Stark Neuroscience Research Institute, Indiana University of School of Medicine, Indianapolis, Indiana 46202, USA.
J Biol Chem. 2012 Mar 9;287(11):8495-506. doi: 10.1074/jbc.M111.323915. Epub 2012 Jan 17.
Aberrant glutamate and calcium signalings are neurotoxic to specific neuronal populations. Calcium/calmodulin-dependent kinase II (CaMKII), a multifunctional serine/threonine protein kinase in neurons, is believed to regulate neurotransmission and synaptic plasticity in response to calcium signaling produced by neuronal activity. Importantly, several CaMKII substrates control neuronal structure, excitability, and plasticity. Here, we demonstrate that CaMKII inhibition for >4 h using small molecule and peptide inhibitors induces apoptosis in cultured cortical neurons. The neuronal death produced by prolonged CaMKII inhibition is associated with an increase in TUNEL staining and caspase-3 cleavage and is blocked with the translation inhibitor cycloheximide. Thus, this neurotoxicity is consistent with apoptotic mechanisms, a conclusion that is further supported by dysregulated calcium signaling with CaMKII inhibition. CaMKII inhibitory peptides also enhance the number of action potentials generated by a ramp depolarization, suggesting increased neuronal excitability with a loss of CaMKII activity. Extracellular glutamate concentrations are augmented with prolonged inhibition of CaMKII. Enzymatic buffering of extracellular glutamate and antagonism of the NMDA subtype of glutamate receptors prevent the calcium dysregulation and neurotoxicity associated with prolonged CaMKII inhibition. However, in the absence of CaMKII inhibition, elevated glutamate levels do not induce neurotoxicity, suggesting that a combination of CaMKII inhibition and elevated extracellular glutamate levels results in neuronal death. In sum, the loss of CaMKII observed with multiple pathological states in the central nervous system, including epilepsy, brain trauma, and ischemia, likely exacerbates programmed cell death by sensitizing vulnerable neuronal populations to excitotoxic glutamate signaling and inducing an excitotoxic insult itself.
异常的谷氨酸和钙信号对特定神经元群体具有神经毒性。钙/钙调蛋白依赖性激酶 II（CaMKII）是神经元中的一种多功能丝氨酸/苏氨酸蛋白激酶，据信它可以调节神经递质传递和突触可塑性，以响应神经元活动产生的钙信号。重要的是，几种 CaMKII 底物控制神经元结构、兴奋性和可塑性。在这里，我们证明使用小分子和肽抑制剂超过 4 小时抑制 CaMKII 会诱导培养的皮质神经元凋亡。长时间 CaMKII 抑制产生的神经元死亡与 TUNEL 染色增加和 caspase-3 切割有关，并用翻译抑制剂环己酰亚胺阻断。因此，这种神经毒性与细胞凋亡机制一致，这一结论进一步得到 CaMKII 抑制时钙信号失调的支持。CaMKII 抑制肽还增强了斜坡去极化产生的动作电位数量，表明 CaMKII 活性丧失时神经元兴奋性增加。长时间抑制 CaMKII 会增加细胞外谷氨酸浓度。细胞外谷氨酸的酶缓冲作用和 NMDA 型谷氨酸受体的拮抗作用可防止与长时间 CaMKII 抑制相关的钙失调和神经毒性。然而，在没有 CaMKII 抑制的情况下，升高的谷氨酸水平不会引起神经毒性，这表明 CaMKII 抑制和升高的细胞外谷氨酸水平的组合导致神经元死亡。总之，在包括癫痫、脑外伤和缺血在内的中枢神经系统的多种病理状态下观察到的 CaMKII 丧失可能通过使易受伤害的神经元群体对兴奋性谷氨酸信号敏感并诱导兴奋性损伤本身，从而加重程序性细胞死亡。
