Chloride channels in stellate cells are essential for uniquely high secretion rates in neuropeptide-stimulated Drosophila diuresis

Abstract

Epithelia frequently segregate transport processes to specific cell types, presumably for improved efficiency and control. The molecular players underlying this functional specialization are of particular interest. In Drosophila, the renal (Malpighian) tubule displays the highest per-cell transport rates known and has two main secretory cell types, principal and stellate. Electrogenic cation transport is known to reside in the principal cells, whereas stellate cells control the anion conductance, but by an as-yet-undefined route. Here, we resolve this issue by showing that a plasma membrane chloride channel, encoded by ClC-a, is exclusively expressed in the stellate cell and is required for Drosophila kinin-mediated induction of diuresis and chloride shunt conductance, evidenced by chloride ion movement through the stellate cells, leading to depolarization of the transepithelial potential. By contrast, ClC-a knockdown had no impact on resting secretion levels. Knockdown of a second CLC gene showing highly abundant expression in adult Malpighian tubules, ClC-c, did not impact depolarization of transepithelial potential after kinin stimulation. Therefore, the diuretic action of kinin in Drosophila can be explained by an increase in ClC-a-mediated chloride conductance, over and above a resting fluid transport level that relies on other (ClC-a-independent) mechanisms or routes. This key segregation of cation and anion transport could explain the extraordinary fluid transport rates displayed by some epithelia.

Transport proteins NHA1 and NHA2 are essential for survival, but have distinct transport modalities

“…Both Drosophila NHAs are widely expressed but at particularly high levels in epithelia, suggesting a role in organismal ion homeostasis (15,16). Given that insect epithelia are energized by an apical plasma membrane H + V-ATPase (17, 18), the NHAs have been hypothesized to act as the partner exchangers, as predicted by Wieczorek and coworkers (17,19), that colocalize with V-ATPase and use the proton electrochemical gradient to achieve transepithelial transport of sodium and potassium, similar to the position in mammalian MDCK cells (20).Consistent with this, overexpression of GFP-tagged Nha1 or Nha2 labels the same apical membrane as the V-ATPase in the Malpighian (renal) tubule, a model epithelium in which transport and control mechanisms are conveniently studied (21,22), and RNAi against one of the exchangers affects fluid secretion (15). Thus, Drosophila provides an ideal system in which to investigate the roles of NHAs in multicellular animals.…”

“…Consistent with this, overexpression of GFP-tagged Nha1 or Nha2 labels the same apical membrane as the V-ATPase in the Malpighian (renal) tubule, a model epithelium in which transport and control mechanisms are conveniently studied (21,22), and RNAi against one of the exchangers affects fluid secretion (15). Thus, Drosophila provides an ideal system in which to investigate the roles of NHAs in multicellular animals.…”

The orphan pentameric ligand-gated ion channelpHCl-2is gated by pH and regulates fluid secretion inDrosophilaMalpighian tubules

“…In Drosophila, the Malpighian tubules consist of two pairs of blind-ended tubes that filter the haemolymph and form the primary urine. The Malpighian tubules are made up of two main cell types, the principal cells, which mediate the transcellular secretion of Na + and K + , and the stellate cells, which contribute to Cl − secretion (Blumenthal, 2003;Cabrero et al, 2014;O'Donnell et al, 1996O'Donnell et al, , 1998. In Drosophila, as in other insects, transepithelial secretion is an active process powered by an electrogenic V-type proton ATPase located in the apical membrane of the principal cells (Beyenbach, 1995;Du et al, 2006;Maddrell and O'Donnell, 1992;Torrie et al, 2004;Weng et al, 2003).…”

The rectal complex and Malpighian tubules of the cabbage looper (Trichoplusia ni): regional variations in Na+ and K+ transport and cation reabsorption by secondary cells

“…Stellate cells in tubules of Drosophila melanogaster are the site of kinin-stimulated Cl − secretion through Clc-a channels (Cabrero et al, 2014) and are an important site of water transport via aquaporins (Drosophila integral protein, DrIP) (Kaufmann et al, 2005). Immunohistochemical studies have provided evidence for multiple Na + and K + transporters in stellate cells of dipterans.…”