Caveolae facilitate muscarinic receptor-mediated intracellular Ca 2+ mobilization and contraction in airway smooth muscle

• Published in: American journal of physiology. Lung cellular and molecular physiology 2007-12, Vol.293 (6), p.L1406-L1418
• Main Authors: Gosens, Reinoud, Stelmack, Gerald L., Dueck, Gordon, Mutawe, Mark M., Hinton, Martha, McNeill, Karol D., Paulson, Angela, Dakshinamurti, Shyamala, Gerthoffer, William T., Thliveris, James A., Unruh, Helmut, Zaagsma, Johan, Halayko, Andrew J.
• Format: Article
• Language: English
• ISSN: 1040-0605; 1522-1504
• DOI: 10.1152/ajplung.00312.2007

Contractile responses of airway smooth muscle (ASM) determine airway resistance in health and disease. Caveolae microdomains in the plasma membrane are marked by caveolin proteins and are abundant in contractile smooth muscle in association with nanospaces involved in Ca 2+ homeostasis. Caveolin-1 can modulate localization and activity of signaling proteins, including trimeric G proteins, via a scaffolding domain. We investigated the role of caveolae in contraction and intracellular Ca 2+ ([Ca 2+ ] i ) mobilization of ASM induced by the physiological muscarinic receptor agonist, acetylcholine (ACh). Human and canine ASM tissues and cells predominantly express caveolin-1. Muscarinic M 3 receptors (M 3 R) and Gα q/11 cofractionate with caveolin-1-rich membranes of ASM tissue. Caveolae disruption with β-cyclodextrin in canine tracheal strips reduced sensitivity but not maximum isometric force induced by ACh. In fura-2-loaded canine and human ASM cells, exposure to methyl-β-cyclodextrin (mβCD) reduced sensitivity but not maximum [Ca 2+ ] i induced by ACh. In contrast, both parameters were reduced for the partial muscarinic agonist, pilocarpine. Fluorescence microscopy revealed that mβCD disrupted the colocalization of caveolae-1 and M 3 R, but [ N-methyl- 3 H]scopolamine receptor-binding assay revealed no effect on muscarinic receptor availability or affinity. To dissect the role of caveolin-1 in ACh-induced [Ca 2+ ] i flux, we disrupted its binding to signaling proteins using either a cell-permeable caveolin-1 scaffolding domain peptide mimetic or by small interfering RNA knockdown. Similar to the effects of mβCD, direct targeting of caveolin-1 reduced sensitivity to ACh, but maximum [Ca 2+ ] i mobilization was unaffected. These results indicate caveolae and caveolin-1 facilitate [Ca 2+ ] i mobilization leading to ASM contraction induced by submaximal concentrations of ACh.