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A Mechanical Force Contributes to the "Osmotic Swelling" of Brush-Border Membrane Vesicles

Martin Kirouac ^* , Vincent Vachon ^* , Mélanie Fortier ^* , Marie-Claude Trudel ^*, Alfred Berteloot ^* , Jean-Louis Schwartz ^*  and Raynald Laprade ^* 

^* Membrane Protein Research Group; Department of Physics; and Department of Physiology, University of Montreal, Montreal, Quebec, Canada

Correspondence: Address reprint requests to Raynald Laprade, Groupe d'étude des protéines membranaires, University of Montreal, PO Box 6128, Centre Ville Station, Montreal, Quebec, H3C 3J7, Canada. Tel.: 514-343-7960; Fax: 514-343-7146; E-mail: raynald.laprade{at}umontreal.ca.

Brush-border membrane vesicles and an osmotic swelling assay have been used extensively to monitor the pore-forming activity of Bacillus thuringiensis toxins. After a hypertonic shock, Manduca sexta midgut brush-border membrane vesicles shrink rapidly and reswell partially to a volume that depends on membrane permeability and toxin concentration rather than regaining their original volume as expected from theoretical models. Because efflux of buffer from the vesicles, as they shrink, could contribute to this phenomenon, vesicles were mixed with a hypertonic solution of the buffer with which they were loaded. Under these conditions, they are not expected to reswell, since the same solute is present on both sides of the membrane. Nevertheless, with several buffers, vesicles reswelled readily, an observation that demonstrates the involvement of an additional restoration force. Reswelling also occurred when, in the absence of toxin, the buffers were replaced by glucose, a solute that diffuses readily across the membrane, but did not occur with rat liver microsomes, despite their permeability to glucose. Unexpected swelling was also observed with rabbit jejunum brush-border membrane vesicles, suggesting that the cytoskeleton, present in brush-border membrane vesicles but absent from microsomes, could be responsible for the restoration force.