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We present a model for mimicking acute aerobic endurance exercise to freshly isolated cardiomyocytes by evoking exercise\u2010like contractions over prolonged periods of time with trains of electrical twitch stimulations. We then investigated immediate contractile, Ca<jats:sup>2+<\/jats:sup>, and metabolic responses to acute exercise in perfused freshly isolated left ventricular rat cardiomyocytes, after a matrix\u2010design optimized protocol and induced a mimic for acute aerobic endurance exercise by trains of prolonged field twitch stimulations. Acute exercise decreased cardiomyocyte fractional shortening 50%\u201380% (<jats:italic>p<\/jats:italic>\u2009&lt;\u2009.01). This was not explained by changes to intracellular Ca<jats:sup>2+<\/jats:sup> handling (<jats:italic>p<\/jats:italic>\u2009&gt;\u2009.05); rather, we observed a weak insignificant Ca<jats:sup>2+<\/jats:sup> transient increase (<jats:italic>p<\/jats:italic>\u2009=\u2009.11), while myofilament Ca<jats:sup>2+<\/jats:sup> sensitivity increased 20%\u201370% (<jats:italic>p<\/jats:italic>\u2009&lt;\u2009.05). Acidic pH 6.8 decreased fractional shortening 20%\u201370% (<jats:italic>p<\/jats:italic>\u2009&lt;\u2009.05) because of 20%\u201330% decreased Ca<jats:sup>2+<\/jats:sup> transients (<jats:italic>p<\/jats:italic>\u2009&lt;\u2009.05), but no difference occurred between control and acute exercise (<jats:italic>p<\/jats:italic>\u2009&gt;\u2009.05). Addition of 1 or 10\u2009mM La<jats:sup>\u2212<\/jats:sup> increased fractional shortening in control (1\u2009mM La<jats:sup>\u2212<\/jats:sup>: no difference, <jats:italic>p<\/jats:italic>\u2009&gt;\u2009.05; 10\u2009mM La<jats:sup>\u2212<\/jats:sup>: 20%\u201330%, <jats:italic>p<\/jats:italic>\u2009&lt;\u2009.05) and acute exercise (1\u2009mM La<jats:sup>\u2212<\/jats:sup>: 40%\u201390%, <jats:italic>p<\/jats:italic>\u2009&lt;\u2009.01; 10\u2009mM La<jats:sup>\u2212<\/jats:sup>: 50%\u2013100%, <jats:italic>p<\/jats:italic>\u2009&lt;\u2009.01) and rendered acute exercise indifferent from control (<jats:italic>p<\/jats:italic>\u2009&gt;\u2009.05). Intrinsic autofluorescence showed a resting NAD<jats:sub>state<\/jats:sub> of 0.59\u2009\u00b1\u20090.04 and FAD<jats:sub>state<\/jats:sub> of 0.17\u2009\u00b1\u20090.03, while acute exercise decreased NADH\/FAD ratio 8% (<jats:italic>p<\/jats:italic>\u2009&lt;\u2009.01), indicating intracellular oxidation. In conclusion, we show a novel approach for studying immediate acute cardiomyocyte responses to aerobic endurance exercise. We find that acute exercise in cardiomyocytes decreases contraction, but Ca<jats:sup>2+<\/jats:sup> handling and myofilament Ca<jats:sup>2+<\/jats:sup> sensitivity compensate for this, while acidosis and reduced energy substrate and mitochondrial ATP generation explain this.<\/jats:p>","DOI":"10.1002\/cbf.3847","type":"journal-article","created":{"date-parts":[[2023,9,4]],"date-time":"2023-09-04T06:40:15Z","timestamp":1693809615000},"page":"1147-1161","update-policy":"https:\/\/doi.org\/10.1002\/crossmark_policy","source":"Crossref","is-referenced-by-count":2,"title":["Electrically stimulated in vitro heart cell mimic of acute exercise reveals novel immediate cellular responses to exercise: Reduced contractility and metabolism, but maintained calcium cycling and increased myofilament calcium sensitivity"],"prefix":"10.1002","volume":"41","author":[{"ORCID":"https:\/\/orcid.org\/0000-0003-3229-5424","authenticated-orcid":false,"given":"Ana Da 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Metabolic Health, College of Medical, Veterinary and Life Sciences University of Glasgow Glasgow UK"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Karen","family":"McGlynn","sequence":"additional","affiliation":[{"name":"School of Cardiovascular and Metabolic Health, College of Medical, Veterinary and Life Sciences University of Glasgow Glasgow UK"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Andrew","family":"McNair","sequence":"additional","affiliation":[{"name":"School of Cardiovascular and Metabolic Health, College of Medical, Veterinary and Life Sciences University of Glasgow Glasgow UK"}],"role":[{"vocabulary":"crossref","role":"author"}]},{"given":"Peter","family":"Bowman","sequence":"additional","affiliation":[{"name":"School of Cardiovascular and Metabolic Health, College of Medical, Veterinary and Life Sciences University of Glasgow Glasgow 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