Effects of azelastine on contraction of guinea pig tracheal smooth muscle

Azelastine [4-(p-chlorobenzyl)-2-(hexahydro-1-methyl-1H-azepine-4-yl)-1(2H)-phthalazinone hydrochloride] is a new anti-asthmatic drug. We examined the mechanism of its inhibitory action on guinea pig tracheal smooth muscle contraction by measuring membrane potential and isometric force using intracellular microelectrodes and a micro-force transducer. The mean resting membrane potential of guinea pig tracheal muscle cells was -54 mV. Perfusion with 20 mM tetraethylammonium (TEA) caused membrane depolarization and elicited spontaneous action potentials. Azelastine (1-100 microM) suppressed both the amplitude and maximal rate of rise of the action potentials in a concentration-dependent manner. Complete abolition occurred at 100 microM. Similarly, azelastine (0.1-100 microM) inhibited and abolished 50 mM KCl-induced contractions. These results suggest that azelastine may inhibit voltage-dependent Ca2+ channels. Next, pretreatment of tracheal muscle (for 15 min) with azelastine (0.01-100 microM) inhibited subsequent acetylcholine (ACh) (0.01-100 microM)-induced contractions. Azelastine, 100 microM, completely abolished the ACh-induced contractions. In contrast, high concentrations of Ca2+ channel antagonists diltiazem (10-100 microM) or nifedipine (20 microM), and Ca2(+)-free solution, only partially depressed the ACh contractions suggesting that azelastine has an additional effect on intracellular Ca2+ release. In Ca2(+)-free solution (containing 0.5 mM EGTA), azelastine (1-100 microM) depressed and abolished the transient contractions induced by 10 microM ACh. We conclude that azelastine inhibits airway constriction by inhibiting both voltage-sensitive Ca2+ slow channels on the cell membrane and Ca2+ release from a intracellular storage site.