Journal
JOURNAL OF CEREBRAL BLOOD FLOW AND METABOLISM
Volume 32, Issue 1, Pages E1-E7Publisher
NATURE PUBLISHING GROUP
DOI: 10.1038/jcbfm.2011.142
Keywords
adenosine; astrocytes; brain ischemia; cell culture; energy metabolism
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Funding
- NIGMS NIH HHS [T32 GM007753] Funding Source: Medline
- NINDS NIH HHS [R01 NS078167, R01 NS075177, P01 NS050315] Funding Source: Medline
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Recent reports based on a chemiluminescent enzymatic assay for detection of adenosine conclude that cultured astrocytes release adenosine during mildly hypoxic conditions. If so, astrocytes may suppress neural activity in early stages of hypoxia. The aim of this study was to reevaluate the observation using high-performance liquid chromatography (HPLC). The HPLC analysis showed that exposure to 20 or 120 minutes of mild hypoxia failed to increase release of adenosine triphosphate (ATP), adenosine diphosphate (ADP), adenosine monophosphate (AMP), and adenosine from cultured astrocytes. Similar results were obtained using a chemiluminescent enzymatic assay. Moreover, since the chemiluminescent enzymatic assay relies on hydrogen peroxide generation, release of free-radical scavengers from hypoxic cells can interfere with the assay. Accordingly, adenosine added to samples collected from hypoxic cultures could not be detected using the chemiluminescent enzymatic assay. Furthermore, addition of free-radical scavengers sharply reduced the sensitivity of adenosine detection. Conversely, use of a single-step assay inflated measured values due to the inability of the assay to distinguish adenosine and its metabolite inosine. These results show that cultured astrocytes do not release adenosine during mild hypoxia, an observation consistent with their high resistance to hypoxia. Journal of Cerebral Blood Flow & Metabolism (2012) 32; doi:10.1038/jcbfm.2011.142; published online 12 October 2011
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