Quantitative Assessment of Polymerized and Depolymerized Platelet Microtubules

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Abstract
The equilibrium between assembled and disassembled microtubules was studied in human platelets exposed to aggregating agents. Soluble and insoluble tubulin were “frozen” by addition of a glycerol-dimethyl sulfoxide-containing medium. The two pools were estimated by measuring the colchicine binding activities of total and polymerized tubulin. Resting platelets were found to contain an average of 56.2 μg tubulin/1 × 109 cells of which 56.7% was in polymerized form. Platelet aggregation induced by thrombin, ADP, epinephrine, or collagen produced a transient decrease in the pool of polymerized tubulin which was evident within 15 s after addition of the aggregating agent. A return to base-line values occurred within 1-4 min depending upon the specific aggregating agent used. Neither secretory release nor aggregation of platelets were found to be prerequisites for the temporary disturbance of the equilibrium between soluble and polymerized tubulin. With thrombin as the aggregating agent a clear threshold concentration could be demonstrated above with a dose-dependent dissociation response of microtubules was evident. We conclude that microtubules exist in a dynamic equilibrium between polymerized and depolymerized forms in human platelets, which is transiently disturbed by their interaction with aggregating agents.