Molecular actions of anesthetic drugs : Differential modulation by polymorphic forms of protein kinase C.

本研究是根据《自然》(自然，364：82,1993)报道麻醉药抑制中枢神经组织中PKC活性的论文制定的计划。我们首先尝试建立一种体外PKC检测体系，以分析麻醉药物的分子作用。由于PKC亚类在脑组织中的差异表达在其他地方也有报道，因此利用成熟的cDNA表达系统研究了PKC亚类对麻醉药物的反应。与我们的估计相反，在第一次体外实验中，我们不可能得到关于PKC活性的重复性结果。氟烷也不能抑制PKC的活性，尽管有报道说氟烷抑制了神经细胞中的PKC活性。我们的方法失败的可能原因有以下几点：1.体外实验系统和体内实验系统麻醉药物作用的差异：我们首次提到的Make a l…报告更多的研究表明，在与麻醉药物平衡的情况下，体内神经系统中的PKC活性受到抑制，报告没有显示药物对神经细胞中的PKC活性有直接影响。报告中的结果可能是对体内复杂神经网络的最终影响。体内和体外研究的差异可能是用于体外基因表达实验的COS7细胞没有发现PKC抑制的原因。2.药物敏感性和/或可能针对细胞谱系和/或物种的信号转导系统的差异：尽管有报道称麻醉药物抑制神经细胞的PKC活性，但我们研究的COS7细胞不是神经细胞，而是猴肾细胞。因此，大鼠/人神经细胞和猴肾细胞在细胞谱系和种类上可能存在很大差异。这种差异可能会在两个细胞系之间引入一些不同的细胞内信号或药物敏感性。从以上描述的可能性中，我们注意到我们的体外方法存在困难，然后我们也同时进行了体内方法。实际上，大鼠脑组织提取液中的PKC活性被检测到，这些大鼠被Nembutal、氟烷或七氟醚麻醉。然而，与以前的报道不同的是，我们在大鼠脑组织中没有发现氟烷对PKC活性的任何抑制。在我们的体内实验中，氟烷增强了大鼠脑内PKC的活性。而七氟醚对大鼠PKC活性有抑制作用。较少

This study was made a plan according to the paper (Nature, 364 : 82,1993) which reported that anesthetic drugs suppressed PKC activities in central nervous tissues. We tried at first to establish an in vitro PKC assay system in order to analyze molecular actions of anesthetic drugs. Since differential expressions of PKC subclasses in brain tissues were reported elsewhere, responsibility of PKC subclasses to anesthetic drugs were examined by using a well-established cDNA expression system. Contrary to our estimation, we could not have reproducible results on PKC activities in the first in vitro experiment. We also could not have any PKC suppression by halothane although it was reported that PKC activity in nerve cells was suppressed by halothane. The following factors are considered to possible reasons why our approach failed.1.Difference in anesthetic drug effects between on in vitro experimental system and on in vivo experimental system :The report which we first referred to make a pl … More an of this study showed that PKC activity was suppressed in vivo in nervous system equilibrated with anesthetic drugs and the report did not show that the drugs affected directly on PKC activity in nerve cells. It is possible that the results in the report were final effects on an in vivo complex neuron network. The difference between in vivo study and in vitro study may be the reason why no PKC suppression was found in COS7 cell lines which were used for in vitro cDNA expression experiment.2.Difference between drug sensitivity and/or signal transduction systems which may be specific to cell-lineage and/or species :Although the anesthetic drugs were reported to suppress PKC activities in nerve cells, the COS7 cell we used in this study is not a nerve cell but a monkey renal cell. Therefore, there could be much difference in cell-lineage and species between rat/human nerve cell and monkey renal cell. This difference might introduce some differential intracellular signaling or drug-sensitivity between two lineages.From those possibilities described above, we noticed a difficulty in our in vitro approach and then we performed an in vivo approach simultaneously too. Actually PKC activities were examined in brain tissue extracts from the rats which were anesthetized with nembutal, halothane, or sevoflurane. Unlike the previous report, however, we could not find any suppressed PKC activity by halothane in the rat brain tissues. Rat brain PKC activity in our in vivo experiment was enhanced by halothane. On the other hand, sevoflurane suppressed rat PKC activity. Less