When platelets are suspended in hypotonic media, they swell, then slowly recover their original volume. This phenomenon has been utilized in an assay system in which platelets in plasma are diluted to water and the time course of the cell size changes followed by measurement of optical density. Recovery from this """"""""hypotonic shock"""""""" has been found to correlate well with other viability measurements of platelets stored for transfusion. Yet, the cellular basis for this correlation, and for the recovery phenomenon itself, are not at all understood. The usual method for performance of the assay involves diluting platelets in plasma in cuvette and observing the optical density change with time. This is slow, cumbersome and severely limits attempts to correlate the volume changes with other cellular events. We have adapted microtiter plate technology to the performance of hypotonic shock assays. This permits performance of 96 assays simultaneously and is a major advance from a technical point of view. We have begun to examine cytoskeletal protein changes at intervals after hyptonic challenge and to correlate these with cellular swelling, as determined by optical density. We will set up other methods for measurement of cell volume in order to verify the optical density readings. Since the recovery response to hypotonic shock decreases with time in storage, we will also attempt to correlate the reaction with other events thought to be related to the storage lesion. One possibility is activation of platelet release mechanisms. Such activation has been followed by release of intracellular constituents and by measurement of entities (such as GMP-140, or PADGEM) which are expressed during activation.
