Ghosh Srijit, Alkawadri Tuleen, McGarvey Lorcan P, Hollywood Mark A, Thornbury Keith D, Sergeant Gerard P
Smooth Muscle Research Centre, Dundalk Institute of Technology, Co. Louth, Ireland.
School of Medicine, Dentistry, and Biomedical Sciences, Queen's University, Belfast, Northern Ireland.
Am J Physiol Lung Cell Mol Physiol. 2025 Feb 1;328(2):L301-L312. doi: 10.1152/ajplung.00188.2024. Epub 2025 Jan 8.
Cholinergic tone is elevated in obstructive lung conditions such as chronic obstructive pulmonary disease (COPD) and asthma, but the cellular mechanisms underlying cholinergic contractions of airway smooth muscle (ASM) are still unclear. Some studies report an important role for L-type Ca channels (LTCC) and Ano1 Ca-activated Cl channels (CACC) in these responses, but others dispute their importance. Cholinergic contractions of ASM involve activation of M3Rs, however, stimulation of M2Rs exerts a profound hypersensitization of these responses. Here, we show that M2R-dependent potentiation of cholinergic nerve-evoked contractions of ASM was reversed by the LTCC blocker nifedipine and the Ano1 CACC inhibitors Ani9 and CaCC-A01. Carbachol induced sustained contractions of ASM that were converted into oscillatory contractions when M3Rs were blocked with 4-DAMP. The 4-DAMP-resistant contractions were absent in preparations taken from M2R knockout (KO) mice. The remaining M2R-dependent responses, observed in wild-type (WT) mice, were abolished by nifedipine and Ani9. Inhibition of sarcoplasmic endoplasmic reticulum Ca ATPases (SERCA) with thapsigargin increased the amplitude of contractions induced by electrical field stimulation (EFS) and these effects were also reversed by nifedipine and Ani9. Thapsigargin also potentiated contractions of ASM induced by the LTCC activator FPL64176. Therefore, contractions of ASM that involved Ca influx via LTCC were enhanced by inhibition of SERCA. Immunocytochemistry experiments revealed prominent SERCA staining around the periphery of ASM cells. These data indicate that M2R-dependent contractions of ASM involve Ano1 CACC and LTCC by a mechanism involving inhibition of buffering of Ca influx by SERCA. The role of L-type Ca channels and Ano1 Ca-activated Cl channels in cholinergic contractions of airway smooth muscle is disputed. Here, we show that both channels are involved in M2 muscarinic receptor-dependent contractions of murine airway smooth muscle via inhibition of buffering of Ca influx by sarcoplasmic endoplasmic reticulum Ca ATPases.
在诸如慢性阻塞性肺疾病（COPD）和哮喘等阻塞性肺部疾病中，胆碱能张力升高，但气道平滑肌（ASM）胆碱能收缩的细胞机制仍不清楚。一些研究报道L型钙通道（LTCC）和Ano1钙激活氯通道（CACC）在这些反应中起重要作用，但其他研究对它们的重要性提出质疑。ASM的胆碱能收缩涉及M3R的激活，然而，M2R的刺激会使这些反应产生深刻的超敏反应。在这里，我们表明，LTCC阻滞剂硝苯地平和Ano1 CACC抑制剂Ani9和CaCC-A01可逆转M2R依赖性增强的ASM胆碱能神经诱发收缩。卡巴胆碱诱导ASM持续收缩，当用4-DAMP阻断M3R时，收缩转变为振荡收缩。从M2R基因敲除（KO）小鼠制备的标本中不存在对4-DAMP耐药的收缩。在野生型（WT）小鼠中观察到的其余M2R依赖性反应被硝苯地平和Ani9消除。用毒胡萝卜素抑制肌浆内质网钙ATP酶（SERCA）可增加电场刺激（EFS）诱导的收缩幅度，这些作用也被硝苯地平和Ani9逆转。毒胡萝卜素还增强了LTCC激活剂FPL64176诱导的ASM收缩。因此，通过抑制SERCA对钙内流的缓冲作用，涉及通过LTCC的钙内流的ASM收缩增强。免疫细胞化学实验显示ASM细胞周边有明显的SERCA染色。这些数据表明，ASM的M2R依赖性收缩通过一种涉及抑制SERCA对钙内流缓冲的机制涉及Ano1 CACC和LTCC。L型钙通道和Ano1钙激活氯通道在气道平滑肌胆碱能收缩中的作用存在争议。在这里，我们表明这两种通道都通过抑制肌浆内质网钙ATP酶对钙内流的缓冲作用参与小鼠气道平滑肌的M2毒蕈碱受体依赖性收缩。
