To accommodate expanding volume (V) during hyposmotic swelling, animal cells change their shape and increase surface area (SA) by drawing extra membrane from surface and intracellular reserves. The relative contributions of these processes, sources and extent of membrane reserves are not well defined. In this study, the SA and V of single substrate-attached A549, 16HBE14o−, CHO and NIH 3T3 cells were evaluated by reconstructing cell three-dimensional topology based on conventional light microscopic images acquired simultaneously from two perpendicular directions. The size of SA reserves was determined by swelling cells in extreme 98% hypotonic (∼6 mOsm) solution until membrane rupture; all cell types examined demonstrated surprisingly large membrane reserves and could increase their SA 3.6 ± 0.2-fold and V 10.7 ± 1.5-fold. Blocking exocytosis (by N-ethylmaleimide or 10°C) reduced SA and V increases of A549 cells to 1.7 ± 0.3-fold and 4.4 ± 0.9-fold, respectively. Interestingly, blocking exocytosis did not affect SA and V changes during moderate swelling in 50% hypotonicity. Thus, mammalian cells accommodate moderate (<2-fold) V increases mainly by shape changes and by drawing membrane from preexisting surface reserves, while significant endomembrane insertion is observed only during extreme swelling. Large membrane reserves may provide a simple mechanism to maintain membrane tension below the lytic level during various cellular processes or acute mechanical perturbations and may explain the difficulty in activating mechanogated channels in mammalian cells.
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This study was supported by the Canadian Institutes of Health Research, the Canadian Cystic Fibrosis Foundation and Natural Sciences and Engineering Research Council of Canada. We thank Dr. Jean-Yves Lapointe for help during the development of our imaging technique; Drs. Cathy Morris, Fred Sachs and Andrzej Kubalski for their comments on the manuscript; and Ms. Helene Chabot for technical assistance.
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Groulx, N., Boudreault, F., Orlov, S.N. et al. Membrane Reserves and Hypotonic Cell Swelling. J Membrane Biol 214, 43–56 (2006) doi:10.1007/s00232-006-0080-8
- Cell volume
- Membrane reserve
- Membrane stretch
- Three-dimensional cell imaging