Cellular mechanisms of dentin sensation by odontoblast as senory receptor cell(expression of TRP and voltage-dependent K^+ channels, and analysis by odontoblast-neuron co-culture system)

成牙本质细胞作为感觉受体细胞的牙本质感觉的细胞机制（TRP和电压依赖性K^通道的表达，以及成牙本质细胞-神经元共培养系统的分析）

• 批准号: 18592050
• 负责人: SHIBUKAWA Yoshiyuki
• 金额: $ 2.36万
• 依托单位: Tokyo Dental College
• 依托单位国家: 日本
• 项目类别: Grant-in-Aid for Scientific Research (C)
• 财政年份: 2006
• 资助国家: 日本
• 起止时间: 2006 至 2007
• 项目状态: 已结题
• 来源: https://kaken.nii.ac.jp/grant/KAKENHI-PROJECT-18592050/
• 关键词: Dentistry; Signal Transduction; Neuroscience; Oral Physiology; Dental Pulp; Odontoblast; TRP(transient receptor notential)Channels; Na^+; Ca^<2+> exchanger(NCX); TRP(transient recetor potential; Ca^<2+> exchanger (NCX); transient receptor poten

This study aimed to clarify the role of odontoblasts in cellular mechanisms of dentin sensation. During the research period from April 2006 to March 2008, we investigated expression of TRPV1 (vanilloid receptor 1; VR1) channel (a transient receptor potential (TRP) family subtype, which contributes essentially for the detection of pain sensation), voltage-dependent K+ channels (Kv channles; which involve in the generation of action potential), and Na^+-Ca^<2+> exchangers (NCX; which regulate ntracellular Ca^<2+> concentartion ([Ca^<2+>]_i) by extrusion of [Ca^<2+>]_i)in rat odontoblasts by immunohistochemical, intracellular Ca^<2+> concentartion measurement and whole-cell patch-clamp technique.1. TRPV1 channels: Immunohistochemical experiments showed localization of TRPV1 on the distal regions of odontoblast membranes. RT-PCR analysis showed that odontoblasts express TRPV subfamily 1, 2, 3. In the fura-2 fluorescence analysis to measure[Ca^<2+>]I, anandamide (AEA; 10 pM; endogenous TRPV … More 1 activator) evoked a transient rise in [Ca^<2+>], in the presence of extracellular Ca^<2+> with slight delay in activation of transient rise after AEA application. However, in the absence of extracellular Ca^<2+>, we could not observe any transient rise in [Ca^<2+>], indicating that AEA activates Ca^<2+> influx. The Ca^<2+> influx was blocked by a TRPV1 channel antagonist, capsazepine. In our previous study, that extracellular application of capsaicin activated inward currents in rat odontoblast (Okumura, et. Al., 2006), but their current amplitude was small (ca, 10 pA). In addition, in the present experiments, rise in [Ca^<2+>], after AEA application accompanied with delay time (ca, 2-3 min)in activation. It has been reported that capsaicin as well as AEA binds intracellular domain of TRPV1 channels. Therefore, we examine whether or not intracellular application of capsaicin activate inward currents in odontoblasts. Under the continuous voltage-clamp condition with holding potential of 0 mV, an intracellular application of 10 μM capsaicin via patch-pipette elicited inward currents, showing transient activation followed by current decaying. Therefore, these results indicate that odontoblasts express functional TRPV1 channels. 2. Kv channels: In acutely isolated odontoblasts, depolarizing steps from a holding potential of -80mV evoked time- and voltage-dependent outward currents. The relatively slow activation kinetics exhibited strong dependence on the membrane potential. Time constant of inactivation was approximately 400 ms at membrane potential of -50 mV. These results indicate that odontoblasts express slowly-activating and -inactivating time- and voltage-dependent K+ currents. 3. NCX: The reverse exchange Ca^<2+>influx in odontoblasts was blocked by an NCX inhibitor (KB-R7943 and SEA0400) in a concentration-dependent manner. The inward currents via forward Na^+-Ca^<2+>exchange had a dependence on external Na+. Immunohistochemical localization of NCX was detected at distal membrane of odontoblasts.These results indicate that odontblasts possess NCX. Our research results indicate significant expression of TRPV1, Kvchannels and NCX in odontoblasts, suggesting that odontoblasts may sirectly respond to the noxious stimuli, and generate action potential by coupling with voltage-dependent Na^+ channels. NCX play an important role in the regulation of intracellular Ca^<2+> levels by excessive internal Ca^<2+>, as well as in Ca-<2+> transport pathway to the dentin mineralizing front by transporting increased intracellular Ca^<2+> as a result of activation of TRPV1. Less

这项研究旨在阐明牙本质感觉的细胞机制中的odontoblasts。依赖性K+通道F动作电位）和Na^+-ca^<2+>交换器（NCX;，它通过挤出[CA ^<2+>]由免疫组织化学，细胞内CA^<2+> tartion蛋白斑点和全细胞斑块技术。1在Fura-2荧光分析中，ODONTOBLAST trPV亚家族1、2、3。用于测量[Ca ^<2+>] I，Anandamide（AEA; AEA; 10 pm; 10 pm;更多1个激活剂）引起了[Ca ^<2 +>] <2+>在没有细胞外Ca^<2+> <2+>的情况下，aightee aea的激活略有延迟<2+>在我们的Providence研究中，TRPV1通道拮抗剂（capsazepine）在大鼠Odontoblast中被激活。此外，在目前的实验中，激活中的[Ca^<2+] In升高。通过贴片的夹具为10μm的夹具，通过斑块向内流动，瞬时腐烂，cate od腐蛋白在-50 mV的膜电位上，缓慢的激活动力学对iNactivaon的时间偏见约为400毫秒。 con依的方式依赖于前进的na^+-ca^<2+>交换对外部Na+的依赖性。 trpv1，kvchannels和ncx在odontoblasts中的表达，对有害的刺激Na^+通道响应，在调节内部CA^<2+>水平中起重要作用。 ，通过转运TRPV1的细胞内矿物质矿物质的运输途径

项目成果

Cerebral cortical dysfunction in patients with temporomandibular disorders in association with jaw movement observation

• DOI: 10.1016/j.pain.2006.10.006
• 发表时间: 2007-03
• 期刊: Pain
• 影响因子: 7.4
• 作者: Y. Shibukawa;T. Ishikawa;Y. Kato;Zhen-kang Zhang;Ting Jiang;M. Shintani;Masaki Shimono;T. Kumai;Takashi Suzuki;Motoichiro Kato;Yoshio Nakamura

Ca2+ signaling in odontoblasts:role in dentinogenesis and sensory transduction

成牙本质细胞中的 Ca2 信号传导：在牙本质发生和感觉转导中的作用

• 发表时间: 2007
• 作者: 新谷 益郎;他;渋川 義幸;Shibukawa Y.
• 通讯作者: Shibukawa Y.

Aberrant component in high frequency range relating mirror neuron dysfunction in schizophrenia patients.

高频范围内的异常成分与精神分裂症患者的镜像神经元功能障碍有关。

• 发表时间: 2007
• 作者: Kato;M;Kato;Y;Shibukawa;Y;Shintani;M
• 通讯作者: M

Somatosensory evoked magnetic fields(SEFs) from buccal and tongue mucosa using piezo-driven tactile stimulation device : a magnetoencephalography study

使用压电驱动触觉刺激装置从颊和舌粘膜产生体感诱发磁场（SEF）：脑磁图研究

• 发表时间: 2007
• 作者: Tamura;Y;Kubo;K;Shintani;M;Tazaki;M;Shibukawa;Y;Ichinohe;T
• 通讯作者: T

エナメル芽細胞の直接的なCa2+輸送はNCXI,3によって仲介される

成釉细胞中的直接 Ca2+ 转运由 NCXI 介导，3

• 发表时间: 2006
• 作者: 奥村 礼二郎;ら
• 通讯作者: ら