Ca2+-activated Cl- channel currents in mammary secretory cells from lactating mouse
Ca2+-activated Cl- channel currents in mammary secretory cells from lactating mouse
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DOI：
10.1152/ajpcell.00050.2016
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发表时间：
2016-11-01
影响因子：
5.5
通讯作者：
Ishikawa, Toru
中科院分区：
文献类型：
--
作者：
Kamikawa, Akihiro;Ichii, Osamu;Ishikawa, Toru
通过Ca2+激活的Cl-通道（CaCC）的Cl-分泌对于唾液腺等外分泌腺的液体分泌至关重要。同样在乳腺中，已经假设CaCC在乳的Cl-和水相的分泌中起重要作用。然而，没有证据表明哺乳动物的天然乳腺分泌（MS）细胞中的CaCC的功能表达。因此，我们使用全细胞膜片钳技术评估了从哺乳期小鼠新鲜分离的MS细胞中的膜电流。在MS细胞中，我们检测到的CaCC电流具有以下特征：1）在亚微摩尔浓度范围内的Ca~（2+）依赖性激活; 2）电压依赖性激活; 3）激活和失活的缓慢动力学; 4）稳态电流的外向整流; 5）阴离子渗透性的顺序为I-> NO_3-> Br-> Cl->>谷氨酸; 6）Cl-通道阻断剂（尼氟灭酸、DIDS和CaCCinh-A01）的抑制。天然CaCC电流的这些特征类似于异源表达的TMEM16A的报道特征。RT-PCR分析显示多个CaCC通道包括TMEM16A、Best1和Best3在哺乳期小鼠乳腺中表达。免疫组织化学染色显示TMEM16A蛋白定位于MS细胞的顶膜。总的来说，我们的数据有力地表明，MS细胞功能性表达CaCC，其至少部分由TMEM16A构成。MS细胞顶膜上的CaCC如TMEM16A可能影响乳汁的数量和/或质量。
The Cl- secretion via Ca2+-activated Cl- channel (CaCC) is critical for fluid secretion in exocrine glands like the salivary gland. Also in the mammary gland, it has been hypothesized that CaCC plays an important role in the secretion of Cl- and aqueous phase of milk. However, there has been no evidence for the functional expression of CaCC in native mammary secretory (MS) cells of lactating animals. We therefore assessed membrane current in MS cells that were freshly isolated from lactating mice using whole cell patch-clamp techniques. In MS cells, we detected CaCC current that exhibited the following characteristics: 1) Ca2+-dependent activation at the concentrations of sub-micromolar range; 2) voltage-dependent activation; 3) slow kinetics for activation and deactivation; 4) outward rectification of the steady-state current; 5) anion permeability in the sequence of I- > NO3- > Br- > Cl- >> glutamate; 6) inhibition by Cl- channel blockers (niflumic acid, DIDS, and CaCCinh-A01). These characteristics of native CaCC current were similar to reported characteristics of heterologously expressed TMEM16A. RT-PCR analyses showed the expression of multiple CaCC channels including TMEM16A, Best1, and Best3 in the mammary glands of lactating mice. Immuno-histochemical staining revealed the localization of TMEM16A protein at the apical membrane of the MS cells. Collectively, our data strongly suggest that MS cells functionally express CaCC, which is at least partly constituted by TMEM16A. The CaCC such as TMEM16A at the apical membrane of the MS cells may influence the quantity and/ or quality of milk.