Dual expression of constitutively active Gα s -protein-coupled receptors differentially establishes the resting activity of the cAMP-gated HCN2 channel in a single compartment
Dual expression of constitutively active Gα s -protein-coupled receptors differentially establishes the resting activity of the cAMP-gated HCN2 channel in a single compartment
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组成型活性 Gα s 蛋白偶联受体的双重表达差异性地建立了单个隔室中 cAMP 门控 HCN2 通道的静息活性
DOI：
10.1016/j.bbrc.2017.10.082
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发表时间：
2017
影响因子：
3.1
通讯作者：
Kanamori Ryosuke
中科院分区：
文献类型：
--
作者：
Nakashima Noriyuki;Nakashima Kie;Nakayama Takeo;Takaku Akiko;Kanamori Ryosuke
超极化激活的环核苷酸门控2（HCN 2）通道是在神经系统中表达的HCN通道家族的主要亚型，其设定膜电位、调节细胞兴奋性并感知细胞外环境的变化。神经元表达各种Gα s蛋白偶联受体（GPCR），其中许多表现出配体非依赖性组成型活性。这些膜结合蛋白在神经元的各种亚细胞区室中表达。因此，一定比例的HCN 2通道响应于由组成型活性GPCR产生的基础cAMP池大小而打开。在这里，我们采用了外源性HEK 293表达系统和电压钳膜片钳记录研究基础HCN 2通道的活动存在下，两个GPCR不同的基础活动在一个单一的隔室。我们将β2-肾上腺素受体（β2AR）与气味受体（OR）一起使用，因为已知这两种GPCR家族均显示出较强的基础活性。因此，β 2 AR单独强烈增强HCN 2通道的活性，和OR的共表达进一步多样化的HCN 2通道的活性，这是完全取消的腺苷酸环化酶inhibitors.因此，我们的结论是，组成型活性GPCR的双重表达建立了不同范围的基础cAMP池大小通过相互添加或抑制的相互作用，即使在没有外部刺激的静息细胞。
The hyperpolarization-activated cyclic nucleotide-gated 2 (HCN2) channel is a major subtype of the HCN channel family expressed in the nervous system that sets the membrane potential, regulates cell excitability and senses changes in the extracellular environment. Neurons express various Gαs-protein-coupled receptors (GPCRs), many of which show ligand-independent constitutive activity. These membrane-bound proteins are expressed in various subcellular compartments of neurons. Therefore, some proportion of HCN2 channels opens in response to the basal cAMP pool size produced by constitutively active GPCRs. Here, we employed an exogenous HEK293 expression system and voltage-clamp patch-clamp recordings to investigate basal HCN2 channel activity in the presence of two GPCRs with diverse basal activities in a single compartment. We utilized the β2-adrenoceptor (β2AR) together with odorant receptors (ORs), as both GPCR families are known to show strong basal activity. Consequently, β2AR alone strongly enhanced the activity of HCN2 channels, and co-expression of ORs further diversified the HCN2 channel activity, which was totally abolished by an adenylate cyclase inhibitor.Thus, we conclude that the dual expression of constitutively active GPCRs establishes the diverse range of the basal cAMP pool size in resting cells through mutual additive or suppressive interactions, even in the absence of external stimulation.