A SLC4-like anion exchanger from renal tubules of the mosquito (Aedes aegypti): evidence for a novel role of stellate cells in diuretic fluid secretion

Abstract

WITH THEIR ENRICHED EXPRESSION and activity of an apical vacuolar (V-type) H ϩ -ATPase (2, 52, 86), the principal cells of renal (Malpighian) tubules in the mosquito (Aedes aegypti) are similar to acid-secreting cells of the mammalian renal collecting tubule (85), mammalian male reproductive tract (10), turtle and amphibian urinary bladders (9), amphibian skin (17), and mammalian bone (76). In these acid-secreting cells, the apical V-type H ϩ -ATPase is often opposed by a basolateral Cl/HCO 3 anion exchanger of the solute-linked carrier 4 (SLC4) superfamily of bicarbonate transporters. The anion exchanger mediates the basolateral extrusion of intracellular HCO 3 Ϫ , thereby removing the intracellular HCO 3 Ϫ that was generated during the ionization of carbonic acid (H 2 CO 3 ).Disulfonic stilbene derivates, such as SITS and DIDS, are potent inhibitors of almost all SLC4 transporters (61). These negatively charged compounds are thought to interact with positively charged lysine residues on the external surface of SLC4 proteins (47, 83), perhaps blocking a conduit for anion translocation. Pharmacological studies using stilbene derivatives suggest the presence of Cl/HCO 3 exchangers in Malpighian tubules of some insects. For example, the addition of SITS to the peritubular bath of isolated Aedes Malpighian tubules lowers the unstimulated rates of fluid secretion (25), and both peritubular SITS and DIDS inhibit the serotoninstimulated rates of fluid secretion in isolated Malpighian tubules of Rhodnius prolixus (23,29). In isolated Malpighian tubules of the cricket (Teleogryllus oceanicus), SITS inhibits unstimulated rates of fluid secretion in the distal and main segments (89); DIDS also inhibits secretion in the distal segment, but it paradoxically stimulates secretion in the main segment (89). Finally, X-ray microanalysis on larval Drosophila Malpighian tubules indicate that DIDS decreases the intracellular Cl Ϫ concentration ([Cl Ϫ ]) in the basal cytoplasm of principal cells, which is consistent with the presence of a Cl/HCO 3 exchanger (88).To date, only one SLC4-like transporter has been cloned and characterized from any insect, i.e., the Na ϩ

Localization and role of inward rectifier K+ channels in Malpighian tubules of the yellow fever mosquito Aedes aegypti

“…Heterologous expression of the AeKir subunits was assessed via Western blotting of total membrane-protein fractions, as described previously (Piermarini et al, 2010. In brief, cleared oocyte lysates were centrifuged at 100,000 g for 60 min at 4 C to isolate the membrane proteins, which were then separated on 12% polyacrylamide gels (10 mg per lane) and transferred to polyvinylidene difluoride (PVDF) membranes (Piermarini et al, 2010).…”

“…In brief, cleared oocyte lysates were centrifuged at 100,000 g for 60 min at 4 C to isolate the membrane proteins, which were then separated on 12% polyacrylamide gels (10 mg per lane) and transferred to polyvinylidene difluoride (PVDF) membranes (Piermarini et al, 2010). The Kir1, Kir2B, and Kir3 antibodies were dissolved in blocking buffer at concentrations of 0.16 mg/ml, 1.74 mg/ml, and 0.345 mg/ml, respectively.…”

“…Crude lysates of Malpighian tubules isolated from~30 adult females (~150 tubules) were prepared as described in previous studies (Piermarini et al, 2010. The samples were boiled for 5 min before separating them on 12% polyacrylamide gels (~50 tubule equivalents per lane).…”

Intracellular Na+, K+ and Cl− activities in Acheta domesticus Malpighian tubules and the response to a diuretic kinin neuropeptide

“…The transcellular pathway may be through principal cells or, in dipteran insects, through stellate cells. The uptake of Cl -from the bathing medium is opposed by the cell-negative basolateral membrane voltage (V b ), and has been attributed to Na + /K + /2Cl -and K + /Cl -cotransporters (Leyssens et al, 1994;Ianowski et al, 2002;Ianowski and O'Donnell, 2004) and a Cl -/HCO 3 -exchanger (Piermarini et al, 2010). The movement of Cl -from cell to lumen is most likely through Cl -channels (Ianowski and O'Donnell, 2006) because this is favoured by the lumen-positive apical membrane voltage (V a ), but an apical K + /Cl -cotransporter contributes to fluid secretion by Malpighian tubules of the yellow fever mosquito, Aedes aegypti (Piermarini et al, 2011).…”

Intracellular pH regulation in unstimulatedCalliphorasalivary glands is Na+ dependent and requires V-ATPase activity

“…The HCO 3 -transporters mentioned here belong to the solute-linked carrier 4 (SLC4)-like transporters. To date, only two SLC4-like transporters from insects have been cloned and identified, the Na + -driven anion exchanger (NDAE) of Drosophila (Romero et al, 2000;Sciortino et al, 2001) and an AE of Aedes aegypti (Piermarini et al, 2010). Drosophila NDAE seems to be analogous in function to the mammalian NDCBE, and to date no orthologs of the mammalian NBCs have been found in insects.…”