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Swimbladder gas gland cells cultured on permeable supports regain their characteristic polarity

Caroline Prem and Bernd Pelster^*

Institut für Zoologie und Limnologie, Universität Innsbruck, A-6020 Innsbruck, Austria

View larger version (122K): [in a new window]   Fig. 1. Electron micrograph of a gas gland cell cultured on a permeable filter membrane. The cell is columnar and shows a clear polarity with some microvilli at the luminal membranes and extensive membrane foldings in both the lateral and the basal membranes. The cytoplasm contains filamentous mitochondria and a Golgi apparatus. Lamellar bodies are most commonly seen near the luminal membranes. g, Golgi apparatus; lb, lamellar body; m, filamentous mitochondria; mv, microvilli; n, nucleus. Scale bar, 1 µm.

View larger version (217K): [in a new window]   Fig. 2. Electron micrograph of gas gland cells forming a pseudostratified epithelium. In this case, only luminal cells show a clear polarity, with some microvilli at the luminal membranes and extensive membrane foldings in the basolateral membranes. bl, basolateral membrane foldings; m, filamentous mitochondria; n, nucleus. Scale bar, 1 µm.

View larger version (47K): [in a new window]   Fig. 3. Immunohistochemical localization of Na+/K+-ATPase in primary cultured eel gas gland cells. The confocal image shows three optical sections of a gas gland cell in the xy (A), the yz (B) and the xz (C) planes. The white bars in A (the xy section) indicate the locations of the other two sections; the white bar in C (the xz section) indicates the location of the xy section in A. Note that Na+/K+-ATPase immunoreactivity is located at lateral cell membranes, but there is no immunoreactivity at the apical membranes. Scale bar, 1 µm.

View larger version (59K): [in a new window]   Fig. 4. Three-dimensional reconstruction of Na+/K+-ATPase in cultured gas gland cells, calculated on the basis of the confocal images shown in Fig. 3. Na+/K+-ATPase is localized to the lateral and to some basolateral membranes. Scale bar, 1 µm.

View larger version (30K): [in a new window]   Fig. 5. A typical western blot of eel swimbladder homogenate after incubation with chicken Na+/K+-ATPase antibody. In negative controls performed without incubation with the primary antibody no bands were detected. Molecular mass standards (in kDa) (Amersham rainbow marker RPN756 was used) have been labelled by hand.

View larger version (15K): [in a new window]   Fig. 6. Changes in transepithelial resistance Rt of cultured swimbladder gas gland cells over time with DMEM medium supplied to the basal membrane and buffer solution to the apical membrane. Cells were seeded onto Costar Transwell 13 membranes at day 0. Values are means ± S.E.M., N=6

View larger version (26K): [in a new window]   Fig. 7. A comparison of the lactate released (as a percentage of total lactate released) from the basal (circles) and apical (triangles) sides of gas gland cells cultured on permeable Anodisc 13 membranes over a period of 2 days. The total rate of lactate release in these experiments was between 0.5 and 0.6 nmol h–1. Values are means ± S.E.M.; N=20.

View larger version (29K): [in a new window]   Fig. 8. Average rate of total lactate release of gas gland cells cultured on permeable supports in a superfusion system with fluid supplied to both sides of the superfusion system (liquid culture) and in an air/liquid system, in which humidified air was supplied to the apical side of the cells. Values are mean ± S.E.M.; N=20.

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