Ikeda Ryo, Ling Jennifer, Cha Myeounghoon, Gu Jianguo G
Department of Anesthesiology and Perioperative Medicine, College of Medicine, University of Alabama at Birmingham, 901 19TH Street South, BMR II 210, Birmingham, AL, 35294, USA.
Department of Orthopaedic Surgery, Jikei University School of Medicine, 3-25-8 Nishi-Shinbashi, Minato-ku, Tokyo, 105-8461, Japan.
Mol Pain. 2015 Apr 25;11:23. doi: 10.1186/s12990-015-0022-5.
Mammals use tactile end-organs to perform sensory tasks such as environmental exploration, social interaction, and tactile discrimination. However, cellular and molecular mechanisms underlying tactile transduction in tactile end-organs remain poorly understood. The patch-clamp recording technique may be the most valuable approach for detecting and studying tactile transduction in tactile end-organs, but it is technically challenging because tactile transduction elements in an end-organ are normally inaccessible by patch-clamp recording electrodes. Here we describe an in situ patch-clamp recording protocol for the study of tactile transduction in Merkel cells of rat whisker hair follicles, one of the most sensitive tactile end-organs in mammals. This technique offers an opportunity to explore the identities and properties of ion channels that are involved in tactile transduction in whisker hair follicles, and it may also lend a useful tool for researchers to study other tactile end-organs. The experimental protocol describes procedures for 1) tissue dissection and whisker hair follicle preparation, 2) device setup and steps for performing patch-clamp recordings from Merkel cells in a whisker hair follicle, 3) methods of delivering mechanical stimuli, and 4) intra-follicle microinjection for receptor knockdown in whisker hair follicles. The main procedures in this protocol, from tissue preparation to whole-cell patch-clamp recordings, can be completed in a few hours.
哺乳动物利用触觉末梢器官来执行诸如环境探索、社交互动和触觉辨别等感觉任务。然而，触觉末梢器官中触觉转导的细胞和分子机制仍知之甚少。膜片钳记录技术可能是检测和研究触觉末梢器官中触觉转导最有价值的方法，但它在技术上具有挑战性，因为末梢器官中的触觉转导元件通常无法通过膜片钳记录电极接触到。在这里，我们描述了一种用于研究大鼠触须毛囊默克尔细胞中触觉转导的原位膜片钳记录方案，触须毛囊是哺乳动物中最敏感的触觉末梢器官之一。这项技术为探索参与触须毛囊触觉转导的离子通道的特性提供了机会，它也可能为研究人员研究其他触觉末梢器官提供一个有用的工具。该实验方案描述了以下步骤：1）组织解剖和触须毛囊制备；2）设备设置以及从触须毛囊中的默克尔细胞进行膜片钳记录的步骤；3）施加机械刺激的方法；4）用于在触须毛囊中敲低受体的毛囊内显微注射。该方案中的主要步骤，从组织制备到全细胞膜片钳记录，几个小时内即可完成。
