Walsh Yvonne, Leach Michael, Veale Emma L, Mathie Alistair
Medway School of Pharmacy, University of Kent and University of Greenwich, Central Avenue, Chatham Maritime, ME4 4TB, UK.
University of Greenwich, Central Avenue, Chatham Maritime, ME4 4TB, UK.
Biochem Biophys Rep. 2021 May 19;26:101021. doi: 10.1016/j.bbrep.2021.101021. eCollection 2021 Jul.
TREK and TRESK K2P channels are widely expressed in the nervous system, particularly in sensory neurons, where they regulate neuronal excitability. In this study, using whole-cell patch-clamp electrophysiology, we characterise the inhibitory effect of the anticonvulsant lamotrigine and two derivatives, sipatrigine and 3,5-diamino-6-(3,5-bistrifluoromethylphenyl)-1,2,4-triazine (CEN-092) on these channels. Sipatrigine was found to be a more effective inhibitor than lamotrigine of TREK-1, TREK-2 and TRESK channels. Sipatrigine was slightly more potent on TREK-1 channels (EC = 16 μM) than TRESK (EC = 34 μM) whereas lamotrigine was equally effective on TREK-1 and TRESK. Sipatrigine was less effective on a short isoform of TREK-2, suggesting the N terminus of the channel is important for both inhibition and subsequent over-recovery. Inhibition of TREK-1 and TREK-2 channels by sipatrigine was reduced by mutation of a leucine residue associated with the norfluoxetine binding site on these channels (L289A and L320A on TREK-1 and TREK-2, respectively) but these did not affect inhibition by lamotrigine. Inhibition of TRESK by sipatrigine and lamotrigine was attenuated by mutation of bulky phenylalanine residues (F145A and F352A) in the inner pore helix. However, phosphorylation mutations did not alter the effect of sipatrigine. CEN-092 was a more effective inhibitor of TRESK channels than TREK-1 channels. It is concluded that lamotrigine, sipatrigine and CEN-092 are all inhibitors of TREK and TRESK channels but do not greatly discriminate between them. The actions of these compounds may contribute to their current and potential use in the treatment of pain and depression.
TREK和TRESK K2P通道在神经系统中广泛表达，尤其是在感觉神经元中，它们在其中调节神经元兴奋性。在本研究中，我们使用全细胞膜片钳电生理学方法，表征了抗惊厥药物拉莫三嗪及其两种衍生物西帕三嗪和3,5-二氨基-6-(3,5-双三氟甲基苯基)-1,2,4-三嗪（CEN-092）对这些通道的抑制作用。结果发现，西帕三嗪对TREK-1、TREK-2和TRESK通道的抑制作用比拉莫三嗪更有效。西帕三嗪对TREK-1通道（EC = 16 μM）的效力略高于TRESK（EC = 34 μM），而拉莫三嗪对TREK-1和TRESK的作用相同。西帕三嗪对TREK-2的短异构体效果较差，这表明通道的N端对抑制作用和随后的过度恢复都很重要。与这些通道上的去甲氟西汀结合位点相关的亮氨酸残基发生突变（TREK-1和TREK-2上分别为L289A和L320A）后，西帕三嗪对TREK-1和TREK-2通道的抑制作用减弱，但这些突变不影响拉莫三嗪的抑制作用。西帕三嗪和拉莫三嗪对TRESK的抑制作用通过内孔螺旋中庞大的苯丙氨酸残基（F145A和F352A）突变而减弱。然而，磷酸化突变并未改变西帕三嗪的作用。CEN-092对TRESK通道的抑制作用比对TREK-1通道更有效。得出的结论是，拉莫三嗪、西帕三嗪和CEN-092都是TREK和TRESK通道的抑制剂，但对它们的区分不大。这些化合物的作用可能有助于它们目前以及潜在地用于治疗疼痛和抑郁症。
