Neurotrophins [e.g., brain-derived neurotrophic factor (BDNF), neurotrophin 4 (NT4)], known to affect neuronal structure and function, are expressed in nonneuronal tissues including the airway. However, their function is unclear. We examined the effect of acute vs. prolonged neurotrophin exposure on regulation of airway smooth muscle (ASM) intracellular Ca2+concentration ([Ca2+]i): sarcoplasmic reticulum (SR) Ca2+release and Ca2+influx (specifically store-operated Ca2+entry, SOCE). Human ASM cells were incubated for 30 min in medium (control) or 1 or 10 nM BDNF, NT3, or NT4 (acute exposure) or overnight in 1 nM BDNF, NT3, or NT4 (prolonged exposure) and imaged after loading with the Ca2+indicator fura-2 AM. [Ca2+]iresponses to ACh, histamine, bradykinin, and caffeine and SOCE following SR Ca2+depletion were compared across cell groups. Force measurements were performed in human bronchial strips exposed to neurotrophins. Basal [Ca2+]i, peak responses to all agonists, SOCE, and force responses to ACh and histamine were all significantly enhanced by both acute and prolonged BDNF exposure (smaller effect of NT4) but decreased by NT3. Inhibition of the BDNF/NT4 receptor trkB by K252a prevented enhancement of [Ca2+]iresponses. ASM cells showed positive immunostaining for BDNF, NT3, NT4, trkB, and trkC (NT3 receptor). These novel data demonstrate that neurotrophins influence ASM [Ca2+]iand force regulation and suggest a potential role for neurotrophins in airway diseases.