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

本研究是根据Nature， 364: 82,1993报道麻醉药物抑制中枢神经组织PKC活性的论文制定的计划。为了分析麻醉药物的分子作用，我们首先尝试建立体外PKC检测系统。由于PKC亚类在脑组织中的差异表达在其他地方被报道，PKC亚类对麻醉药物的作用通过一个完善的cDNA表达系统来研究。与我们的估计相反，我们无法在第一次体外实验中获得可重复的PKC活性结果。尽管有报道称神经细胞中的PKC活性被氟烷抑制，但我们也没有发现氟烷对PKC的抑制作用。以下因素被认为是我们的方法失败的可能原因。体外实验系统与体内实验系统麻醉药物作用的差异：我们首次引用的报告进一步表明，在体内麻醉药物平衡的神经系统中PKC活性受到抑制，但报告并未表明药物直接影响神经细胞PKC活性。报告中的结果可能是对体内复杂神经元网络的最终影响。用于体外cDNA表达实验的COS7细胞系未发现PKC抑制的原因可能是体内研究与体外研究的差异。药物敏感性和/或信号转导系统之间的差异可能是细胞谱系和/或物种特异性的：尽管有报道称麻醉药物可以抑制神经细胞中的PKC活性，但我们在本研究中使用的COS7细胞不是神经细胞，而是猴子肾细胞。因此，大鼠/人神经细胞与猴肾细胞在细胞系和种类上可能存在很大差异。这种差异可能会在两个谱系之间引入一些不同的细胞内信号传导或药物敏感性。从上述可能性中，我们注意到体外方法的困难，然后我们也同时进行了体内方法。实际上，PKC的活性是在被麻醉的大鼠的脑组织提取物中检测的，这些大鼠分别被麻醉了镇静剂、氟烷和七氟烷。然而，与之前的报道不同，我们没有发现氟烷在大鼠脑组织中抑制PKC活性。在我们的体内实验中，氟烷增强了大鼠脑PKC的活性。另一方面，七氟醚抑制大鼠PKC活性。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