NHE8 is an intracellular cation/H+exchanger in renal tubules of the yellow fever mosquitoAedes aegypti

The goal of this study was to identify and characterize the hypothesized apical cation/H(+) exchanger responsible for K(+) and/or Na(+) secretion in the renal (Malpighian) tubules of the yellow fever mosquito Aedes aegypti. From Aedes Malpighian tubules, we cloned "AeNHE8," a full-length cDNA encoding an ortholog of mammalian Na(+)/H(+) exchanger 8 (NHE8). The expression of AeNHE8 transcripts is ubiquitous among mosquito tissues and is not enriched in Malpighian tubules. Western blots of Malpighian tubules suggest that AeNHE8 is expressed primarily as an intracellular protein, which was confirmed by immunohistochemical localizations in Malpighian tubules. AeNHE8 immunoreactivity is expressed in principal cells of the secretory, distal segments, where it localizes to a subapical compartment (e.g., vesicles or endosomes), but not in the apical brush border. Furthermore, feeding mosquitoes a blood meal or treating isolated tubules with dibutyryl-cAMP, both of which stimulate a natriuresis by Malpighian tubules, do not influence the intracellular localization of AeNHE8 in principal cells. When expressed heterologously in Xenopus laevis oocytes, AeNHE8 mediates EIPA-sensitive Na/H exchange, in which Li(+) partially and K(+) poorly replace Na(+). The expression of AeNHE8 in Xenopus oocytes is associated with the development of a conductive pathway that closely resembles the known endogenous nonselective cation conductances of Xenopus oocytes. In conclusion, AeNHE8 does not mediate cation/H(+) exchange in the apical membrane of Aedes Malpighian tubules; it is more likely involved with an intracellular function.