Zamzow Christina R, Xiong Wei, Parkinson Fiona E
Department of Pharmacology and Therapeutics, University of Manitoba, Winnipeg, Manitoba, Canada.
J Neurosci Res. 2008 Sep;86(12):2641-9. doi: 10.1002/jnr.21718.
Adenosine (ADO) is produced by cultured neurons and astrocytes, albeit by different pathways, during in vitro stroke models (Parkinson and Xiong [2004] J. Neurochem. 88:1305-1312). Expression of ecto-5' nucleotidase (e-N), the enzyme responsible for extracellular dephosphorylation of AMP to ADO, is more abundant in astrocytes than neurons. Therefore, we tested the hypothesis that N-methyl-D-aspartate (NMDA) evokes ADO release per se from neurons, whereas dephosphorylation of extracellular adenine nucleotides contributes to NMDA-evoked ADO production in the presence of astrocytes. We used four different cell preparations-cortical rat neurons, cortical rat astrocytes, cocultures of neurons and astrocytes, and transient cocultures of neurons with astrocytes on transwell filters-to show that astrocytes contribute to NMDA-evoked increases in extracellular ADO. NMDA significantly increased ADO and inosine (INO) production from cultured cortical neurons but only increased extracellular INO production from cocultures. In neurons, the equilibrative nucleoside transport (ENT) inhibitor dipyridamole (DPR) prevented NMDA-evoked ADO and INO production, whereas the e-N inhibitor alpha,beta-methylene ADP (AOPCP) had no effect. Conversely, from both cocultures and transient cocultures DPR significantly decreased NMDA-evoked INO but not ADO generation. AOPCP inhibited NMDA-evoked production of both ADO and INO from transient cocultures. In the absence of astrocytes, NMDA evoked release of intracellular ADO and INO from cultured cortical neurons through ENT. However, in the presence of astrocytes, extracellular conversion of adenine nucleotides to ADO contributed significantly to NMDA-evoked production of this purine.
在体外中风模型中（帕金森和熊[2004]《神经化学杂志》88:1305 - 1312），腺苷（ADO）由培养的神经元和星形胶质细胞产生，尽管产生途径不同。胞外5'核苷酸酶（e - N）负责将AMP细胞外去磷酸化为ADO，其在星形胶质细胞中的表达比神经元中更丰富。因此，我们检验了以下假设：N - 甲基 - D - 天冬氨酸（NMDA）本身会引起神经元释放ADO，而在有星形胶质细胞存在时，细胞外腺嘌呤核苷酸的去磷酸化有助于NMDA诱导的ADO产生。我们使用了四种不同的细胞制剂——大鼠皮质神经元、大鼠皮质星形胶质细胞、神经元与星形胶质细胞的共培养物以及神经元与星形胶质细胞在Transwell滤器上的瞬时共培养物——来表明星形胶质细胞有助于NMDA诱导的细胞外ADO增加。NMDA显著增加了培养的皮质神经元中ADO和肌苷（INO）的产生，但仅增加了共培养物中细胞外INO的产生。在神经元中，平衡核苷转运（ENT）抑制剂双嘧达莫（DPR）可阻止NMDA诱导的ADO和INO产生，而e - N抑制剂α,β - 亚甲基ADP（AOPCP）则没有作用。相反，在共培养物和瞬时共培养物中，DPR显著降低了NMDA诱导的INO产生，但不影响ADO的生成。AOPCP抑制了瞬时共培养物中NMDA诱导的ADO和INO产生。在没有星形胶质细胞的情况下，NMDA通过ENT引起培养的皮质神经元释放细胞内ADO和INO。然而，在有星形胶质细胞存在时，腺嘌呤核苷酸向ADO的细胞外转化对NMDA诱导的这种嘌呤的产生有显著贡献。
