Regulation of brain functions by Ca^<2+>/calmodulin-dependent protein kinase II activity

Ca^2+/钙调素依赖性蛋白激酶II（CaMKII）是中枢神经系统中最丰富的蛋白激酶之一，是细胞内Ca^2+响应各种刺激的关键介质。CaMKII参与许多神经元功能，包括突触可塑性和高级脑功能。然而，CaMKII的哪种蛋白质功能，即，蛋白激酶活性、钙调蛋白结合能力或与其他蛋白质相互作用的多聚体结构，是导致此类神经元功能的原因仍有待确定。为了明确CaMKII的蛋白激酶活性及其底物蛋白磷酸化的作用，我们通过用Arg-42替换Lys-42，用失活的CaMKII a亚基工程化敲入小鼠。正如预期的那样，CaMKII活性选择性降低，而CaMKII亚基蛋白水平与纯合突变体中野生型对照的水平相当。原位杂交显示CaMK Ⅱ亚基mRNA的正常表达模式，包括纯合突变体中CaMK Ⅱ α mRNA的树突状定位。使用急性海马脑片的电生理学分析和使用培养的海马神经元的细胞生物学研究揭示了突触可塑性异常，但在纯合突变体海马中的正常基础突触传递。此外，在这些小鼠中还观察到了海马依赖性学习缺陷。这些结果表明，CaMK Ⅱ α的蛋白激酶活性及其底物蛋白磷酸化对于维持正常海马功能是必需的。这种功能缺陷的CaMKIIα突变小鼠将作为一个强大的工具，在寻找活动依赖性突触可塑性的分子细节。

Ca^<2+>/calmodulin-dependent protein kinase II (CaMKII) is one of the most abundant protein kinase in the central nervous system and a key mediator of intracellular Ca^<2+> in response to various stimuli. CaMKII is involved in many neuronal functions including synaptic plasticity and higher brain functions. However, which protein function of CaMKII, i.e., protein kinase activity, calmodulin-binding capacity or multimeric structure interacting with other proteins, is responsible for such neuronal functions remains to be determined yet. To clarify specifically the role of protein kinase activity of CaMKII and its substrate protein phosphorylation, we engineered knock-in mice with the inactivated a subunit of CaMKII by replacing Lys-42 with Arg-42. As expected, CaMKII activity was selectively reduced, while CaMKII subunit protein levels were comparable to those of wild type controls in homozygous mutants. In situ hybridization revealed normal expression patterns of CaMKII subunit mRNAs, including dendritic localization of CaMKIIα mRNA in homozygous mutants. Electrophysiological analysis using acute hippocampal slices and cell biological study using cultured hippocampal neurons revealed abnormal synaptic plasticity but normal basal synaptic transmission in hippocampus in homozygous mutants. In addition, hippocampus-dependent learning deficit was also observed in these mice. These results indicate that protein kinase activity of CaMKIIα and its substrate protein phosphorylation are essential for maintaining normal hippocampal functions. Such a functionally deficient CaMKIIα mutant mouse will serve as a powerful tool in search for molecular details for activity-dependent synaptic plasticity.