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Environmental and Stress Physiology

The Mechanics of Injury to Isolated Protoplasts following Osmotic Contraction and Expansion ¹

Department of Agronomy, Cornell University, Ithaca, New York 14853-0144, School of Physics, University of New South Wales, Kensington New South Wales 2033 Australia

Micro-osmotic manipulation was used to determine the influence of osmotic contraction on the expansion potential of individual protoplasts isolated from rye (Secale cereale L. cv Puma) leaves. For protoplasts isolated from leaves of nonacclimated plants (NA protoplasts), osmotic contraction in sufficiently hypertonic solutions (>1.53 osmolal) predisposed the protoplasts to lysis during osmotic expansion when they were returned to isotonic conditions (0.53 osmolal). In contrast, for protoplasts isolated from leaves of cold acclimated plants (ACC protoplasts), osmotic contraction in either 2.6 or 4.0 osmolal solutions was readily reversible. Following osmotic contraction, the resting tension (_r) of NA protoplasts was similar to that determined for protoplasts in isotonic solutions (i.e. 110 ± 22 micronewtons per meter). In contrast, _r of ACC protoplasts decreased from 164 ± 27 micronewtons per meter in isotonic solutions to values close to zero in hypertonic solutions. Following expansion in hypotonic solutions, _r's of both NA and ACC protoplasts were similar for area expansions over the range of 1.3 to 1.6. Following osmotic contraction and reexpansion of NA protoplasts, hysteresis was observed in the relationship between _r and surface area—with higher values of _r at a given surface area. In contrast, no hysteresis was observed in this relationship for ACC protoplasts. Direct measurements of plasma membrane tension () during osmotic expansion of NA protoplasts from hypertonic solutions (1.53 osmolal) revealed that increased rapidly after small increments in surface area, and lysis occurred over a range of 1.2 to 8 millinewtons per meter. During osmotic expansion of ACC protoplasts from hypertonic solutions (2.6 osmolal), there was little increase in until after the isotonic surface area was exceeded. These results are discussed in relation to the differences in the behavior of the plasma membrane of NA and ACC protoplasts during osmotic contraction (i.e. endocytotic vesiculation versus exocytotic extrusion) and provide a mechanistic interpretation to account for the differential sensitivity of NA and ACC protoplasts to osmotic expansion from hypertonic solutions.

¹ This material is, in part, based on work supported by the United States Department of Energy under Contract No. DE-AC02-81ER10917 and Grant No. DE-FG03-84ER13214. Department of Agronomy series paper No. 1573.

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