ANO1 电流参与了大鼠尾侧附睾 PDGFRα 阳性特化平滑肌细胞的起搏过程

IF 3.2 3区 生物学 Q3 CELL BIOLOGY Cell and Tissue Research Pub Date : 2024-04-08 DOI:10.1007/s00441-024-03890-x
Wataru Kudo, Retsu Mitsui, Hikaru Hashitani
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引用次数: 0

摘要

附睾管表现出自发性阶段性收缩(SPC),以储存和运输精子。在这里,我们探索了驱动尾侧附睾管 SPC 的起搏器细胞的分子鉴定,并以 ANO1 Ca2+ 激活的 Cl- 通道(CaCC)为重点,研究了 SPC 背后的起搏器电流特性。通过免疫组织化学方法观察了大鼠尾侧附睾管中血小板衍生生长因子受体α(PDGFRα)或ANO1阳性细胞的分布。酶切分离的附睾细胞采用穿孔全细胞膜片钳技术,SPCs则采用视频边缘跟踪技术记录。免疫组化显示,α-平滑肌肌动蛋白(α-SMA)阳性细胞分布在最内层的平滑肌层,同时表达PDGFRα和ANO1。大约三分之一的离体附睾细胞在保持电位-60 mV时表现出自发瞬态内向电流(STIC)。根据细胞内 Cl- 浓度的不同,STIC 的反向电位接近计算出的氯等效电位。ANO1特异性抑制剂Ani9(3 µM)可降低STIC的振幅和频率,而肌浆/内质网Ca2+-ATP酶(SERCA)抑制剂环哌嗪酸(CPA,30 µM)可消除STIC。Ani9(3 或 10 µM)降低了 SPCs 的频率,但不改变其振幅。因此,PDGFRα+、ANO1+特化平滑肌细胞(SMC)似乎可作为起搏器细胞,通过产生依赖 ANO1 的 STIC,以电驱动附睾 SPC。STIC 由细胞内 Ca2+ 储存库自发释放 Ca2+ 和 ANO1 随后打开导致去极化,去极化扩散到邻近的 SMC,在那里 L 型电压依赖性 Ca2+ 通道被激活,从而形成 SPC。
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Involvement of ANO1 currents in pacemaking of PDGFRα-positive specialised smooth muscle cells in rat caudal epididymis

The epididymal duct exhibits spontaneous phasic contractions (SPCs) to store and transport sperm. Here, we explored molecular identification of pacemaker cells driving SPCs in the caudal epididymal duct and also investigated properties of pacemaker currents underlying SPCs focusing on ANO1 Ca2+-activated Cl channels (CaCCs). Immunohistochemistry was performed to visualise the distribution of platelet-derived growth factor receptor α (PDGFRα)- or ANO1-positive cells in the rat caudal epididymal duct. Perforated whole-cell patch clamp technique was applied to enzymatically isolated epididymal cells, while SPCs were recorded with video edge-tracking technique. Immunohistochemistry revealed the distribution of α-smooth muscle actin (α-SMA)-positive cells co-expressing both PDGFRα and ANO1 in the innermost smooth muscle layer. Approximately one-third of isolated epididymis cells exhibited spontaneous transient inward currents (STICs) at the holding potential −60 mV. The reversal potential for STICs was close to the calculated chloride equivalent potential depending on intracellular Cl concentrations. Ani9 (3 µM), the ANO1 specific inhibitor, decreased both amplitude and frequency of STICs, while cyclopiazonic acid (CPA, 30 µM), a sarco-/endoplasmic reticulum Ca2+-ATPase (SERCA) inhibitor, abolished STICs. Ani9 (3 or 10 µM) reduced the frequency of SPCs without changing their amplitude. Thus, PDGFRα+, ANO1+ specialised smooth muscle cells (SMCs) appear to function as pacemaker cells to electrically drive epididymal SPCs by generating ANO1-dependnet STICs. STICs arising from spontaneous Ca2+ release from intracellular Ca2+ store and subsequent opening of ANO1 result in depolarisations that spread into adjacent SMCs where L-type voltage-dependent Ca2+ channels are activated to develop SPCs.

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来源期刊
Cell and Tissue Research
Cell and Tissue Research 生物-细胞生物学
CiteScore
7.00
自引率
2.80%
发文量
142
审稿时长
1 months
期刊介绍: The journal publishes regular articles and reviews in the areas of molecular, cell, and supracellular biology. In particular, the journal intends to provide a forum for publishing data that analyze the supracellular, integrative actions of gene products and their impact on the formation of tissue structure and function. Submission of papers with an emphasis on structure-function relationships as revealed by recombinant molecular technologies is especially encouraged. Areas of research with a long-standing tradition of publishing in Cell & Tissue Research include: - neurobiology - neuroendocrinology - endocrinology - reproductive biology - skeletal and immune systems - development - stem cells - muscle biology.
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