Functional occurrence of the interaction of tissue plasminogen activator with the NR1 Subunit of N-methyl-D-aspartate receptors during stroke.
OBJECTIVE
Despite its fibrinolytic effect, tissue-type plasminogen activator (tPA) displays deleterious effects in the brain, including proexcitotoxicity, that can reduce the overall benefit from thrombolysis during stroke. We have proposed that tPA potentiates excitotoxicity by interacting with and cleaving the aminoterminal end of the NR1 subunit of N-methyl-d-aspartate receptors, leading to an increased calcium influx, Erk1/2 activation, and neurotoxicity. Because this mechanism is debated, our aim was to demonstrate its in vivo occurrence and relevance. Because tPA is released under ischemic conditions, we hypothesized that if it indeed processes NR1, then the released fragment should reactivate the immune system in animals that had been immunized long before with recombinant aminoterminal end of the NR1. This effect should be exacerbated in ischemic animals thrombolysed with recombinant tPA.


METHODS
At a time when specific antibodies could not be detected any longer, mice previously vaccinated with recombinant aminoterminal end of the NR1 were subjected to thromboembolic stroke induced by injecting thrombin in the middle cerebral artery alone or with intravenous thrombolysis.


RESULTS
Stroke performed 1 year after active immunization induced the reappearance of antibodies against the aminoterminal end of the NR1 in the plasma, an effect significantly increased when ischemia was followed by recombinant tPA-induced reperfusion. Moreover, immunization preventing the interaction of tPA with aminoterminal end of the NR1 reduced ischemic brain damages and extended the therapeutic window of tPA-induced thrombolysis.


CONCLUSIONS
We demonstrate that the tPA-dependent interaction and cleavage of the NR1 subunit of N-methyl-d-aspartate receptors occurs in vivo after stroke and that this interaction is a relevant therapeutic target for stroke treatment.
INSERM U919 serine proteases and pathophysiology of the neurovascular unit, UMR CNRS 6232 Cinaps, GIP Cyceron, Bd Becquerel, BP5229, 14074 Caen, France.
Richard
R
Macrez
Laurent
L
Bezin
Brigitte
B
Le Mauff
Carine
C
Ali
Denis
D
Vivien
D000818:Animals; D001323:Autoantibodies / Q000096:biosynthesis; D007114:Immunization; D007150:Immunohistochemistry; D051379:Mice; D009419:Nerve Tissue Proteins / Q000096:biosynthesis / Q000235:genetics; D016194:Receptors, N-Methyl-D-Aspartate / Q000235:genetics / Q000276:immunology / Q000502:physiology*; D011994:Recombinant Proteins / Q000096:biosynthesis / Q000235:genetics; D020521:Stroke / Q000188:drug therapy / Q000276:immunology / Q000503:physiopathology*; D013917:Thrombin / Q000494:pharmacology; D015912:Thrombolytic Therapy; D010959:Tissue Plasminogen Activator / Q000276:immunology / Q000494:pharmacology / Q000502:physiology*
D001323:Autoantibodies; C409466:NR1 NMDA receptor; D009419:Nerve Tissue Proteins; D016194:Receptors, N-Methyl-D-Aspartate; D011994:Recombinant Proteins; D013917:Thrombin; D010959:Tissue Plasminogen Activator