Involvement of Allosteric Effect and K(Ca) Channels in Crosstalk between β(2)-Adrenergic and Muscarinic M(2) Receptors in Airway Smooth Muscle

To advance the development of bronchodilators for asthma and chronic obstructive pulmonary disease (COPD), this study was designed to investigate the mechanism of functional antagonism between β(2)-adrenergic and muscarinic M(2) receptors, focusing on allosteric effects and G proteins/ion channels coupling. Muscarinic receptor antagonists (tiotropium, glycopyrronium, atropine) synergistically enhanced the relaxant effects of β(2)-adrenergic receptor agonists (procaterol, salbutamol, formoterol) in guinea pig trachealis. This crosstalk was inhibited by iberitoxin, a large-conductance Ca(2+)-activated K(+) (K(Ca)) channel inhibitor, whereas it was increased by verapamil, a L-type voltage-dependent Ca(2+) (VDC) channel inhibitor; additionally, it was enhanced after tissues were incubated with pertussis or cholera toxin. This synergism converges in the G proteins (G(i), G(s))/K(Ca) channel/VDC channel linkages. Muscarinic receptor antagonists competitively suppressed, whereas, β(2)-adrenergic receptor agonists noncompetitively suppressed muscarinic contraction. In concentration-inhibition curves for β(2)-adrenergic receptor agonists with muscarinic receptor antagonists, EC(50) was markedly decreased, and maximal inhibition was markedly increased. Hence, muscarinic receptor antagonists do not bind to allosteric sites on muscarinic receptors. β(2)-Adrenergic receptor agonists bind to allosteric sites on these receptors; their intrinsic efficacy is attenuated by allosteric modulation (partial agonism). Muscarinic receptor antagonists enhance affinity and efficacy of β(2)-adrenergic action via allosteric sites in β(2)-adrenergic receptors (synergism). In conclusion, K(Ca) channels and allosterism may be novel targets of bronchodilator therapy for diseases such as asthma and COPD.