Excitatory cholinergic responses in mouse primary bronchial smooth muscle require both Ca2+ entry via L-type Ca2+ channels and store operated Ca2+ entry via Orai channels  

Malfunctions in airway smooth muscle Ca2+-signalling leads to airway hyperresponsiveness in asthma and chronic obstructive pulmonary disease. Ca2+-release from intracellular stores is important in mediating agonist-induced contractions, but the role of influx via L-type Ca2+ channels is controversial. We re-examined roles of the sarcoplasmic reticulum Ca2+ store, refilling of this store via store-operated Ca2+ entry (SOCE) and L-type Ca2+ channel pathways on carbachol (CCh, 0.1-10 μM)-induced contractions of mouse bronchial rings and intracellular Ca2+ signals of mouse bronchial myocytes. In tension experiments, the ryanodine receptor (RyR) blocker dantrolene (100 μM) reduced CCh-responses at all concentrations, with greater effects on sustained rather than initial components of contraction. 2-Aminoethoxydiphenyl borate (2-APB, 100 μM), in the presence of dantrolene, abolished CCh-responses, suggesting sarcoplasmic reticulum Ca2+ store is essential for contraction. The SOCE blocker GSK-7975A (10 μM) reduced CCh-contractions, with greater effects at higher (e.g. 3 and 10 μM) CCh concentrations. Nifedipine (1 μM), abolished remaining contractions in GSK-7975A (10 μM). A similar pattern was observed on intracellular Ca2+-responses to 0.3 μM CCh, where GSK-7975A (10 μM) substantially reduced Ca2+ transients induced by CCh and nifedipine (1 μM) abolished remaining responses. When nifedipine (1 μM) was applied alone it had less effect, reducing tension responses at all CCh concentrations by 25% - 50%, with greater effects at lower (e.g. 0.1 and 0.3 μM) CCh concentrations. When nifedipine (1 μM) was examined on the intracellular Ca2+-response to 0.3 μM CCh, it only modestly reduced Ca2+ signals, while GSK-7975A (10 μM) abolished remaining responses. In conclusion, Ca2+-influx from both SOCE and L-type Ca2+ channels contribute to excitatory cholinergic responses in mouse bronchi. The contribution of L-type Ca2+ channels was especially pronounced at lower doses of CCh, or when SOCE was blocked. This suggests L-type Ca2+ channels might be a potential target for bronchoconstriction under certain circumstances.