P-type Na+/K+-ATPase and V-type H+-ATPase expression patterns in the osmoregulatory organs of larval and adult mosquito Aedes aegypti

doi:10.1242/jeb.02551

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First published online November 17, 2006
Journal of Experimental Biology 209, 4638-4651 (2006)
Published by The Company of Biologists 2006
doi: 10.1242/jeb.02551

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P-type Na⁺/K⁺-ATPase and V-type H⁺-ATPase expression patterns in the osmoregulatory organs of larval and adult mosquito Aedes aegypti

Marjorie L. Patrick^*, Karlygash Aimanova, Heather R. Sanders and Sarjeet S. Gill

Department of Cell Biology and Neuroscience, University of California, Riverside, CA 92521-0146, USA

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Fig. 1. Immunolocalization of P-type Na⁺/K⁺-ATPase (red) and V-type H⁺-ATPase (blue) in the different regions of the midgut of fourth instar A. aegypti larvae. In the whole mounts of gastric caeca (GC; A), the proximal two thirds of each caecum expressed high levels of P-type Na⁺/K⁺-ATPase and the distal third expressed V-type H⁺-ATPase. (B-D) In the paraffin sections of caeca P-type Na⁺/K⁺-ATPase was localized to the basal membrane of the proximal segment only and V-type H⁺-ATPase was on the apical membrane throughout the caecum and also on the basal membrane on the distal caecum. In the very anterior of the midgut, high P-type Na⁺/K⁺-ATPase expression was found in the very anterior region of the midgut (E, white arrow) on the basal membrane (G, white arrow). The apical membrane in this region did not label for either ATPase (G). In the anterior midgut (AMG) there was high V-type H⁺-ATPase expression on the basal membrane (E) and P-type Na⁺/K⁺-ATPase was expressed apically (H). In the posterior midgut (PMG; F) V-type H⁺-ATPase was apically, and P-type Na⁺/K⁺-ATPase basally, expressed. All images were overlays of both Cy3 and Cy5 signals. Scale bars, 100 µm (A,B,E-G), 50 µm (C,D,H,I).

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Fig. 2. Immunolocalization of P-type Na⁺/K⁺-ATPase (red) and V-type H⁺-ATPase (blue) in the fourth instar larval A. aegypti Malpighian tubules, rectum and anal papillae. In the distal Malpighian tubule (MT) whole mounts (A,D,E) P-type Na⁺/K⁺-ATPase was highly expressed in the stellate cells (SC; D, white arrow), whereas V-type H⁺-ATPase was expressed only in the principal cells (PC; E, white arrow). V-type H⁺-ATPase was expressed in the distal principal cell on the apical membrane and cytoplasmically as evident in paraffin sections (H). In the proximal Malpighian tubules (B,C), there were no stellate cells and the principal cells had P-type Na⁺/K⁺-ATPase and V-type H⁺-ATPase on the basal and apical membranes, respectively, as seen in the paraffin sections (G). The rectal epithelium expressed high levels of P-type Na⁺/K⁺-ATPase (F) on the basal membrane (I, white arrow) whereas the anterior hindgut (AHG) showed little expression of either ATPase (F). P-type Na⁺/K⁺-ATPase and V-type H⁺-ATPase were localized to the basal and apical aspects respectively of the anal papillae (J,K). A,F,G,I-K are overlays of Cy3 and Cy5 signal and in B-E, H, the signals have been separated. Scale bars, 100 µm (A,F), 50 µm (B-E,I,J), 10 µm (G,H,K).

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Fig. 3. Immunolocalization of P-type Na⁺/K⁺-ATPase (red) and V-type H⁺-ATPase (blue) in the stomach and Malpighian tubules (MTs) of adult female A. aegypti. In a mid-longitudinal optical section of a whole mount of the stomach (A,B) labelling of P-type Na⁺/K⁺-ATPase was present on the lateral membranes and V-type H⁺-ATPase was present cytoplasmically. In the paraffin cross sections of the stomach epithelium, P-type Na⁺/K⁺-ATPase was present basolaterally and V-type H⁺-ATPase was present apically (C,D). In whole mounts of Malpighian tubules, the distal principal cells expressed only V-type H⁺-ATPase (F) and the stellate cells P-type Na⁺/K⁺-ATPase (E, white arrow). There were no stellate cells observed in the proximal region of the Malpighian tubules and the principal cells expressed both P-type Na⁺/K⁺-ATPase and V-type H⁺-ATPase (G). In the sections of the proximal principal cells (H), P-type Na⁺/K⁺-ATPase and V-type H⁺-ATPase were expressed on the basal (white arrow) and apical (white arrowhead) membranes, respectively. Also in H, the anterior hindgut (AHG) expressed P-type Na⁺/K⁺-ATPase on the basal membrane and V-type H⁺-ATPase on the apical membrane. In C,D,G,H, the Cy3 and Cy5 signals are overlayed and in A,B,E,F, the signals are separated. Scale bars, 100 µm (D), 50 µm (A-C,G,H), 10 µm (E,F).

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Fig. 4. Immunolocalization of P-type Na⁺/K⁺-ATPase (red) and V-type H⁺-ATPase (blue) in paraffin sections (A-D,F) and optical sections from whole mounts (E) of the anterior hindgut (AHG), rectum and rectal pads (RPs) of adult female A. aegypti. P-type Na⁺/K⁺-ATPase was localized to the basolateral membrane and V-type H⁺-ATPase apically of the of anterior hindgut epithelium (A,B,E,F). The rectal epithelium itself exhibited very low expression, whereas the rectal pads had very intense labeling of P-type Na⁺/K⁺-ATPase on the basolateral membrane, and V-type H⁺-ATPase was labelled on the apical membrane (C,D). All images are overlays of Cy3 and Cy5 signals. Scale bars, 100 µm (A,B), 50 µm (C,D,F), 10 µm (E).

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Fig. 5. Distribution of V-type H⁺-ATPase (866 bp band), (820 bp band) P-type Na⁺/K⁺-ATPase and 18s (201 and 100 bp bands) gene expression in (A) larval A. aegypti midgut, Malpighian tubules (MT), rectum and anal papillae, and (B) adult female A. aegypti Malpighian tubules (MT), anterior hindgut (AHG), rectum, and midgut. The right lane shows the no reverse transcriptase control in both figures.