Division of Cell Signaling, Okazaki Institute for Integrative Bioscience (National Institute for Physiological Sciences), National Institutes of Natural Sciences, 5-1 Higashiyama, Myodaiji, Okazaki, Aichi, 444-8787, Japan.
Department of Physiological Sciences, The Graduate University for Advanced Studies (SOKENDAI), 5-1 Higashiyama, Myodaiji, Okazaki, Aichi, 444-8787, Japan.
Pflugers Arch. 2017 Oct;469(10):1313-1323. doi: 10.1007/s00424-017-1998-7. Epub 2017 Jun 13.
The sensation of itching can be defined as "an unpleasant cutaneous sensation that provokes a desire to scratch." The perception of itching is not critical for the maintenance of life, but persistent itching can be extremely irritating and decreases the quality of life. Crotamiton (N-ethyl-o-crotonotoluidide) has been used as an anti-itch agent for humans for around 70 years. In spite of the long use of crotamiton, its mechanism of action remains unknown. We hypothesized that crotamiton might have effects on transient receptor potential (TRP) channels expressed in the peripheral nervous system and the skin. We first examined the effects of crotamiton on TRP channels by whole-cell patch-clamp recordings. We found that crotamiton strongly inhibited TRPV (vanilloid) 4 channels followed by large currents after crotamiton washout. In mice, crotamiton inhibited itch-related behaviors induced by a TRPV4-selective agonist (GSK1016790A). We biophysically investigated the large TRPV4 currents after crotamiton washout. Comparing single-channel open probabilities and current amplitudes of TRPV4, increases in both parameters were found to contribute to the large washout currents of TRPV4. Because the change in current amplitudes suggested pore dilation of TRPV4, we examined this possibility with cation replacement experiments and by measuring changes in reversal potentials. Greater cation influxes and changes in reversal potentials upon crotamiton washout were observed, suggesting that the TRPV4 pore dilated in its uninhibited state. From these results, we identified the molecular target of crotamiton as TRPV4 and demonstrated pore dilation of TRPV4 upon crotamiton washout.
瘙痒的感觉可以被定义为“一种不愉快的皮肤感觉，会引起搔抓的欲望”。瘙痒的感觉对于维持生命并不重要，但持续的瘙痒会非常令人不适，降低生活质量。克罗米通（N-乙基-O-克罗米通）已经被人类用作抗瘙痒剂约 70 年。尽管克罗米通已经被长期使用，但它的作用机制仍然未知。我们假设克罗米通可能对周围神经系统和皮肤中表达的瞬时受体电位（TRP）通道有影响。我们首先通过全细胞膜片钳记录来检查克罗米通对 TRP 通道的影响。我们发现克罗米通强烈抑制 TRPV（香草素）4 通道，随后在克罗米通洗脱后出现大电流。在小鼠中，克罗米通抑制由 TRPV4 选择性激动剂（GSK1016790A）诱导的瘙痒相关行为。我们从生物物理角度研究了克罗米通洗脱后的大 TRPV4 电流。比较 TRPV4 的单通道开放概率和电流幅度，发现这两个参数的增加都有助于 TRPV4 的大洗脱电流。由于电流幅度的变化提示 TRPV4 的孔扩张，我们通过阳离子替换实验和测量反转电位的变化来检查这种可能性。在克罗米通洗脱后观察到更大的阳离子内流和反转电位的变化，这表明 TRPV4 孔在未被抑制的状态下扩张。根据这些结果，我们确定了克罗米通的分子靶标为 TRPV4，并证明了 TRPV4 在克罗米通洗脱后孔扩张。
