Characterization of Transepithelial Transport in the Osmoregulation of Freshwater Dipteran Larvae
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In the osmoregulatory epithelia of freshwater (FW) animals, transcellular solute transport relies on at least one of the primary ionomotive pumps, Na+/K+-ATPase (NKA) and V-type H+-ATPase (VA), but there are no data on their distribution in FW chironomid larvae. The paracellular solute movement in invertebrate epithelia is controlled by the septate junctions (SJs). However, nothing is known about the SJ components in aquatic insects. In the present set of studies, larvae of the FW chironomid Chironomus riparius and FW mosquito Aedes aegypti were used to examine a role for NKA and VA and SJ proteins, respectively, in the maintenance of salt and water balance in aquatic dipterans. Spatial distribution and activity of NKA and VA along the alimentary canal of C. riparius larvae revealed the importance of the rectum in the ionoregulatory homeostasis. It was found that the rectum absorbed relatively high amounts of K+ into the hemolymph under dilute conditions and decreased K+ absorption in brackish water (BW). This rectal K+ absorption was dependent on the activities of both NKA and VA. Next, genes encoding transmembrane SJ proteins megatrachea, sinuous, kune-kune (Kune), neurexin IV, snakeskin (Ssk), mesh and gliotactin (Gli) were identified in A. aegypti and shown to exhibit tissue specific transcript abundance in larval osmoregulatory epithelia. Ssk and mesh expression was restricted to smooth SJ bearing midgut and Malpighian tubules (MT) whereas Gli was detected in all tissues examined. Kune was confined to SJs in the posterior midgut and rectum and apical membrane domain of the syncytial anal papilla epithelium. Rearing A. aegypti larvae in BW caused an increase in Kune and Gli protein and Ssk and mesh mRNA abundance in the midgut and MT which occurred in conjunction with increased midgut and decreased MT permeability. Paracellular MT permeability was further modulated by leucokinin. When dsRNA was used to reduce Gli abundance in the midgut, paracellular permeability was decreased. Together, this research provides a better understanding of the physiology of transepithelial ion and water transport in aquatic insects and offers significant insight into the role of NKA, VA and SJ proteins in the osmoregulation of FW dipteran larvae.