Chronic adenosine A1R stimulation in hypoxia leads to persistent hippocampal synaptic depression, while unopposed adenosine A2AR receptor stimulation during hypoxia/reperfusion triggers adenosine-induced post-hypoxia synaptic potentiation (APSP) and increased neuronal death. Still, the mechanisms responsible for this adenosine-mediated neuronal damage following hypoxia need to be fully elucidated. We tested the hypothesis that A1R and A2AR regulation by protein kinase casein kinase 2 (CK2) and clathrin-dependent endocytosis of AMPARs both contribute to APSPs and neuronal damage. The APSPs following a 20-min hypoxia recorded from CA1 layer of rat hippocampal slices were abolished by A1R and A2AR antagonists and by broad-spectrum AMPAR antagonists. The inhibitor of GluA2 clathrin-mediated endocytosis Tat-GluA2-3Y peptide and the dynamin-dependent endocytosis inhibitor dynasore both significantly inhibited APSPs. The CK2 antagonist DRB also inhibited APSPs and, like hypoxic treatment, caused opposite regulation of A1R and A2AR surface expression. APSPs were abolished when calcium-permeable AMPAR (CP-AMPAR) antagonist (IEM or philanthotoxin) or non-competitive AMPAR antagonist perampanel was applied 5 min after hypoxia. In contrast, perampanel, but not CP-AMPAR antagonists, abolished APSPs when applied during hypoxia/reperfusion. To test for neuronal viability after hypoxia, propidium iodide staining revealed significant neuroprotection of hippocampal CA1 pyramidal neurons when pretreated with Tat-GluA2-3Y peptide, CK2 inhibitors, dynamin inhibitor, CP-AMPAR antagonists (applied 5 min after hypoxia), and perampanel (either at 5 min hypoxia onset or during APSP). These results suggest that the A1R-CK2-A2AR signaling pathway in hypoxia/reperfusion injury model mediates increased hippocampal synaptic transmission and neuronal damage via calcium-permeable AMPARs that can be targeted by perampanel for neuroprotective stroke therapy.

缺氧条件下的慢性腺苷 A1R 刺激导致持续的海马突触抑制，而缺氧/再灌注过程中无对抗的腺苷 A2AR 受体刺激会触发腺苷诱导的缺氧后突触增强 (APSP) 和神经元死亡增加。尽管如此，需要充分阐明导致缺氧后这种腺苷介导的神经元损伤的机制。我们测试了以下假设：蛋白激酶酪蛋白激酶 2 (CK2) 对 A1R 和 A2AR 的调节和 AMPAR 的网格蛋白依赖性内吞作用都有助于 APSP 和神经元损伤。从大鼠海马切片的 CA1 层记录的 20 分钟缺氧后的 APSP 被 A1R 和 A2AR 拮抗剂以及广谱 AMPAR 拮抗剂消除。GluA2 网格蛋白介导的内吞作用 Tat-GluA2-3Y 肽抑制剂和动力蛋白依赖性内吞作用抑制剂 dynasore 均显着抑制 APSP。CK2 拮抗剂 DRB 也抑制 APSP，并且像缺氧处理一样，引起 A1R 和 A2AR 表面表达的相反调节。当缺氧 5 分钟后应用钙渗透性 AMPAR (CP-AMPAR) 拮抗剂（IEM 或 philanthotoxin）或非竞争性 AMPAR 拮抗剂 perampanel 时，APSPs 被废除。相比之下，在缺氧/再灌注期间应用时，perampanel，但不是 CP-AMPAR 拮抗剂，消除了 APSP。为了测试缺氧后的神经元活力，碘化丙啶染色显示，当用 Tat-GluA2-3Y 肽、CK2 抑制剂、动力蛋白抑制剂、CP-AMPAR 拮抗剂（缺氧后 5 分钟应用）和吡仑帕奈（缺氧 5 分钟或 APSP 期间）。这些结果表明，缺氧/再灌注损伤模型中的 A1R-CK2-A2AR 信号通路通过渗透钙的 AMPARs 介导增加的海马突触传递和神经元损伤，perampanel 可以靶向这些 AMPARs 进行神经保护性卒中治疗。