Deletion of Grin1 in mouse megakaryocytes reveals NMDA receptor role in platelet function and proplatelet formation.

James I Hearn, Taryn N Green, Colin L Hisey, Markus Bender, Emma C Josefsson, Nicholas Knowlton, Juliane Baumann, Raewyn C Poulsen, Stefan K Bohlander, Maggie L Kalev-Zylinska

Blood 2022 Apr 28

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The process of proplatelet formation (PPF) requires coordinated interaction between megakaryocytes (MKs) and the extracellular matrix (ECM), followed by a dynamic reorganization of the actin and microtubule cytoskeleton. Localized fluxes of intracellular calcium ions (Ca2+) facilitate MK-ECM interaction and PPF. Glutamate-gated N-methyl-D-aspartate receptor (NMDAR) is highly permeable to Ca2+. NMDAR antagonists inhibit MK maturation ex vivo; however, there are no in vivo data. Using the Cre-loxP system, we generated a platelet lineage-specific knockout mouse model of reduced NMDAR function in MKs and platelets (Pf4-Grin1-/- mice). Effects of NMDAR deletion were examined using well-established assays of platelet function and production in vivo and ex vivo. We found that Pf4-Grin1-/- mice had defects in megakaryopoiesis, thrombopoiesis, and platelet function, which manifested as reduced platelet counts, lower rates of platelet production in the immune model of thrombocytopenia, and prolonged tail bleeding time. Platelet activation was impaired to a range of agonists associated with reduced Ca2+ responses, including metabotropic like, and defective platelet spreading. MKs showed reduced colony and proplatelet formation. Impaired reorganization of intracellular F-actin and α-tubulin was identified as the main cause of reduced platelet function and production. Pf4-Grin1-/- MKs also had lower levels of transcripts encoding crucial ECM elements and enzymes, suggesting NMDAR signaling is involved in ECM remodeling. In summary, we provide the first genetic evidence that NMDAR plays an active role in platelet function and production. NMDAR regulates PPF through a mechanism that involves MK-ECM interaction and cytoskeletal reorganization. Our results suggest that NMDAR helps guide PPF in vivo. © 2022 by The American Society of Hematology.

Citation

James I Hearn, Taryn N Green, Colin L Hisey, Markus Bender, Emma C Josefsson, Nicholas Knowlton, Juliane Baumann, Raewyn C Poulsen, Stefan K Bohlander, Maggie L Kalev-Zylinska. Deletion of Grin1 in mouse megakaryocytes reveals NMDA receptor role in platelet function and proplatelet formation. Blood. 2022 Apr 28;139(17):2673-2690