Zawieja Scott D, Pea Grace A, Broyhill Sarah E, Patro Advaya, Bromert Karen H, Norton Charles E, Kim Hae Jin, Sivasankaran Sathesh Kumar, Li Min, Castorena-Gonzalez Jorge A, Drumm Bernard T, Davis Michael J
Department of Medical Pharmacology & Physiology, University of Missouri, Columbia, United States.
Bioinformatics and Analytics Core, Division of Research, Innovation and Impact, University of Missouri, Columbia, United States.
Elife. 2025 Sep 11;12:RP90679. doi: 10.7554/eLife.90679.
Collecting lymphatic vessels (cLVs) exhibit spontaneous contractions with a pressure-dependent frequency, but the identity of the lymphatic pacemaker cell is still debated. Here, we combined immunofluorescence and scRNAseq analyses with electrophysiological methods to examine the cellular constituents of the mouse cLV wall and assess whether any cell type exhibited morphological and functional processes characteristic of pacemaker cells. We employed inducible Cre mouse models to target-specific cell populations including CkitCreER to target interstitial cells of Cajal-like cells, PdgfrβCreER to target pericyte-like cells; PdgfrαCreER to target CD34 adventitial cells; and Myh11CreER to target lymphatic muscle cells (LMCs) directly. These inducible Cre lines were crossed to the fluorescent reporter , the genetically encoded Ca sensor GCaMP6f, and the light-activated cation channel rhodopsin2 (ChR2). Only LMCs consistently, but heterogeneously, displayed spontaneous Ca events during the diastolic period of the contraction cycle, and whose frequency was modulated in a pressure-dependent manner. Further, optogenetic depolarization with ChR2 induced propagated contractions only in LMCs. Membrane potential recordings in LMCs demonstrated that the rate of diastolic depolarization significantly correlated with contraction frequency. These findings support the conclusion that LMCs, or a subset of LMCs, are responsible for mouse cLV pacemaking.
集合淋巴管（cLVs）表现出与压力相关频率的自发收缩，但淋巴管起搏细胞的身份仍存在争议。在这里，我们将免疫荧光和单细胞RNA测序分析与电生理方法相结合，以检查小鼠cLV壁的细胞成分，并评估是否有任何细胞类型表现出起搏细胞特有的形态和功能过程。我们使用诱导型Cre小鼠模型来靶向特定细胞群体，包括使用CkitCreER靶向类 Cajal 间质细胞，使用PdgfrβCreER靶向类周细胞；使用PdgfrαCreER靶向CD34外膜细胞；以及使用Myh11CreER直接靶向淋巴管平滑肌细胞（LMCs）。这些诱导型Cre品系与荧光报告基因、基因编码的钙传感器GCaMP6f和光激活阳离子通道视紫红质2（ChR2）杂交。只有LMCs在收缩周期的舒张期持续但不均匀地显示出自发性钙事件，并且其频率以压力依赖的方式受到调节。此外，用ChR2进行光遗传学去极化仅在LMCs中诱导传播性收缩。LMCs的膜电位记录表明，舒张期去极化速率与收缩频率显著相关。这些发现支持了LMCs或LMCs的一个子集负责小鼠cLV起搏的结论。
