Yin Zeyuan, Torre Eleonora, Marrot Manon, Peters Colin H, Feather Amy, Nichols William G, Logantha Sunil Jit R J, Arshad Areej, Martis Simran Agnes, Ozturk Nilay Tugba, Chen Weixuan, Liu Jiaxuan, Qu Jingmo, Zi Min, Cartwright Elizabeth J, Proenza Catherine, Torrente Angelo, Mangoni Matteo E, Dobrzynski Halina, Atkinson Andrew J
Division of Cardiovascular Sciences, School of Medical Sciences, University of Manchester, Manchester, United Kingdom.
Institut de Génomique Fonctionnelle, Université de Montpellier CNRS, INSERM, Montpellier, France.
Front Med (Lausanne). 2024 Dec 5;11:1488478. doi: 10.3389/fmed.2024.1488478. eCollection 2024.
The sinoatrial node (SN) generates the heart rate (HR). Its spontaneous activity is regulated by a complex interplay between the modulation by the autonomic nervous system (ANS) and intrinsic factors including ion channels in SN cells. However, the systemic and intrinsic regulatory mechanisms are still poorly understood. This study aimed to elucidate the sex-specific differences in heart morphology and SN function, particularly focusing on basal HR, expression and function of hyperpolarization-activated HCN4 and HCN1 channels and mRNA abundance of ion channels and mRNA abundance of ion channels contributing to diastolic depolarization (DD) and spontaneous action potentials (APs).
Body weight, heart weight and tibia length of 2- to 3-month-old male and female mice were measured. Conscious HR of male and female mice was recorded via electrocardiography (ECG). Unconscious HR, stroke volume (SV) and ejection fraction (EF) were recorded via echocardiography. HR was measured via Langendorff apparatus. Volume of atria, ventricles and whole hearts were measured from the hearts by microcomputed tomography (micro-CT). Immunohistochemistry targeting HCN4 and HCN1 was conducted in the SN and RA tissues from both male and female hearts. The funny current ( ) of SN cells in 1 nM and following wash-on of 1 μM isoproterenol (ISO) were recorded via whole cell patch clamp. The APs of SN tissue were recorded via sharp microelectrode and optical mapping of membrane voltage. The relative abundance of mRNAs was measured in male and female mice by qPCR.
Heart weight to tibia length ratio and heart volume of females were significantly smaller than males. Unconscious HR in male mice was higher than that in females. Conscious HR, HR, SV, and EF showed no notable difference between male and female mice. Immunohistochemistry revealed HCN4, HCN1, and the sum of HCN4 and HCN1, expression in the SN was notably elevated compared with the RA in both male and females, but there was no sex difference in these channels expression. There were also no significant sex differences in the of in SN cells in the presence of 1 nM ISO, however wash-on 1 μM ISO in the same cells induced a significantly increased shift of to more positive voltages in males than in females. The expression of mRNA coding for adrenergic receptor beta-1 (Adrb1) and cholinergic receptors muscarinic 2 (chrm2) in male mice was higher compared with that in female mice. Early diastolic depolarization (EDD) rate in APs from peripheral SN (pSN) from male mice were higher than these in female mice. Mice of both sexes showed equivalent frequency of SN APs and spatial localization of the leading site in control, and similar significant response to ISO 100 nM superfusion.
Males display faster HR, but not HR, than females associated with increased expression of Adrb1 in male versus female. This suggests a possible difference in the β-adrenergic modulation in males and females, possibly related to the greater ISO response of observed in cells from males. The role of hormonal influences or differential expression of other ion channels may explain these sex-specific variations in HR dynamics. Further investigations are necessary to pinpoint the precise molecular substrates responsible for these differences.
窦房结（SN）产生心率（HR）。其自发活动受自主神经系统（ANS）调节与包括窦房结细胞离子通道在内的内在因素之间复杂相互作用的调控。然而，全身和内在调节机制仍知之甚少。本研究旨在阐明心脏形态和窦房结功能的性别特异性差异，尤其关注基础心率、超极化激活的HCN4和HCN1通道的表达与功能以及参与舒张期去极化（DD）和自发动作电位（APs）的离子通道的mRNA丰度。
测量2至3月龄雄性和雌性小鼠的体重、心脏重量和胫骨长度。通过心电图（ECG）记录雄性和雌性小鼠的清醒心率。通过超声心动图记录无意识心率、心搏量（SV）和射血分数（EF）。通过Langendorff装置测量心率。通过微型计算机断层扫描（micro-CT）测量雄性和雌性心脏的心房、心室和全心的体积。对雄性和雌性心脏的窦房结和右心房（RA）组织进行针对HCN4和HCN1的免疫组织化学检测。通过全细胞膜片钳记录1 nM时窦房结细胞的起搏电流（If）以及在加入1 μM异丙肾上腺素（ISO）后的情况。通过尖锐微电极和膜电压光学映射记录窦房结组织的动作电位。通过qPCR测量雄性和雌性小鼠中mRNA的相对丰度。
雌性小鼠的心脏重量与胫骨长度之比和心脏体积显著小于雄性。雄性小鼠的无意识心率高于雌性。雄性和雌性小鼠的清醒心率、心率、心搏量和射血分数无显著差异。免疫组织化学显示，雄性和雌性的窦房结中HCN4、HCN1以及HCN4与HCN1之和的表达均显著高于右心房，但这些通道的表达无性别差异。在存在1 nM ISO的情况下，窦房结细胞的If也无显著性别差异，然而在相同细胞中加入1 μM ISO后，雄性细胞的If向更正电压的显著偏移幅度大于雌性。雄性小鼠中肾上腺素能受体β-1（Adrb1）和胆碱能毒蕈碱2型受体（chrm2）编码mRNA的表达高于雌性小鼠。雄性小鼠外周窦房结（pSN）动作电位的早期舒张期去极化（EDD）速率高于雌性小鼠。在对照中，两性小鼠的窦房结动作电位频率和主导位点的空间定位相同，对100 nM ISO灌注的显著反应也相似。
与雌性相比，雄性表现出更快的心率储备（HRR）而非静息心率（HR），这与雄性中Adrb1表达增加有关。这表明雄性和雌性在β-肾上腺素能调节方面可能存在差异，可能与雄性细胞中观察到的对ISO更大反应有关。激素影响或其他离子通道差异表达的作用可能解释了心率动态中的这些性别特异性变化。需要进一步研究以确定导致这些差异的确切分子底物。
