{"status":"ok","message-type":"work","message-version":"1.0.0","message":{"indexed":{"date-parts":[[2025,4,24]],"date-time":"2025-04-24T04:50:23Z","timestamp":1745470223318,"version":"3.40.4"},"reference-count":26,"publisher":"S. Karger AG","issue":"4","license":[{"start":{"date-parts":[[2009,1,1]],"date-time":"2009-01-01T00:00:00Z","timestamp":1230768000000},"content-version":"vor","delay-in-days":0,"URL":"https:\/\/www.karger.com\/Services\/SiteLicenses"},{"start":{"date-parts":[[2009,1,1]],"date-time":"2009-01-01T00:00:00Z","timestamp":1230768000000},"content-version":"tdm","delay-in-days":0,"URL":"https:\/\/www.karger.com\/Services\/SiteLicenses"}],"content-domain":{"domain":[],"crossmark-restriction":false},"short-container-title":["Pharmacology"],"published-print":{"date-parts":[[2009]]},"abstract":"Muscarinic acetylcholine receptor (mAChR)-mediated guanine nucleotide-binding regulatory protein (G protein) activation and the functional interaction between receptors and the respective G proteins were investigated using an agonist-induced [<sup>35<\/sup>S]guanosine-5\u2032-<i>O<\/i>-(\u03b3-thiotriphosphate) ([<sup>35<\/sup>S]GTP\u03b3S)-binding approach in membranes of 3 native equine airway segments (trachea, bronchus and lung), which differ tremendously in mAChR density and subtype distribution; especially subtypes that couple negatively to adenylyl cyclase through G<sub>i\/0<\/sub> proteins, i.e. M<sub>2<\/sub> receptors. The assay was initially optimized by determining the influence of incubation time, guanosine 5\u2032-diphosphate (GDP), MgCl<sub>2<\/sub> and NaCl on basal and agonist-stimulated [<sup>35<\/sup>S]GTP\u03b3S binding. In standard assays, the presence of 10 \u03bcmol\/l GDP, 10 mmol\/l MgCl<sub>2<\/sub> and 200 mmol\/l NaCl increased carbachol-induced specific [<sup>35<\/sup>S]GTP\u03b3S binding in a segment- and receptor-density-dependent manner. Hereby, mAChR agonists stimulated [<sup>35<\/sup>S]GTP\u03b3S binding with a rank order of potency (oxotremorine M > carbachol > acetylcholine), and in a specific segment- and receptor-density-dependent manner (trachea > bronchus > lung). The increase in the specific [<sup>35<\/sup>S]GTP\u03b3S binding was potently inhibited by the mAChR antagonist atropine. Pertussis toxin and <i>N<\/i>-ethylmaleimide as G<sub>i\/0<\/sub> protein ADP-ribosylating and alkylating agents reduced basal and agonist-stimulated [<sup>35<\/sup>S]GTP\u03b3S binding. The mAChR-stimulated [<sup>35<\/sup>S]GTP\u03b3S binding serves as a useful method for investigating the functional interaction between mAChRs and their respective G proteins in native airway tissue membranes of equines.<\/jats:p>","DOI":"10.1159\/000209254","type":"journal-article","created":{"date-parts":[[2009,3,25]],"date-time":"2009-03-25T14:16:54Z","timestamp":1237990614000},"page":"247-258","source":"Crossref","is-referenced-by-count":0,"title":["Segment-Dependent Activation of Muscarinic Acetylcholine Receptor-Mediated [<sup>35<\/sup>S]Guanosine-5\u2032-<i>O<\/i>-(\u03b3-Thiotriphosphate) Binding in Airway Tissue Membranes"],"prefix":"10.1159","volume":"83","author":[{"given":"Peter","family":"Hajek","sequence":"first","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Fritz Rupert","family":"Ungemach","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]},{"given":"Getu","family":"Abraham","sequence":"additional","affiliation":[],"role":[{"role":"author","vocabulary":"crossref"}]}],"member":"127","published-online":{"date-parts":[[2009,3,19]]},"reference":[{"key":"ref1","doi-asserted-by":"publisher","DOI":"10.1016%2F0163-7258%2893%2990027-B"},{"key":"ref2","doi-asserted-by":"publisher","DOI":"10.1016\/j.siny.2006.07.005"},{"key":"ref3","doi-asserted-by":"publisher","DOI":"10.1016%2FS0014-2999%2898%2900585-8"},{"key":"ref4","doi-asserted-by":"publisher","DOI":"10.1007%2Fs11259-006-3396-z"},{"key":"ref5","doi-asserted-by":"publisher","DOI":"10.1016%2F0014-2999%2888%2990589-4"},{"key":"ref6","doi-asserted-by":"publisher","DOI":"10.1016%2F0024-3205%2882%2990449-0"},{"key":"ref7","doi-asserted-by":"publisher","DOI":"10.1016%2F0024-3205%2893%2990310-Y"},{"key":"ref8","doi-asserted-by":"publisher","DOI":"10.1016%2FS0024-3205%2897%2900048-9"},{"key":"ref9","doi-asserted-by":"publisher","DOI":"10.1016%2FS1471-4892%2801%2900039-X"},{"key":"ref10","doi-asserted-by":"publisher","DOI":"10.1016%2FS0024-3205%2801%2901053-0"},{"key":"ref11","doi-asserted-by":"publisher","DOI":"10.1378%2Fchest.96.6.1285"},{"key":"ref12","doi-asserted-by":"publisher","DOI":"10.1016%2Fj.lfs.2003.07.005"},{"key":"ref13","doi-asserted-by":"publisher","DOI":"10.1016%2F0922-4106%2889%2990006-0"},{"key":"ref14","doi-asserted-by":"publisher","DOI":"10.1046%2Fj.1471-4159.1996.67062549.x"},{"key":"ref15","doi-asserted-by":"publisher","DOI":"10.1007%2FBF00169072"},{"key":"ref16","doi-asserted-by":"publisher","DOI":"10.1016%2F0922-4106%2892%2990026-R"},{"key":"ref17","doi-asserted-by":"publisher","DOI":"10.1016%2FS0014-2999%2899%2900827-4"},{"key":"ref18","doi-asserted-by":"publisher","DOI":"10.1073%2Fpnas.82.2.580"},{"key":"ref19","doi-asserted-by":"publisher","DOI":"10.1016%2FS0165-6147%2802%2900027-5"},{"key":"ref20","doi-asserted-by":"publisher","DOI":"10.1074%2Fjbc.273.27.16865"},{"key":"ref21","doi-asserted-by":"publisher","DOI":"10.1111%2Fj.1432-1033.1992.tb16891.x"},{"key":"ref22","doi-asserted-by":"publisher","DOI":"10.1016%2FS0076-6879%2894%2937048-6"},{"key":"ref23","doi-asserted-by":"publisher","DOI":"10.1016%2Fj.phrs.2008.04.006"},{"key":"ref24","doi-asserted-by":"publisher","DOI":"10.1146%2Fannurev.bi.56.070187.003151"},{"key":"ref25","doi-asserted-by":"publisher","DOI":"10.1038%2Fsj.bjp.0703382"},{"key":"ref26","doi-asserted-by":"publisher","DOI":"10.1038%2F288709a0"}],"container-title":["Pharmacology"],"original-title":[],"language":"en","link":[{"URL":"https:\/\/www.karger.com\/Article\/Pdf\/209254","content-type":"application\/pdf","content-version":"vor","intended-application":"text-mining"},{"URL":"https:\/\/www.karger.com\/Article\/Pdf\/209254","content-type":"unspecified","content-version":"vor","intended-application":"similarity-checking"}],"deposited":{"date-parts":[[2025,4,23]],"date-time":"2025-04-23T23:57:31Z","timestamp":1745452651000},"score":1,"resource":{"primary":{"URL":"https:\/\/karger.com\/article\/doi\/10.1159\/000209254"}},"subtitle":[],"short-title":[],"issued":{"date-parts":[[2009]]},"references-count":26,"journal-issue":{"issue":"4","published-online":{"date-parts":[[2009,4,1]]}},"URL":"https:\/\/doi.org\/10.1159\/000209254","archive":["Portico"],"relation":{},"ISSN":["0031-7012","1423-0313"],"issn-type":[{"type":"print","value":"0031-7012"},{"type":"electronic","value":"1423-0313"}],"subject":[],"published":{"date-parts":[[2009]]}}}