Effects of anesthesia on immunohistochemical detection of phosphorylated extracellular signal-regulated kinase in cerebral cortex
Effects of anesthesia on immunohistochemical detection of phosphorylated extracellular signal-regulated kinase in cerebral cortex
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DOI：
10.1016/j.jneumeth.2008.01.005
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发表时间：
2008-05
影响因子：
3
通讯作者：
Hironori Takamura;S. Ichisaka;Kana Watanabe;Masahito Toigawa;Y. Hata
中科院分区：
文献类型：
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作者：
Hironori Takamura;S. Ichisaka;Kana Watanabe;Masahito Toigawa;Y. Hata
在尝试用化学方法检测大脑皮层细胞外信号调节激酶（ERK）磷酸化状态的过程中，我们确定了深度麻醉安乐死是否会干扰ERK的磷酸化状态，因为麻醉可能会影响ERK的活性依赖性磷酸化。本文比较了戊巴比妥钠短（2和5min）和长（>10min）麻醉对大鼠视皮层和内嗅皮层磷酸化ERK免疫反应性的影响。长时间麻醉使磷酸化ERK免疫阳性细胞的密度急剧降低至短时间麻醉条件下的约15%。在所有皮质区域均观察到减少。pERK免疫反应性在2和5min组之间无显著差异。异氟醚长时间麻醉诱导了相似程度的降低。即使给予相似的麻醉持续时间，免疫组织化学结果也可能包含由于对麻醉剂敏感性的个体差异而引起的变化。我们证明，pERK免疫阳性细胞密度在视觉皮层的变化显着减少，通过标准化的值在非视觉区的内嗅皮层的密度，从而使我们能够检测动物之间的差异，在不同的视觉条件下，具有更高的灵敏度。因此，可以通过计算感兴趣区域的免疫反应性与作为参考的其他皮质区域的免疫反应性的比率来减少差异。
During attempts to examine the phosphorylation status of extracellular signal-regulated kinase (ERK) in cerebral cortex immunohistochemically, we determined whether deep anesthesia for euthanasia disturbs the phosphorylation status of ERK, because the anesthesia might influence activity-dependent phosphorylation of ERK. We compared effects of short (2 and 5min) and long (>10min) anesthesia by pentobarbital on the immunoreactivity for phosphorylated ERK in the visual and entorhinal cortices of rat. The long anesthesia drastically reduced the density of phosphorylated ERK immunopositive cells to about 15% of the short anesthesia condition. The reduction was observed in all cortical regions. We found no significant difference in pERK immunoreactivity between 2 and 5min groups. A reduction of similar degree was induced by long anesthesia with isoflurane. Even if a similar duration of anesthesia is given, the immunohistochemical results possibly contain a variation due to the individual difference in the sensitivity to the anesthetics. We demonstrated that the variation of pERK immunopositive cell density in the visual cortex was significantly reduced by normalizing the values to the density in the nonvisual area in the entorhinal cortex, thus enabling us to detect differences between animals under different visual conditions with higher sensitivity. Therefore, the variation could be reduced by calculating the ratio of immunoreactivity in the area of interest to that in other cortical area as reference.