AIM:This study aims to investigate the impact of TWIK-1 channels on abnormal pacemaker activi-ties induced by hypokalemia and to elucidate the underlying mechanisms.METHODS:The gene sequences encoding hu-man TWIK-1,specific TWIK-1 shRNA and TWIK-1-T118i mutant were synthesized and subsequently subcloned into lenti-viral vectors.To knock down the TWIK-1 gene in human induced pluripotent stem cell-derived cardiomyocytes(hiPSC-CMs),the cells were transduced with lentivirus carrying the specific TWIK-1 shRNA sequences.For the overexpression of TWIK-1 or the TWIK-1-T118i mutant in HL-1 mouse cardiomyocytes,the cells received lentiviral transduction containing the respective gene sequences.Patch-clamp techniques were employed to assess the effects of 1 mmol/L extracellular K+on the membrane potentials and whole-cell currents of the cardiomyocytes.RESULTS:Under conditions of 1 mmol/L extra-cellular K+,depolarization of membrane potentials was observed in the hiPSC-CMs and the HL-1 mouse cardiomyocytes ex-pressing human TWIK-1 channel,leading to the induction of abnormal pacemaker activities.This phenomenon could be reversibly abolished by the removal of extracellular Na+or inhibited through TWIK-1 knockdown.In contrast,the mem-brane potentials of HL-1 mouse cardiomyocytes expressing human TWIK-1-T118i mutant hyperpolarized,with no occur-rence of abnormal pacemaker activities.The hiPSC-CMs exhibiting abnormal pacemaker activities at 1 mmol/L extracellu-lar K+demonstrated TWIK-1-like Na+leak currents,which were blocked by quinine,a non-selective blocker of TWIK-1.CONCLUSION:The TWIK-1 channels play a critical role in the development of hypokalemia-induced abnormal pace-maker activities in human cardiomyocytes by facilitating Na+leak currents.

AIM:This study aims to investigate the impact of TWIK-1 channels on abnormal pacemaker activi-ties induced by hypokalemia and to elucidate the underlying mechanisms.METHODS:The gene sequences encoding hu-man TWIK-1,specific TWIK-1 shRNA and TWIK-1-T118i mutant were synthesized and subsequently subcloned into lenti-viral vectors.To knock down the TWIK-1 gene in human induced pluripotent stem cell-derived cardiomyocytes(hiPSC-CMs),the cells were transduced with lentivirus carrying the specific TWIK-1 shRNA sequences.For the overexpression of TWIK-1 or the TWIK-1-T118i mutant in HL-1 mouse cardiomyocytes,the cells received lentiviral transduction containing the respective gene sequences.Patch-clamp techniques were employed to assess the effects of 1 mmol/L extracellular K+on the membrane potentials and whole-cell currents of the cardiomyocytes.RESULTS:Under conditions of 1 mmol/L extra-cellular K+,depolarization of membrane potentials was observed in the hiPSC-CMs and the HL-1 mouse cardiomyocytes ex-pressing human TWIK-1 channel,leading to the induction of abnormal pacemaker activities.This phenomenon could be reversibly abolished by the removal of extracellular Na+or inhibited through TWIK-1 knockdown.In contrast,the mem-brane potentials of HL-1 mouse cardiomyocytes expressing human TWIK-1-T118i mutant hyperpolarized,with no occur-rence of abnormal pacemaker activities.The hiPSC-CMs exhibiting abnormal pacemaker activities at 1 mmol/L extracellu-lar K+demonstrated TWIK-1-like Na+leak currents,which were blocked by quinine,a non-selective blocker of TWIK-1.CONCLUSION:The TWIK-1 channels play a critical role in the development of hypokalemia-induced abnormal pace-maker activities in human cardiomyocytes by facilitating Na+leak currents.

Objective:To observe the neuroprotective effect of wind medicine (Puerariae Lobatue Radix and Acori Tatarinowii Rhizoma) in combination with Panax ginseng total saponin (PNS) on cerebral ischemia-reperfusion rats; To elucidate the mechanism of "wind medicine increasing effect".Methods:Totally 140 male SD rats were divided into sham-operation group, model group, PNS group, Puerariae Lobatue Radix group, Acori Tatarinowii Rhizoma group, Puerariae Lobatue Radix + PNS group, Acori Tatarinowii Rhizoma + PNS group according to the random number table method, with 20 rats in each group. Except for the sham-operation group, the cerebral ischemia/reperfusion rat model was established using the modified Longa line bolus method in the remaining groups. After 7 d of administration of the appropriate pharmacologic intervention in each group, neurological dysfunction was evaluated by Zea-longa score after final administration, cerebral infarct volume was determined by TTC staining; blood brain barrier (BBB) permeability of brain tissue on the ischemic side was detected by Evans blue content; BBB ultrastructure of each group of rats was observed by transmission electron microscopy; Claudin 5 protein expression level was detected by immunohistochemistry; Zonula occludens-1 (ZO-1), major facilitator supeffamily domain-containing protein 2a (Mfsd2a), Occludin, P-glycoprotein (P-gp), Monocarboxylate Transporters-1 (MCT1) and breast cancer resistance protein (BCRP) protein expression levels were detected by Western-blot.Results:Compared with the model group, the rat neurological function scores were reduced in each administration group ( P<0.05), infarct volume was reduced ( P<0.05), EB content of brain tissue decreased ( P<0.05), protein expressions of Claudin 5, ZO-1, Mfsd2a and Occludin in brain tissue were elevated ( P<0.05), the protein expressions of P-gp, BCRP and MCT1 were reduced ( P<0.05), and the protein expressions of Claudin 5, Mfsd2a, and Occludin was higher in the Puerariae Lobatue Radix + PNS group and Acori Tatarinowii Rhizoma + PNS group than that of each group of medication alone ( P<0.05), and the protein expression of MCT1 was lower than that of each group of medication alone ( P<0.05); the protein expression level of ZO-1 in the Puerariae Lobatue Radix + PNS group was higher than that of the group of medication alone ( P<0.05); P-gp protein expression was lower in Acori Tatarinowii Rhizoma + PNS group than in the PNS group and Acori Tatarinowii Rhizoma group ( P<0.05). Conclusion:Wind medicine (Puerariae Lobatue Radix and Acori Tatarinowii Rhizoma) may potentiate the neuroprotective effect of PNS on cerebral ischemia-reperfusion rats, and the mechanism may be related to the protection of BBB structural integrity and maintenance of central barrier properties, while regulating substance transport proteins and increasing the intracerebral content of the drug.