Cited by 4.3k
University of London
Publishes on Antiplatelet Therapy and Cardiovascular Diseases, Platelet Disorders and Treatments, Blood properties and coagulation. 164 papers and 11.8k citations.
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A new technique was developed for the measurement of extracellular free ATP in very small samples of whole blood using the luciferin-luciferase enzyme system. The method had a very low background corresponding to approximately 10(-16) mol ATP. ATP was measured in blood as it emerged during haemostasis following precise puncture of rat and rabbit arteries and after standardized incisions of human skin by the Simplate device. The initial concentration of free ATP in blood emerging 2-4 s after vascular injury was about 2 X 10(-7) M in rats and rabbits and about 2 X 10(-6) M in humans. The free-ATP concentration increased to 2 X 10(-5) M 3-5 min after injury and these increases could be prevented by heparin (20 u./ml). The source of the initial free ATP was identified as damaged cells in the injured vessel wall. Sufficient ADP, both released as such with ATP and generated by enzymic dephosphorylation of ATP, would be present at the site of injury to initiate haemostatic aggregation of platelets.
1. An optical method used for measuring platelet aggregation was adapted for measurements of the change in shape of platelets that rapidly follows the addition of the aggregating agent adenosine diphosphate (ADP).2. Measurements were made using dilute suspension of platelets with sufficient EDTA to prevent their aggregation.3. Measurements of the velocity and magnitude of the optical effects of the shape change were highly reproducible.4. Volumetric measurements showed that the shape change is not associated with an increase in mean platelet volume.5. The velocity of the shape change had a temperature coefficient of about 4.5.6. The velocity and magnitude of the shape change were not affected by pH between 5.8 and 9.2.7. The dependence of the velocity of the shape change on the ADP concentration was in accordance with Michaelis-Menton kinetics. The K(m) was about 7.2 x 10(-7)M.8. The velocity of the shape change was inhibited competitively by ATP, adenosine and 2-chloroadenosine but not by AMP. When the inhibitors were added after the maximum of the shape change they caused a concentration-dependent diminution in the record of the change.9. The results suggest that the shape change is initiated by reaction of the agonist ADP with specific receptor sites on the platelet membrane which leads to energy-requiring changes in the structures responsible for maintaining the disk shape of normal platelets.