The NMDA receptor regulates integrin activation, ATP release and arterial thrombosis through store-operated Ca2+ entry in platelets

Friedrich Reusswig; Münteha Yilmaz; Marius Brechtenkamp; Irena Krueger; Lisa Maria Metz; Nikolaj Klöcker; Eckhard Lammert; Margitta Elvers

doi:10.3389/fcvm.2023.1171831

The NMDA receptor regulates integrin activation, ATP release and arterial thrombosis through store-operated Ca²⁺ entry in platelets

Front Cardiovasc Med. 2023 May 12:10:1171831. doi: 10.3389/fcvm.2023.1171831. eCollection 2023.

Authors

Friedrich Reusswig¹, Münteha Yilmaz¹, Marius Brechtenkamp¹, Irena Krueger¹, Lisa Maria Metz¹, Nikolaj Klöcker², Eckhard Lammert^{3

4

5}, Margitta Elvers¹

Affiliations

¹ Department of Vascular- and Endovascular Surgery, University Hospital Düsseldorf, Heinrich-Heine University, Düsseldorf, Germany.
² Institute of Neural and Sensory Physiology, Medical Faculty and University Hospital Düsseldorf, Heinrich-Heine University, Düsseldorf, Germany.
³ Institute for Vascular and Islet Cell Biology, German Diabetes Center (DDZ), Leibniz Center for Diabetes Research at Heinrich-Heine University, Düsseldorf, Germany.
⁴ German Center for Diabetes Research (DZD e.V.), Helmholtz Zentrum München, Neuherberg, Germany.
⁵ Institute of Metabolic Physiology, Heinrich Heine University, Düsseldorf, Germany.

Abstract

Introduction: Platelet activation and thrombus formation is crucial for hemostasis, but also trigger arterial thrombosis. Calcium mobilization plays an important role in platelet activation, because many cellular processes depend on the level of intracellular Ca²⁺ ([Ca²⁺](i)), such as integrin activation, degranulation, cytoskeletal reorganization. Different modulators of Ca²⁺ signaling have been implied, such as STIM1, Orai1, CyPA, SGK1, etc. Also, the N-methyl-D-aspartate receptor (NMDAR) was identified to contribute to Ca²⁺ signaling in platelets. However, the role of the NMDAR in thrombus formation is not well defined.

Methods: In vitro and in vivo analysis of platelet-specific NMDAR knock-out mice.

Results: In this study, we analyzed Grin1^fl/fl-Pf4-Cre⁺ mice with a platelet-specific knock-out of the essential GluN1 subunit of the NMDAR. We found reduced store-operated Ca²⁺ entry (SOCE), but unaltered store release in GluN1-deficient platelets. Defective SOCE resulted in reduced Src and PKC substrate phosphorylation following stimulation of glycoprotein (GP)VI or the thrombin receptor PAR4 followed by decreased integrin activation but unaltered degranulation. Consequently, thrombus formation on collagen under flow conditions was reduced ex vivo, and Grin1^fl/fl-Pf4-Cre⁺ mice were protected against arterial thrombosis. Results from human platelets treated with the NMDAR antagonist MK-801 revealed a crucial role of the NMDAR in integrin activation and Ca²⁺ homeostasis in human platelets as well.

Conclusion: NMDAR signaling is important for SOCE in platelets and contributes to platelet activation and arterial thrombosis. Thus, the NMDAR represents a novel target for anti-platelet therapy in cardiovascular disease (CVD).

Keywords: ATP; NMDAR; calcium; integrin; platelets; thrombosis.

Grants and funding

The study was funded by the Deutsche Forschungsgemeinschaft (DFG, German Research Foundation), Collaborative Research Centre TRR259 (Aortic Disease)—Grant No. 397484323, TP A07 to ME.