title: Localization and role of inward rectifier K(+) channels in Malpighian tubules of the yellow fever mosquito Aedes aegypti.
abstract: Malpighian tubules of adult female yellow fever mosquitoes Aedes aegypti express three inward rectifier K(+) (Kir) channel subunits: AeKir1, AeKir2B and AeKir3. Here we 1) elucidate the cellular and membrane localization of these three channels in the Malpighian tubules, and 2) characterize the effects of small molecule inhibitors of AeKir1 and AeKir2B channels (VU compounds) on the transepithelial secretion of fluid and electrolytes and the electrophysiology of isolated Malpighian tubules. Using subunit-specific antibodies, we found that AeKir1 and AeKir2B localize exclusively to the basolateral membranes of stellate cells and principal cells, respectively; AeKir3 localizes within intracellular compartments of both principal and stellate cells. In isolated tubules bathed in a Ringer solution containing 34 mM K(+), the peritubular application of VU590 (10 μM), a selective inhibitor of AeKir1, inhibited transepithelial fluid secretion 120 min later. The inhibition brings rates of transepithelial KCl and fluid secretion to 54% of the control without a change in transepithelial NaCl secretion. VU590 had no effect on the basolateral membrane voltage (Vbl) of principal cells, but it significantly reduced the cell input conductance (gin) to values 63% of the control within ∼90 min. In contrast, the peritubular application of VU625 (10 μM), an inhibitor of both AeKir1 and AeKir2B, started to inhibit transepithelial fluid secretion as early as 60 min later. At 120 min after treatment, VU625 was more efficacious than VU590, inhibiting transepithelial KCl and fluid secretion to ∼35% of the control without a change in transepithelial NaCl secretion. Moreover, VU625 caused the Vbl and gin of principal cells to respectively drop to values 62% and 56% of the control values within only ∼30 min. Comparing the effects of VU590 with those of VU625 allowed us to estimate that AeKir1 and AeKir2B respectively contribute to 46% and 20% of the transepithelial K(+) secretion when the tubules are bathed in a Ringer solution containing 34 mM K(+). Thus, we uncover an important role of AeKir1 and stellate cells in transepithelial K(+) transport under conditions of peritubular K(+) challenge. The physiological role of AeKir3 in intracellular membranes of both stellate and principal cells remains to be determined.
authors.affiliation: Department of Entomology, Ohio Agricultural Research and Development Center, The Ohio State University, Wooster, OH 44691, USA. Electronic address: piermarini.1@osu.edu.
authors.forename: Peter M
authors.initials: PM
authors.lastname: Piermarini
authors.affiliation: Department of Entomology, Ohio Agricultural Research and Development Center, The Ohio State University, Wooster, OH 44691, USA.
authors.forename: Sonja M
authors.initials: SM
authors.lastname: Dunemann
authors.affiliation: Department of Entomology, Ohio Agricultural Research and Development Center, The Ohio State University, Wooster, OH 44691, USA.
authors.forename: Matthew F
authors.initials: MF
authors.lastname: Rouhier
authors.affiliation: Department of Entomology, Ohio Agricultural Research and Development Center, The Ohio State University, Wooster, OH 44691, USA.
authors.forename: Travis L
authors.initials: TL
authors.lastname: Calkins
authors.affiliation: Department of Anesthesiology, Vanderbilt University School of Medicine, Nashville, TN 37232, USA; Department of Pharmacology, Vanderbilt University School of Medicine, Nashville, TN 37232, USA.
authors.forename: Rene
authors.initials: R
authors.lastname: Raphemot
authors.affiliation: Department of Anesthesiology, Vanderbilt University School of Medicine, Nashville, TN 37232, USA; Department of Pharmacology, Vanderbilt University School of Medicine, Nashville, TN 37232, USA.
authors.forename: Jerod S
authors.initials: JS
authors.lastname: Denton
authors.affiliation: Department of Biomedical Sciences, VRT 8004, Cornell University, Ithaca, NY 14853, USA.
authors.forename: Rebecca M
authors.initials: RM
authors.lastname: Hine
authors.affiliation: Department of Biomedical Sciences, VRT 8004, Cornell University, Ithaca, NY 14853, USA.
authors.forename: Klaus W
authors.initials: KW
authors.lastname: Beyenbach
mesh_terms: D000330:Aedes / Q000187:drug effects / Q000378:metabolism*; D000818:Animals; D001692:Biological Transport / Q000187:drug effects; D005260:Female; D006573:Heterocyclic Compounds, 1-Ring / Q000494:pharmacology; D008317:Malpighian Tubules / Q000187:drug effects / Q000378:metabolism*; D008564:Membrane Potentials; D011188:Potassium / Q000378:metabolism*; D024661:Potassium Channels, Inwardly Rectifying / Q000187:drug effects / Q000378:metabolism*; D011189:Potassium Chloride / Q000378:metabolism; D012965:Sodium Chloride / Q000378:metabolism
chemical_list: C544855:7,13-bis(4-nitrobenzyl)-1,4,10-trioxa-7,13-diazacyclopentadecane; D006573:Heterocyclic Compounds, 1-Ring; D024661:Potassium Channels, Inwardly Rectifying; D012965:Sodium Chloride; D011189:Potassium Chloride; D011188:Potassium
keywords: Electrophysiology; Immunolocalization; Pharmacology; Potassium channels; Small molecules; Transepithelial fluid secretion