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Hyperactivation of RyR by genetic mutations or posttranslational modification can cause various skeletal muscle and arrhythmogenic heart diseases. Inhibitors of RyR are therefore expected to be potential drugs for treatment of such diseases. This article describes assays to evaluate RyR channel activity, including an ER Ca<jats:sup>2+<\/jats:sup> measurement assay that is compatible with high\u2010throughput screening and a [<jats:sup>3<\/jats:sup>H]\u2010ryanodine binding assay that provides a quantitative measure of RyR channel activity as a second screen for compound hits. \u00a9 2019 by John Wiley &amp; Sons, Inc.<\/jats:p><jats:p><jats:bold>Basic Protocol 1<\/jats:bold>: [Ca<jats:sup>2+<\/jats:sup>]<jats:sub>ER<\/jats:sub> assay for ryanodine receptor (RyR) channel activity<\/jats:p><jats:p><jats:bold>Support Protocol 1<\/jats:bold>: Determination of dose dependence and isoform selectivity of RyR inhibitors using [Ca<jats:sup>2+<\/jats:sup>]<jats:sub>ER<\/jats:sub> assay<\/jats:p><jats:p><jats:bold>Basic Protocol 2<\/jats:bold>: [<jats:sup>3<\/jats:sup>H]\u2010Ryanodine binding assay for RyR channel activity<\/jats:p><jats:p><jats:bold>Support Protocol 2<\/jats:bold>: Isolation of microsomes from RyR\u2010expressing HEK293 cells<\/jats:p>","DOI":"10.1002\/cpph.71","type":"journal-article","created":{"date-parts":[[2019,12,13]],"date-time":"2019-12-13T14:29:43Z","timestamp":1576247383000},"update-policy":"https:\/\/doi.org\/10.1002\/crossmark_policy","source":"Crossref","is-referenced-by-count":25,"title":["Assays for Modulators of Ryanodine Receptor (RyR)\/Ca<sup>2+<\/sup> Release Channel Activity for Drug Discovery for Skeletal Muscle and Heart Diseases"],"prefix":"10.1002","volume":"87","author":[{"given":"Takashi","family":"Murayama","sequence":"first","affiliation":[{"name":"Department of Pharmacology Juntendo University School of Medicine  Tokyo Japan"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Nagomi","family":"Kurebayashi","sequence":"additional","affiliation":[{"name":"Department of Pharmacology Juntendo University School of Medicine  Tokyo 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