摘要
超极化活化环核苷酸门控(hyperpolarization-activated cyclic-nucleotide-gated,HCN)通道参与调制心脏跳动的节律和速率。与HCN1和HCN2有所不同,慢通道HCN4可能不存在电压依赖的滞后现象。本研究采用单细胞膜片钳方法,在稳定转染hHCN4的HEK293细胞上进行电生理记录,观察hHCN4通道是否存在滞后现象,以及cAMP对其的调制作用;同时采用实时定量RT-PCR方法检测窦房结和心房组织中HCNs的表达。电压钳实验结果显示hHCN4电流(Ih)激活随着保持电位超极化的变化而向去极化方向移动。三角电位变化钳(triangular ramp)和动作电位钳的结果也显示了hHCN4的滞后现象。cAMP增加Ih电流幅度,且使电流激活向去极化方向移动,从而改变内源性hHCN4滞后行为。RT-PCR结果显示,人窦房结组织主要表达HCN4,占75%,HCN1占21%,HCN2占3%,HCN3占0.7%。以上结果提示,人窦房结组织主要表达HCN4亚型,hHCN4的Ih存在电压依赖性的滞后现象,且受cAMP调制。由此推断,hHCN4通道的滞后现象可能在窦房结起搏活动中起到了关键作用。
The hyperpolarization-activated cyclic nucleotide-gated （HCN） channels modulate and regulate cardiac rhythm and rate.It has been suggested that,unlike the HCN1 and HCN2 channels,the slower HCN4 channel may not exhibit voltage-dependent hysteresis.We studied the electrophysiological properties of human HCN4 （hHCN4） channels and its modulation by cAMP to determine whether hHCN4 exhibits hysteresis,by using single-cell patch-clamp in HEK293 cells stably transfected with hHCN4.Quantitative real-time RT-PCR was also used to determine levels of expression of HCNs in human cardiac tissue.Voltage-clamp analysis revealed that hHCN4 current （Ih） activation shifted in the depolarizing direction with more hyperpolarized holding potentials.Triangular ramp and action potential clamp protocols also revealed hHCN4 hysteresis.cAMP enhanced Ih and shifted activation in the depolarizing direction,thus modifying the intrinsic hHCN4 hysteresis behavior.Quantitative PCR analysis of human sinoatrial node （SAN） tissue showed that HCN4 accounts for 75% of the HCNs in human SAN while HCN1 （21%）,HCN2 （3%）,and HCN3 （0.7%） constitute the remainder.Our data suggest that HCN4 is the predominant HCN subtype in the human SAN and that Ih exhibits voltage-dependent hysteresis behavior that can be modified by cAMP.Therefore,hHCN4 hysteresis potentially plays a crucial role in human SAN pacemaking activity.
出处
《生理学报》
CAS
CSCD
北大核心
2010年第1期1-13,共13页
Acta Physiologica Sinica
基金
funded by Medtronic,Inc
