title: Dynamics of calcium spiking, mitochondrial collapse and phosphatidylserine exposure in platelet subpopulations during activation.
abstract: Essentials The sequence and logic of events leading to platelet procoagulant activity are poorly understood. Confocal time-lapse microscopy was used to investigate activation of single adherent platelets. Platelet transition to the procoagulant state followed cytosolic calcium oscillations. Mitochondria did not collapse simultaneously and membrane potential loss could be reversible.
  
  
  
  Background Activated platelets form two subpopulations, one of which is able to efficiently aggregate, and another that externalizes phosphatidylserine (PS) and thus accelerates membrane-dependent reactions of blood coagulation. The latter, procoagulant subpopulation is characterized by a high cytosolic calcium level and the loss of inner mitochondrial membrane potential, and there are conflicting opinions on their roles in its formation. Methods We used confocal microscopy to investigate the dynamics of subpopulation formation by imaging single, fibrinogen-bound platelets with individual mitochondria in them upon loading with calcium-sensitive and mitochondrial potential-sensitive dyes. Stimulation was performed with thrombin or the protease-activated receptor (PAR) 1 agonist SFLLRN. Stochastic simulations with a computational systems biology model of PAR1 calcium signaling were employed for analysis. Results Platelet activation resulted in a series of cytosolic calcium spikes and mitochondrial calcium uptake in all platelets. The frequency of spikes decreased with time for SFLLRN stimulation, but remained high for a long period of time for thrombin. In some platelets, uptake of calcium by mitochondria led to the mitochondrial permeability transition pore opening and inner mitochondrial membrane potential loss, which could be either reversible or irreversible. The latter resulted in an increase in the cytosolic calcium level and PS exposure. These platelets had higher cytosolic calcium levels before activation, and their mitochondria collapsed not simultaneously but one after another. Conclusions These results support a model of procoagulant subpopulation development following a series of stochastic cytosolic calcium spikes that are accumulated by mitochondria, leading to a collapse, and suggest important roles of individual platelet reactivity and signal exchange between different mitochondria of a platelet.
authors.affiliation: Federal Research and Clinical Center of Pediatric Hematology, Oncology and Immunology, Moscow, Russia.|Center for Theoretical Problems of Physicochemical Pharmacology, Moscow, Russia.
authors.forename: S I
authors.initials: SI
authors.lastname: Obydennyy
authors.affiliation: Federal Research and Clinical Center of Pediatric Hematology, Oncology and Immunology, Moscow, Russia.|Center for Theoretical Problems of Physicochemical Pharmacology, Moscow, Russia.|Faculty of Physics, Lomonosov Moscow State University, Moscow, Russia.|Therapeutic Faculty, Pirogov Russian National Research Medical University, Moscow, Russia.
authors.forename: A N
authors.identifier: 0000-0003-4720-7319
authors.initials: AN
authors.lastname: Sveshnikova
authors.affiliation: Federal Research and Clinical Center of Pediatric Hematology, Oncology and Immunology, Moscow, Russia.|Center for Theoretical Problems of Physicochemical Pharmacology, Moscow, Russia.|Faculty of Physics, Lomonosov Moscow State University, Moscow, Russia.
authors.forename: F I
authors.initials: FI
authors.lastname: Ataullakhanov
authors.affiliation: Federal Research and Clinical Center of Pediatric Hematology, Oncology and Immunology, Moscow, Russia. mapanteleev@yandex.ru.|Center for Theoretical Problems of Physicochemical Pharmacology, Moscow, Russia. mapanteleev@yandex.ru.|Faculty of Physics, Lomonosov Moscow State University, Moscow, Russia. mapanteleev@yandex.ru.|Department of Biological and Medical Physics, Moscow Institute of Physics and Technology, Moscow, Russia. mapanteleev@yandex.ru.
authors.forename: M A
authors.initials: MA
authors.lastname: Panteleev
mesh_terms: D001777:Blood Coagulation; D001792:Blood Platelets / Q000378:metabolism; D002118:Calcium / Q000737:chemistry*; D020013:Calcium Signaling; D003198:Computer Simulation; D003600:Cytosol / Q000378:metabolism; D006801:Humans; D053078:Membrane Potential, Mitochondrial; D018613:Microscopy, Confocal; D008928:Mitochondria / Q000378:metabolism*; D033681:Mitochondrial Membrane Transport Proteins / Q000378:metabolism; D000083162:Mitochondrial Permeability Transition Pore; D010446:Peptide Fragments / Q000737:chemistry; D010718:Phosphatidylserines / Q000737:chemistry*; D015539:Platelet Activation; D010973:Platelet Adhesiveness; D015398:Signal Transduction; D013269:Stochastic Processes; D013917:Thrombin / Q000378:metabolism; D010975:Platelet Aggregation Inhibitors
chemical_list: D033681:Mitochondrial Membrane Transport Proteins; D000083162:Mitochondrial Permeability Transition Pore; D010446:Peptide Fragments; D010718:Phosphatidylserines; C082835:thrombin receptor peptide (42-47); D013917:Thrombin; D002118:Calcium; D010975:Platelet Aggregation Inhibitors
keywords: blood platelets; calcium signaling; mitochondria; platelet activation; protease-activated receptors