Vangeel Laura, Benoit Melissa, Miron Yannick, Miller Paul E, De Clercq Katrien, Chaltin Patrick, Verfaillie Catherine, Vriens Joris, Voets Thomas
Laboratory of Ion Channel Research, VIB-KU Leuven Center for Brain & Disease Research, Leuven, Belgium.
Department of Cellular and Molecular Medicine, KU Leuven, Leuven, Belgium.
Br J Pharmacol. 2020 Jun;177(12):2683-2695. doi: 10.1111/bph.14994. Epub 2020 Mar 5.
The transient receptor potential (TRP) ion channel TRPM3 functions as a noxious heat sensor, plays a key role in acute pain sensation and inflammatory hyperalgesia in rodents. Despite its potential as a novel analgesic drug target, little is known about the expression, function and modulation in the humans.
We studied TRPM3 in freshly isolated human dorsal root ganglion (hDRG) neurons and human stem cell-derived sensory (hSCDS) neurons. Expression was analysed at the mRNA level using RT-qPCR. Channel function was assessed using Fura-2-based calcium imaging and whole-cell patch-clamp recordings.
TRPM3 was detected at the mRNA level in both hDRG and hSCDS neurons. The TRPM3 agonists pregnenolone sulphate (PS) and CIM0216 evoked robust intracellular Ca responses in 52% of hDRG and 58% of hSCDS neurons. Whole-cell patch-clamp recordings in hSCDS neurons revealed pregnenolone sulphate (PS)- and CIM0216-evoked currents exhibiting the characteristic current-voltage relation of TRPM3. PS-induced calcium responses in hSCDS neurons were reversed in a dose-dependent manner by the flavonoid isosakuranetin and by antiseizure drug primidone. Finally, the μ-opioid receptor agonist DAMGO and the GABA receptor agonist baclofen inhibited PS-evoked TRPM3 responses in a subset of hSCDS neurons.
These results provide the first direct evidence of functional expression of the pain receptor TRPM3 in human sensory neurons, largely mirroring the channel's properties observed in mouse sensory neurons. hSCDS neurons represent a valuable and readily accessible in vitro model to study TRPM3 regulation and pharmacology in a relevant human cellular context.
瞬时受体电位（TRP）离子通道TRPM3作为一种有害热感受器，在啮齿动物的急性疼痛感觉和炎症性痛觉过敏中起关键作用。尽管它有潜力成为一种新型镇痛药物靶点，但对其在人类中的表达、功能和调节知之甚少。
我们在新鲜分离的人背根神经节（hDRG）神经元和人干细胞衍生的感觉（hSCDS）神经元中研究TRPM3。使用RT-qPCR在mRNA水平分析表达情况。使用基于Fura-2的钙成像和全细胞膜片钳记录评估通道功能。
在hDRG和hSCDS神经元的mRNA水平均检测到TRPM3。TRPM3激动剂硫酸孕烯醇酮（PS）和CIM0216在52%的hDRG神经元和58%的hSCDS神经元中诱发强烈的细胞内钙反应。hSCDS神经元的全细胞膜片钳记录显示，硫酸孕烯醇酮（PS）和CIM0216诱发的电流呈现出TRPM3特征性的电流-电压关系。黄酮类化合物异樱花素和抗癫痫药物扑米酮以剂量依赖的方式逆转了PS在hSCDS神经元中诱导的钙反应。最后，μ-阿片受体激动剂DAMGO和GABA受体激动剂巴氯芬在一部分hSCDS神经元中抑制了PS诱发的TRPM3反应。
这些结果首次直接证明了疼痛受体TRPM3在人感觉神经元中的功能性表达，很大程度上反映了在小鼠感觉神经元中观察到的该通道特性。hSCDS神经元是一种有价值且易于获得的体外模型，可在相关的人类细胞环境中研究TRPM3的调节和药理学。
