 4.21 mM, respectively. Chloride was higher in the hemolymph, 57.6 +/- 7.84, than in the lumen, 3.51 +/- 2.58, whereas bicarbonate was higher in the lumen, 58.1 +/- 7.34, than the hemolymph, 3.96 +/- 2.89. Time-lapse video assays of pH profiles in vivo revealed that ingestion of the carbonic anhydrase inhibitor acetazolamide and the ion exchange inhibitor DIDS (4,4'-diisothiocyanatostilbene-2,2'-disulfonic acid), at 10(-4) M eliminates lumen alkalinization. Basal application of these inhibitors in situ also reduced gradients recorded with self-referencing pH-sensitive microelectrodes near the basal membrane by approximate to 65% and 85% respectively. Self-referencing chloride-selective microelectrodes revealed a specific spatial profile of transepithelial chloride transport with an efflux maximum in anterior midgut. Both acetazolamide and DIDS reduced chloride effluxes. These data suggest that an H+ V-ATPase-energized anion exchange occurs across the apical membrane of the epithelial cells and implicate an electrophoretic Cl-/HCO3- exchanger and carbonic anhydrase as crucial components of the steady-state alkalinization in anterior midgut of mosquito larvae.
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