Strong alkalinization in the anterior midgut of larval yellow fever mosquitoes (Aedes aegypti): involvement of luminal Na⁺/K⁺‐ATPase

Abstract

Recently, Na + /K + -ATPase has been detected in the luminal membrane of the anterior midgut of larval yellow fever mosquitoes (Aedes aegypti) with immunohistochemical techniques. In this study, the possible involvement of this ATPase in strong alkalinization was investigated on the level of whole larvae, isolated and perfused midgut preparations and on the molecular level of the Na + /K + -ATPase protein. Ouabain (5 mM) did not inhibit the capability of intact larval mosquitoes to alkalinize their anterior midgut. Also in isolated and perfused midgut preparations the perfusion of the lumen with ouabain (5 mM) did not result in a significant change of the transepithelial voltage or the capacity of luminal alkalinization. Na + /K + -ATPase activity was completely abolished when KCl was substituted with choline chloride, suggesting that the enzyme cannot act as an ATP-driven Na + /H + -exchanger. Altogether the results of the present investigation indicate that apical Na + /K + -ATPase is not of direct importance for strong luminal alkalinization in the anterior midgut of larval yellow fever mosquitoes.Strong luminal alkalinization of up to pH values of 12 has been observed in midgut regions of many larvae of endopterygote insects, including members of the orders Coleoptera, Diptera, Trichoptera and Lepidoptera (for references see Clark, '99). Two tissues have been studied especially intensively with regard to their mechanisms and capacities of alkalinization during the past 20 years: the midgut of the tobacco hornworm (Manduca sexta) and the anterior midgut of larval yellow fever mosquitoes (Aedes aegypti). For the lepidopteran larvae, a transport model has been established in which luminal V-type H + -pumps (V-ATPases) energize luminal, electrogenic K + /2H + -exchangers, resulting in active K + secretion and strong luminal alkalinization (for a review see Wieczorek et al., '99). Unfortunately, this mechanism can so far not be tested with the isolated epithelium, because it loses the capacity for strong luminal alkalinization after isolation, whereas active K + secretion continues in vitro at a high rate (Clark et al., '98). Strong luminal alkalinization in the anterior midgut of larval yellow fever mosquitoes appears to significantly differ from lepidopterans, including the methodological aspect that it can readily be observed with isolated and perfused midgut preparations (Onken et al., 2008). In larval A. aegypti, the V-ATPase is not localized in the luminal membrane of the anterior midgut, but instead in the basolateral membrane (Zhuang et al., '99). The epithelium of larval yellow fever mosquitoes generates a lumen negative transepithelial voltage (Clark et

Revisiting the cellular mechanisms of strong luminal alkalinization in the anterior midgut of larval mosquitoes

“…It has been known that the ATPase can accept H + instead of Na + and/or K + (Polvani and Blostein, 1988). However, when we tested this hypothesis by addition of ouabain (5mmoll −1 ), a specific inhibitor of the Na + /K + ATPase, to the luminal perfusate, there was no significant effect on V te , and alkalinization was not impaired (Onken et al, 2009). …”

Ion and solute transport by Prestin in Drosophila and Anopheles

“…The luminal pH of the Drosophila middle midgut is less than 3.0, while the pH of the posterior midgut is greater than 10.0 (Boudko et al, 2001a; Boudko et al, 2001b; Jagadeshwaran et al, 2010; Onken and Moffett, 2009; Onken et al, 2008; Onken et al, 2009; Shanbhag and Tripathi, 2005, 2009). Moreover, the luminal pH of the gut segment between the gastric caeca and the anterior midgut in mosquito larvae is over 10.5 (Linser et al, 2011).…”