Active ammonia absorption in the midgut of the Tobacco hornworm Manduca sexta L.: transport studies and mRNA expression analysis of a Rhesus-like ammonia transporter.
In this study the mid- and hindgut of Manduca sexta larvae were tested for their ammonia transport properties using a custom-made Ussing chamber. In the presence of 0.1 mmoll(-1) ammonia on both sides of the isolated epithelium, active transepithelial ammonia absorption (aTEPA) was observed in all midgut sections, with greatest transport rates (ca. 140 nmol cm(-2)h(-1)) detected in the median midgut. The hindgut showed no aTEPA. In the median midgut inhibition of energy metabolism by azide blocked aTEPA completely, whereas inhibition of vacuolar H(+)-ATPase by bafilomycin A(1) reduced the active transport by 50%. The imposition of a luminal-directed NH(3)-gradient (pH 6.5 apical, pH 8.5 basal) lowered the aTEPA by approximately 50% but did not reverse its direction. Apical addition of amiloride reduced aTEPA by 90%, suggesting a role of carrier-mediated ammonia transport across the apical membrane via a member of the NHE family. Inhibition of the microtubule network by colchicine reduced aTEPA by ca. 50%. In contrast, blocking basal K(+) channels by Ba(2+) had no effect on aTEPA. Using molecular methods, evidence for intestinal expression of a Rhesus-like ammonia transporter (RhMS) was found with low mRNA expression in midgut tissues, but high expression levels in the hindgut, Malpighian tublules and ganglia.
Weihrauch
Dirk
D
Department of Biology/Chemistry, Division of Animal Physiology, University of Osnabrück, 49076 Osnabrück, Germany. weihrauchblues@gmx.net <weihrauchblues@gmx.net>
D000584:Amiloride / Q000494:pharmacology; D000641:Ammonia / Q000097:blood / Q000378:metabolism*; D000818:Animals; D017609:Barium Compounds / Q000494:pharmacology; D001692:Biological Transport; D003078:Colchicine / Q000494:pharmacology; D017952:Ganglia, Invertebrate / Q000187:drug effects / Q000378:metabolism; D006458:Hemolymph / Q000378:metabolism; D019476:Insect Proteins / Q000235:genetics / Q000378:metabolism*; D007413:Intestinal Mucosa / Q000378:metabolism; D008317:Malpighian Tubules / Q000187:drug effects / Q000378:metabolism; D018402:Manduca / Q000378:metabolism*; D026901:Membrane Transport Proteins / Q000235:genetics / Q000378:metabolism*; D026902:Potassium Channel Blockers / Q000494:pharmacology; D012333:RNA, Messenger / Q000378:metabolism; D026941:Sodium Channel Blockers / Q000494:pharmacology; D050257:Tubulin Modulators / Q000494:pharmacology
D017609:Barium Compounds; D019476:Insect Proteins; D026901:Membrane Transport Proteins; D026902:Potassium Channel Blockers; D012333:RNA, Messenger; D026941:Sodium Channel Blockers; D050257:Tubulin Modulators; D000641:Ammonia; D000584:Amiloride; D003078:Colchicine