Macrovacuolation induced by cytochalasin: its relation to the cytoskeleton; morphological and cytochemical...

At higher doses of cytochalasin (e.g. 3 micrograms/ml for 3-20 hr), cells of the rat fibroblastoid line, Hmf, undergo extreme retraction, arborization, and subsequent rounding, and develop big cystic vacuoles. These vacuoles are always closely invested by microfilamentous masses, the CD-induced derivatives of the actin-based cytoskeleton, which aggregate in the endoplasm. Vacuolation is progressive (e.g. 12% cells at 6 hr; greater than 80% at 18 hr), related to total dose (concentration X time); and to congener (CD greater than CB). Vacuole membranes have the same dimension (85 A), surface marker 5'-nucleotidase, and junctional specializations as those found at the cell surface; they lack the membrane markers associated with endomembrane systems (e.g. AcPase, TPPase, IDPase) and are not lysosomal. Vacuoles represent internalized plasma membrane; they apparently result from retention in the endoplasm, and fusion, of pinocytotic vesicles originating at the cell surface. Vacuole membrane is always in intimate relation to the actin-based microfilament aggregates that surround the vacuoles, and actin-membrane linker proteins fodrin and vinculin are localized at the vacuole boundaries. Vacuoles and their enveloping actin-filament aggregates are surrounded by arrays of vimentin-based intermediate filaments. A new membranous compartment with characteristics of plasma membrane is thus formed within the cell under the influence of CD. Rounding brought about by other means causes no vacuolization. Macrovacuolation, like the other changes caused by CD, is completely reversible on restoration of cells to normal medium.