The alpha-2 receptor agonist Dexmedetomidine (Dex) protects the heart against ischemia–reperfusion injury. We investigated the signaling cascade underlying Dex-induced acute cardioprotection, with special emphasis on large-conductance Ca2+-sensitive potassium (BKCa) channels. Rats were anesthetized with pentobarbital. Hearts were isolated, mounted on a Langendorff system and perfused with Krebs–Henseleit buffer. Hearts underwent 33 minutes of ischemia followed by 60 minutes of reperfusion. Before the beginning of ischemia, Dex was administered at different doses (0.1–30 nM) for characterization of a dose-effect relationship. In another set of experiments, Dex (3 nM) was administered together with the BKCa channel inhibitor paxilline and the connexin-43 inhibitor peptide Gap27. Also, the BKCa channel opener NS1619 was administered. In control animals, infarct size was 49% ± 5%. Dex at 3–30 nM reduced infarct size to ∼22%, whereas lower (0.1–1 nM) doses reduced infarct size to ∼38%. Paxilline (1 μM) and GAP27 (6 μM) blocked the Dex-induced cardioprotection. NS1619 (10 μM) reduced infarct size to about the same magnitude as did the higher doses of Dex. Functional heart parameters and coronary flow were not different between the study groups. In male rats, the Dex-induced protection against ischemia–reperfusion injury involves connexin-43 and activation of BKCa channels.