Ontogenic Changes and Differential Localization of T-type Ca(2+) Channel Subunits Cav3.1 and Cav3.2 in Mouse Hippocampus and Cerebellum.
Ontogenic Changes and Differential Localization of T-type Ca(2+) Channel Subunits Cav3.1 and Cav3.2 in Mouse Hippocampus and Cerebellum.
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DOI：
10.3389/fnana.2016.00083
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发表时间：
2016
影响因子：
2.9
通讯作者：
Luján R
中科院分区：
文献类型：
--
作者：
Aguado C;García-Madrona S;Gil-Minguez M;Luján R
T型钙通道在调节脑膜兴奋性方面起着重要作用。虽然T型电流对神经元输出的贡献通常被认为反映了T型通道亚型在躯体-树突间的不同分布，但它们在中枢神经元中的准确亚细胞分布并不完全确定。应用组织印迹技术和高分辨免疫电子显微镜技术，研究了T型Cav3.1和Cav3.2通道亚单位在成人脑中的表达、区域分布和亚细胞定位，以及出生后发育过程中表达的个体发育。组织印迹分析表明，Cav3.1和Cav3.2蛋白在大脑中广泛表达，且大多不重叠。Cav3.1在小脑分子层表达水平最高，Cav3.2在海马区和小脑分子层表达最强。在发育过程中，Cav3.1和Cav3.2的表达水平随着年龄的增长而增加，尽管它们的表达存在明显的地区和发育阶段差异。在细胞和亚细胞水平，免疫电子显微镜显示Cav3.1存在于海马区中间神经元和浦肯野细胞(PC)的体树突域，而Cav3.2存在于CA1锥体细胞、海马区中间神经元和PC的体树突域。Cav3.1和Cav3.2的免疫颗粒大多与细胞膜或细胞内膜相关，不同的隔室有显著差异。因此，Cav3.1主要定位于中间神经元的质膜，而Cav3.2主要定位于树突棘的质膜，主要分布在树突的胞内。在PC中，Cav3.1和Cav3.2显示出相似的分布模式。除了主要的突触后分布外，在建立兴奋性突触的轴突终末中也检测到Cav3.2而不是Cav3.1。这些结果为T型通道亚基的亚细胞定位提供了新的证据，并为Cav3.1和Cav3.2亚单位在中枢神经元质膜上的不均匀分布提供了证据，这可能解释了T型介导电流的功能异质性。
T-type calcium (Ca2+) channels play a central role in regulating membrane excitability in the brain. Although the contributions of T-type current to neuron output is often proposed to reflect a differential distribution of T-type channel subtypes to somato-dendritic compartments, their precise subcellular distributions in central neurons are not fully determined. Using histoblot and high-resolution immunoelectron microscopic techniques, we have investigated the expression, regional distribution and subcellular localization of T-type Cav3.1 and Cav3.2 channel subunits in the adult brain, as well as the ontogeny of expression during postnatal development. Histoblot analysis showed that Cav3.1 and Cav3.2 proteins were widely expressed in the brain, with mostly non-overlapping patterns. Cav3.1 showed the highest expression level in the molecular layer (ml) of the cerebellum (Cb), and Cav3.2 in the hippocampus (Hp) and the ml of Cb. During development, levels of Cav3.1 and Cav3.2 increased with age, although there were marked region- and developmental stage-specific differences in their expression. At the cellular and subcellular level, immunoelectron microscopy showed that labeling for Cav3.1 was present in somato-dendritic domains of hippocampal interneurons and Purkinje cells (PCs), while Cav3.2 was present in somato-dendritic domains of CA1 pyramidal cells, hippocampal interneurons and PCs. Most of the immunoparticles for Cav3.1 and Cav3.2 were either associated with the plasma membrane or the intracellular membranes, with notable differences depending on the compartment. Thus, Cav3.1 was mainly located in the plasma membrane of interneurons, whereas Cav3.2 was mainly located in the plasma membrane of dendritic spines and had a major intracellular distribution in dendritic shafts. In PCs, Cav3.1 and Cav3.2 showed similar distribution patterns. In addition to its main postsynaptic distribution, Cav3.2 but not Cav3.1 was also detected in axon terminals establishing excitatory synapses. These results shed new light on the subcellular localization of T-type channel subunits and provide evidence for the non-uniform distribution of Cav3.1 and Cav3.2 subunits over the plasma membrane of central neurons, which may account for the functional heterogeneity of T-type mediated current.