Serotoninergic Mechanisms Play a Major Role on Platelet-Mediated Thrombogenicity: Studies under Flow Conditions and Effects of Inhibitory Strategies.

Serotoninergic mechanisms are reciprocally implicated in depression and cardiovascular disorders. Serotonin (5-HT) may facilitate the development of a subpopulation of platelets with increased procoagulant activity. We have investigated the involvement of serotoninergic mechanisms in adhesive, cohesive and procoagulant function of platelets. Furthermore, we evaluated the antithrombotic properties of selective serotonin reuptake inhibitors (SSRI). For these purposes we used a series of experimental strategies including standard aggregometry, flow cytometry, perfusion techniques and determination of coagulation parameters. Serotonin concentrations ranging 0.5–5 μM were used in these studies. Citalopram, a selective serotonin reuptake inhibitor (SSRI), was used at concentrations equivalent to those reached in clinical practice.

Aggregation studies indicate that 5-HT is a weak agonist for platelets in comparison with standard activating agents. Only the highest concentrations of 5-HT tested (5 μM) caused minimal and reversible platelet aggregation. Despite this modest effect on aggregation, 5-HT potentiated the aggregation induced by low concentrations of ADP (0.5 μM) in PRP samples (16.3±5.9 % vs. 28.1±5.5 %). Potentiation of aggegation was more evident when PRP was obtained from blood anticoagulated with LMWH (p<0.05), where some thrombin could be generated. Citalopram at 0.6 μM caused statistical reductions in % of maximal aggregation induced by ADP (2 or 0.5 μM) and COL (2.5 μg/ml) (p<0.05). Platelet aggregation was maximally inhibited with citalopram at concentrations exceeding clinical doses (6 μM). Flow cytometry studies revealed a mild increase of P-selectin expression in washed platelets in the first minute after activation with 5-HT. In isolated platelets, 5-HT produced irreversible aggregation when studies were performed in the presence of human-TF enriched microvesicles. A significant increase in Annexin V binding and presence of factor Va was detected by flow cytometry in the latter studies. Further studies performed in perfusion models using human blood flowing though chambers exposing a surface rich in collagen and TF demonstrated that presence of 5-HT in the perfusates increased thrombin generation (p<0.05) as demonstrated by elevations of F1+2 levels. Presence of citalopram in the perfusates significantly inhibited the formation of large platelet aggregates on the perfused thrombogenic surfaces (20.2±3.4 % vs. 11.5±1.7 %; p<0.05) and prevented elevations of F1+2 levels observed in previous studies. Our studies confirm that 5-HT is a weak agonist for platelets, though it possesses a marked ability to enhance the activation induced by other agonists. Moreover, 5-HT showed a powerful capacity to potentiate procoagulant responses of platelets under different experimental conditions and to enhance the thrombogenesis on damaged vascular surfaces. The fact that SSRI inhibited the responses elicited by 5-HT in the previous situations further confirm the implications of serotoninergic mechanisms in platelet activation. Our data reinforce the opinion that serotoninergic mechanisms play an important role in platelet-mediated thrombogenicity, suggesting that modulation of 5-HT mediated responses may offer a new potential target for the development of more powerful antithrombotic drugs.