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Various GABAAR (\u03b3-aminobutyric acid type A receptor) activators, including benzodiazepines, relax ASM. The GABAAR is a ligand-operated Cl\u2212 channel best known for its role in inhibitory neurotransmission in the central nervous system. Although ASM cells express GABAARs, affording a seemingly logical site of action, the mechanism(s) by which GABAAR ligands relax ASM remains unclear. PI320, a novel imidazobenzodiazepine designed for tissue selectivity, is a promising asthma drug candidate. Here, we show that PI320 alleviates methacholine (MCh)-induced bronchoconstriction in vivo and relaxes peripheral airways preconstricted with MCh ex vivo using the forced oscillation technique and precision-cut lung slice experiments, respectively. Surprisingly, the peripheral airway relaxation demonstrated in precision-cut lung slices does not appear to be GABAAR-dependent, as it is not inhibited by the GABAAR antagonist picrotoxin or the benzodiazepine antagonist flumazenil. Furthermore, we demonstrate here that PI320 inhibits MCh-induced airway constriction in the absence of external Ca2, suggesting that PI320-mediated relaxation is not mediated by inhibition of Ca2+ influx in ASM. However, PI320 does inhibit MCh-induced intracellular Ca2+ oscillations in peripheral ASM, a key mediator of contraction that is dependent on sarcoplasmic reticulum Ca2+ mobilization. Furthermore, PI320 inhibits peripheral airway constriction induced by experimentally increasing the intracellular concentration of inositol triphosphate (IP3). These novel data suggest that PI320 relaxes murine peripheral airways by inhibiting intracellular Ca2+ mobilization in ASM, likely by inhibiting Ca2+ release through IP3Rs (IP3 receptors).<\/jats:p>","DOI":"10.1165\/rcmb.2022-0084oc","type":"journal-article","created":{"date-parts":[[2022,7,1]],"date-time":"2022-07-01T11:57:55Z","timestamp":1656676675000},"page":"482-490","update-policy":"https:\/\/doi.org\/10.1164\/crossmarkpolicies","source":"Crossref","is-referenced-by-count":7,"title":["Imidazobenzodiazepine PI320 Relaxes Mouse Peripheral Airways by Inhibiting Calcium Mobilization"],"prefix":"10.1093","volume":"67","author":[{"given":"Jose F.","family":"Perez-Zoghbi","sequence":"first","affiliation":[{"name":"Department of Anesthesiology, Vagelos College of Physicians and Surgeons, Columbia University, New York, New York; and"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Dannah Rae","family":"Sajorda","sequence":"additional","affiliation":[{"name":"Department of Anesthesiology, Vagelos College of Physicians and Surgeons, Columbia University, New York, New York; and"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Daniel A.","family":"Webb","sequence":"additional","affiliation":[{"name":"Department of Chemistry and Biochemistry and"},{"name":"Milwaukee Institute for Drug Discovery, University of Wisconsin\u2013Milwaukee, Milwaukee, Wisconsin"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Leggy A.","family":"Arnold","sequence":"additional","affiliation":[{"name":"Department of Chemistry and Biochemistry and"},{"name":"Milwaukee Institute for Drug Discovery, University of Wisconsin\u2013Milwaukee, Milwaukee, Wisconsin"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Charles W.","family":"Emala","sequence":"additional","affiliation":[{"name":"Department of Anesthesiology, Vagelos College of Physicians and Surgeons, Columbia University, New York, New York; and"}],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Gene T.","family":"Yocum","sequence":"additional","affiliation":[{"name":"Department of Anesthesiology, Vagelos College of Physicians and Surgeons, Columbia University, New York, New York; and"}],"role":[{"role":"author","vocabulary":"crossref"}]}],"member":"286","published-online":{"date-parts":[[2022,10,1]]},"reference":[{"key":"2026012220022187700_bib1","doi-asserted-by":"crossref","first-page":"435","DOI":"10.1016\/j.mcna.2018.12.004","article-title":"Asthma in the primary care setting","volume":"103","author":"Wu","year":"2019","journal-title":"Med Clin North Am"},{"key":"2026012220022187700_bib2","author":"National Center for Health Statistics","year":"2020"},{"key":"2026012220022187700_bib3","doi-asserted-by":"crossref","first-page":"1766","DOI":"10.1021\/acs.molpharmaceut.7b01013","article-title":"A novel orally available asthma drug candidate that reduces smooth muscle constriction and inflammation by 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