Upregulated Connexin 43 Induced by Loss-of-Functional S284L-Mutant α4 Subunit of Nicotinic ACh Receptor Contributes to Pathomechanisms of Autosomal Dominant Sleep-Related Hypermotor Epilepsy

To study the pathomechanism and pathophysiology of autosomal dominant sleep-related hypermotor epilepsy (ADSHE), this study determined functional abnormalities of glutamatergic transmission in the thalamocortical motor pathway, from the reticular thalamic nucleus (RTN), motor thalamic nuclei (MoTN) tosecondary motor cortex (M2C) associated with the S286L-mutant alpha 4 beta 2-nicotinic acetylcholine receptor (nAChR) and the connexin43 (Cx43) hemichannel of transgenic rats bearing the rat S286L-mutant Chrna4 gene (S286L-TG), which corresponds to the human S284L-mutant CHRNA4 gene using multiprobe microdialysis, primary cultured astrocytes and a Simple Western system. Expression of Cx43 in the M2C plasma membrane fraction of S286L-TG was upregulated compared with wild-type rats. Subchronic nicotine administration decreased Cx43 expression of wild-type, but did not affect that of S286L-TG; however, zonisamide (ZNS) decreased Cx43 in both wild-type and S286L-TG. Primary cultured astrocytes of wild-type were not affected by subchronic administration of nicotine but was decreased by ZNS. Upregulated Cx43 enhanced glutamatergic transmission during both resting and hyperexcitable stages in S286L-TG. Furthermore, activation of glutamatergic transmission associated with upregulated Cx43 reinforced the prolonged Cx43 hemichannel activation. Subchronic administration of therapeutic-relevant doses of ZNS compensated the upregulation of Cx43 and prolonged reinforced activation of Cx43 hemichannel induced by physiological hyperexcitability during the non-rapid eye movement phase of sleep. The present results support the primary pathomechanisms and secondary pathophysiology of ADSHE seizures of patients with S284L-mutation.

为研究常染色体显性遗传性睡眠相关运动过度性癫痫（ADSHE）的发病机制和病理生理学，本研究采用多探头微透析技术、原代培养星形胶质细胞及Simple Western系统，测定了携带大鼠S286L突变Chrna4基因（S286L - TG，对应人类S284L突变CHRNA4基因）的转基因大鼠，其丘脑皮质运动通路（从丘脑网状核（RTN）、丘脑运动核（MoTN）到次级运动皮层（M2C））中与S286L突变型α4β2烟碱型乙酰胆碱受体（nAChR）及连接蛋白43（Cx43）半通道相关的谷氨酸能传递功能异常情况。与野生型大鼠相比，S286L - TG大鼠M2C质膜部分中Cx43的表达上调。亚慢性给予尼古丁可降低野生型大鼠Cx43的表达，但对S286L - TG大鼠无影响；然而，唑尼沙胺（ZNS）可降低野生型和S286L - TG大鼠的Cx43表达。亚慢性给予尼古丁对野生型大鼠原代培养的星形胶质细胞无影响，但ZNS可使其Cx43表达降低。在S286L - TG大鼠静息和过度兴奋阶段，上调的Cx43均增强了谷氨酸能传递。此外，与Cx43上调相关的谷氨酸能传递激活，强化了Cx43半通道的长时间激活。亚慢性给予治疗相关剂量的ZNS，可补偿在睡眠非快速眼动期生理过度兴奋所诱导的Cx43上调及Cx43半通道长时间强化激活。本研究结果支持S284L突变患者ADSHE癫痫发作的原发性发病机制和继发性病理生理过程。