title: Real-time imaging of inflation-induced ATP release in the ex vivo rat lung.
abstract: Extracellular ATP and other nucleotides are important autocrine/paracrine mediators that regulate diverse processes critical for lung function, including mucociliary clearance, surfactant secretion, and local blood flow. Cellular ATP release is mechanosensitive; however, the impact of physical stimuli on ATP release during breathing has never been tested in intact lungs in real time and remains elusive. In this pilot study, we investigated inflation-induced ATP release in rat lungs ex vivo by real-time luciferin-luciferase (LL) bioluminescence imaging coupled with simultaneous infrared tissue imaging to identify ATP-releasing sites. With LL solution introduced into air spaces, brief inflation of such edematous lung (1 s, ∼20 cmH2O) induced transient (<30 s) ATP release in a limited number of air-inflated alveolar sacs during their recruitment/opening. Released ATP reached concentrations of ∼10-6M, relevant for autocrine/paracrine signaling, but it remained spatially restricted to single alveolar sacs or their clusters. ATP release was stimulus dependent: prolonged (100 s) inflation evoked long-lasting ATP release that terminated upon alveoli deflation/derecruitment while cyclic inflation/suction produced cyclic ATP release. With LL introduced into blood vessels, inflation induced transient ATP release in many small patchlike areas the size of alveolar sacs. Findings suggest that inflation induces ATP release in both alveoli and the surrounding blood capillary network; the functional units of ATP release presumably consist of alveolar sacs or their clusters. Our study demonstrates the feasibility of real-time ATP release imaging in ex vivo lungs and provides the first direct evidence of inflation-induced ATP release in lung air spaces and in pulmonary blood capillaries, highlighting the importance of purinergic signaling in lung function.
authors.affiliation: Mechanobiology Laboratory, Graduate School of Medicine, Nagoya University, Nagoya, Japan.
authors.forename: Kishio
authors.initials: K
authors.lastname: Furuya
authors.affiliation: Centre de recherche, Centre hospitalier de l'Université de Montréal (CRCHUM), Montreal, Quebec, Canada.
authors.forename: Ju Jing
authors.initials: JJ
authors.lastname: Tan
authors.affiliation: Centre de recherche, Centre hospitalier de l'Université de Montréal (CRCHUM), Montreal, Quebec, Canada.
authors.forename: Francis
authors.initials: F
authors.lastname: Boudreault
authors.affiliation: Mechanobiology Laboratory, Graduate School of Medicine, Nagoya University, Nagoya, Japan.
authors.forename: Masahiro
authors.initials: M
authors.lastname: Sokabe
authors.affiliation: Department of Medicine, Université de Montréal, Montreal, Quebec, Canada; and.|Institut de recherches cliniques de Montréal (IRCM), Quebec, Canada.
authors.forename: Yves
authors.initials: Y
authors.lastname: Berthiaume
authors.affiliation: Centre de recherche, Centre hospitalier de l'Université de Montréal (CRCHUM), Montreal, Quebec, Canada; ryszard.grygorczyk@umontreal.ca.|Department of Medicine, Université de Montréal, Montreal, Quebec, Canada; and.
authors.forename: Ryszard
authors.identifier: 0000-0002-9798-7651
authors.initials: R
authors.lastname: Grygorczyk
mesh_terms: D000255:Adenosine Triphosphate / Q000378:metabolism*; D000818:Animals; D002196:Capillaries / Q000378:metabolism; D003199:Computer Systems; D021621:Imaging, Three-Dimensional; D007202:Indicators and Reagents; D008168:Lung / Q000098:blood supply / Q000378:metabolism*; D008297:Male; D011312:Pressure; D011650:Pulmonary Alveoli / Q000378:metabolism; D011654:Pulmonary Edema / Q000378:metabolism / Q000473:pathology; D017208:Rats, Wistar
chemical_list: D007202:Indicators and Reagents; D000255:Adenosine Triphosphate
keywords: infrared imaging; luminescence microscopy; lung ATP release; lung inflation; mechanical stimulation