ADP-Induced Refractory State of Platelets in Vitro II. Functional and Ultra Studies on Gel Filtered Platelets

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Abstract
Gel filtered platelets (GFP) [human] in Ca free Tyrode solution containing albumin, glucose and adenosine deaminase were preincubated with 1 .mu.M 14C-ADP or 0.15 M NaCl (control) at 37.degree. C. The breakdown of extracellular 14C-ADP was markedly inhibited in this medium. No aggregation took place without fibrinogen, but the platelets underwent a disc to sphere transformation with development of refractoriness towards ADP. Presence of 2 mM CaCl2 in the incubation medium did not prevent refractoriness as reported earlier with washed rabbit platelets. When the ADP degrading enzyme, apyrase, was added at 30 min of incubation, a partial recovery of the aggregability was observed. EM studies showed that the partial restoration of the aggregation response, due to ADP degradation by apyrase, was accompanied by a return of discoidal morphology of the platelets. The ultrastructural studies showed further that spherical form with large number of pseudopods is not by itself a necessary or sufficient indication of platelets in a refractory state. Spherical platelets are apparently more vulnerable to external factors. Refractoriness was mainly caused by a direct effect on the platelets by ADP itself, but the studies also suggested that deteriorating, irreversible, intracellular changes may take place when platelets are in spherical shape. An artificial medium, mechanical stress, incubation at 37.degree. C are factors that probably speed up these changes.

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