Department of Pharmacology and Therapeutics, University of Manitoba, Winnipeg, Manitoba, Canada.
PLoS One. 2012;7(6):e39772. doi: 10.1371/journal.pone.0039772. Epub 2012 Jun 25.
Adenosine, through activation of its A(1) receptors, has neuroprotective effects during hypoxia and ischemia. Recently, using transgenic mice with neuronal expression of human equilibrative nucleoside transporter 1 (hENT1), we reported that nucleoside transporter-mediated release of adenosine from neurons was not a key mechanism facilitating the actions of adenosine at A(1) receptors during hypoxia/ischemia. The present study was performed to test the importance of CD73 (ecto-5'-nucleotidase) for basal and hypoxic/ischemic adenosine production. Hippocampal slice electrophysiology was performed with CD73(+/+) and CD73(-/-) mice. Adenosine and ATP had similar inhibitory effects in both genotypes, with IC(50) values of approximately 25 µM. In contrast, ATP was a less potent inhibitor (IC(50) = 100 µM) in slices from mice expressing hENT1 in neurons. The inhibitory effects of ATP in CD73(+/+) and CD73(-/-) slices were blocked by the adenosine A(1) receptor antagonist 8-cyclopentyl-1,3-dipropylxanthine (DPCPX) and were enhanced by the nucleoside transport inhibitor S-(4-nitrobenzyl)-6-thioinosine (NBTI), consistent with effects that are mediated by adenosine after metabolism of ATP. AMP showed a similar inhibitory effect to ATP and adenosine, indicating that the response to ATP was not mediated by P2 receptors. In comparing CD73(-/-) and CD73(+/+) slices, hypoxia and oxygen-glucose deprivation produced similar depression of synaptic transmission in both genotypes. An inhibitor of tissue non-specific alkaline phosphatase (TNAP) was found to attenuate the inhibitory effects of AMP and ATP, increase basal synaptic activity and reduce responses to oxygen-glucose deprivation selectively in slices from CD73(-/-) mice. These results do not support an important role for CD73 in the formation of adenosine in the CA1 area of the hippocampus during basal, hypoxic or ischemic conditions, but instead point to TNAP as a potential source of extracellular adenosine when CD73 is absent.
腺苷通过激活其 A(1)受体，在缺氧和缺血期间具有神经保护作用。最近，使用神经元表达人平衡核苷转运蛋白 1 (hENT1)的转基因小鼠，我们报道了核苷转运蛋白介导的神经元中腺苷的释放不是促进缺氧/缺血期间腺苷在 A(1)受体上作用的关键机制。本研究旨在测试 CD73（外核苷酸酶）对基础和缺氧/缺血时腺苷产生的重要性。使用 CD73(+/+)和 CD73(-/-)小鼠进行海马切片电生理学研究。在两种基因型中，腺苷和 ATP 具有相似的抑制作用，IC(50)值约为 25 µM。相比之下，在表达神经元中 hENT1 的小鼠的切片中，ATP 的抑制作用较弱（IC(50) = 100 µM）。ATP 在 CD73(+/+)和 CD73(-/-)切片中的抑制作用被腺苷 A(1)受体拮抗剂 8-环戊基-1,3-二丙基黄嘌呤 (DPCPX) 阻断，并被核苷转运抑制剂 S-(4-硝基苄基)-6-硫代肌苷 (NBTI) 增强，这与代谢 ATP 后由腺苷介导的作用一致。AMP 表现出与 ATP 和腺苷相似的抑制作用，表明对 ATP 的反应不是由 P2 受体介导的。在比较 CD73(-/-)和 CD73(+/+)切片时，缺氧和氧葡萄糖剥夺在两种基因型中均产生相似的突触传递抑制。发现组织非特异性碱性磷酸酶 (TNAP) 的抑制剂可减弱 AMP 和 ATP 的抑制作用，增加基础突触活性，并选择性地减少 CD73(-/-)小鼠切片中对氧葡萄糖剥夺的反应。这些结果不支持 CD73 在基础、缺氧或缺血条件下在海马 CA1 区形成腺苷中起重要作用，但表明在没有 CD73 时，TNAP 可能是细胞外腺苷的潜在来源。
