Activity-dependent CREB/ERK2 Activation are Reduced in Hippocalcin Deficient Mice

海马钙蛋白（HIP）是神经元钙感受器（NCS）家族的一员，主要表达于海马锥体细胞。我们通过同源重组产生了HIP缺陷小鼠。光镜下海马形态结构发育正常。场EPSC，成对脉冲易化和早期LTP检测HIP缺陷海马没有任何差异。但HIP缺陷小鼠的光辨别和Morris水迷宫学习能力受损。在本研究中，我们使用了由海马切片组成的半体内系统，并通过使用抗磷酸化Thr^ -<183>Tyr^<185>ERK 2和抗磷酸化Ser^<133>CREB抗体来评估Glu诱导的基因表达级联。NMDA处理导致野生型小鼠中ERK 2以时间和浓度依赖性方式的稳健的进行性磷酸化。在HEP缺陷型小鼠中，NMDA诱导的ERK 2磷酸化显著低于野生型小鼠。钙离子载体的作用类似于NMDA。相反，用蛋白激酶C-和cAMP依赖性蛋白激酶激活剂处理使磷酸化ERK 2的水平在野生型和HEP缺陷小鼠中均增加了相似的水平。NMDA治疗也增加了野生型小鼠磷酸化CREB的水平。已知ERK 2活化是CREB磷酸化的关键组分，CREB磷酸化启动c-fos mRNA合成。这些结果表明，海马钙素通过促进钙介导的ERK 2激活过程，促进活动依赖性基因表达，从而在长期记忆的巩固中发挥重要作用。

Hippocalcin (HIP) is a member of the neuronal calcium sensors (NCS) family predominantly expressed in the hippocampal pyramidal cells. We have generated HIP deficient mice by homologous recombination. The morphological structure of HIP deficient hippocampus developed normally in light microscopic levels. Field EPSC, paired pulse facilitation and early-phase LTP were detected without any differences in HIP deficient hippocampus. However, learning abilities of the light-discrimination and Morris water maze were impaired in HIP deficient mice. In the present study, we used a semi-in vivo system composed of hippocampal slices, and Gluinduced gene expression cascades were assessed by using anti-phospho-Thr^<183>-Tyr^<185> ERK2 and anti-phospho-Ser^<133> CREB antibodies. NMDA treatment led a robust, progressive phosphorylation of ERK2 in wild type mice in time and concentration dependent manners. In HEP deficient mice, the NMDA-induced ERK2 phosphorylation was significantly lower than that in wild type mice. Effect of calcium ionophore mimiced the NMDA. On the contrary, treatments with protein kinase C- and cAMP dependent protein kinase- activators increased levels of phosphorylated ERK2 by similar levels both in wild type and HEP deficient mice. NMDA treatment also increased levels of phosphorylated CREB in wild type mice. ERK2 activation is known to be a key component for CREB phosphorylation which initiates c-fos mRNA synthesis. The levels of NMDA-induced CREB phosphorylation were significantly lower in HIP deficient mice.These results indicate that hippocalcin facilitates the activity-dependent gene expression, which plays an essential role in consolidation of long-term memory, by promoting the calcium-mediated ERK2 activation process.