Direct measurement of adenosine release during hypoxia in the CA1 region of the rat hippocampal slice
Direct measurement of adenosine release during hypoxia in the CA1 region of the rat hippocampal slice
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DOI：
10.1111/j.1469-7793.2000.00143.x
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发表时间：
2000-07-01
影响因子：
5.5
通讯作者：
Frenguelli, BG
中科院分区：
文献类型：
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作者：
Dale, N;Pearson, T;Frenguelli, BG
1.. 我们使用基于酶的双管传感器来测量大鼠海马切片 CA1 区域缺氧期间的腺苷释放，并结合兴奋性突触传递的同步细胞外场记录。2。当负载腺苷脱氨酶、核苷磷酸化酶和黄嘌呤氧化酶的组合时，传感器在10 nM 至20 mu M 的浓度范围内对外源腺苷呈线性响应。3。在没有酶的情况下，传感器放置在海马切片表面时，在缺氧期间记录了 40 +/- 43 pA 的非常小的净信号（平均值 +/- S.E.M.；n = 7）。只有当一个桶装载了完整的酶序列，而另一个桶装载了级联中的最后两个时，传感器才会在缺氧期间记录到大的净差异信号（1226 +/- 423 pA；n = 7）。4。该信号在缺氧期间逐渐增加，与同时记录的场兴奋性突触后电位的缺氧抑制成比例，并被辅福霉素 (0.5-2 μM) 大大降低 (67 +/- 6.5%; n = 9)，辅福霉素是腺苷脱氨酶的选择性抑制剂，腺苷脱氨酶是传感器内酶级联中的第一种酶。5。对于 5 分钟的缺氧事件，传感器记录的腺苷峰值浓度为 5.6 +/- 1.2 μM (n = 16)，抑制传输的 IC(50) 约为 3 μM.6。在名义上不含 Ca(2+) 的人工脑脊液中预孵育 3-6 小时的切片中，缺氧 5 分钟导致腺苷释放量增加约 9 倍（48.9 +/- 17.7 mu M；n = 6）。7。高细胞外Ca(2+) (4 mM) 既减少缺氧期间传感器记录的腺苷信号(3.5 +/- 0.6 muM; n = 4)，又延迟兴奋性突触传递的缺氧抑制。
1.. We have used an enzyme-based, twin-barrelled sensor -to measure adenosine release during hypoxia in the CA1 region of rat hippocampal slices in conjunction with simultaneous extracellular field recordings of excitatory synaptic transmission.2. When loaded with a combination of adenosine deaminase, nucleoside phosphorylase and xanthine oxidase, the sensor responded linearly to exogenous adenosine over the concentration range 10 nM to 20 mu M.3. Without enzymes, the sensor when placed on the surface of hippocampal slices recorded a very small net signal during hypoxia of 40 +/- 43 pA (mean +/- S.E.M.; n = 7). Only when one barrel was loaded with the complete sequence of enzymes and the other with the last two in the cascade did the sensor record a large net difference signal during hypoxia (1226 +/- 423 pA; n = 7).4. This signal increased progressively during the hypoxic episode, scaled with the hypoxic depression of the simultaneously recorded field excitatory postsynaptic potential and was greatly reduced (67 +/- 6.5%; n = 9) by coformycin (0.5-2 mu M), a selective inhibitor of adenosine deaminase, the first enzyme in the enzymic cascade within the sensor.5. For 5 min hypoxic episodes, the sensor recorded a peak concentration of adenosine of 5.6 +/- 1.2 mu M (n = 16) with an IC(50) for the depression of transmission of approximately 3 mu M.6. In slices pre-incubated for 3-6 h in nominally Ca(2+)-free artificial cerebrospinal fluid, 5 min of hypoxia resulted in an approximately 9-fold greater release of adenosine (48.9 +/- 17.7 mu M; n = 6).7. High extracellular Ca(2+) (4 mM) both reduced the adenosine signal recorded by the sensor during hypoxia (3.5 +/- 0.6 mu M; n = 4) and delayed the hypoxic depression of excitatory synaptic transmission.