We previously characterized 1-ethyl-2-benzimidazolinone (1-EBIO), as well as the clinically useful benzoxazoles, chlorzoxazone (CZ), and zoxazolamine (ZOX), as pharmacological activators of the intermediate-conductance Ca²⁺-activated K⁺ channel, hIK1. The mechanism of activation of hIK1, as well as the highly homologous small-conductance, Ca²⁺-dependent K⁺ channel, rSK2, was determined following heterologous expression inXenopus oocytes using two-electrode voltage clamp (TEVC) and excised, inside-out patch-clamp techniques. 1-EBIO, CZ, and ZOX activated both hIK1 and rSK2 in TEVC and excised inside-out patch-clamp experiments. In excised, inside-out patches, 1-EBIO and CZ induced a concentration-dependent activation of hIK1, with half-maximal (K _1/2) values of 84 μM and 98 μM, respectively. Similarly, CZ activated rSK2 with a K _1/2 of 87 μM. In the absence of CZ, the Ca²⁺-dependent activation of hIK1 was best fit with a K _1/2 of 700 nM and a Hill coefficient (n) of 2.0. rSK2 was activated by Ca²⁺ with a K _1/2 of 700 nM and ann of 2.5. Addition of CZ had no effect on either theK _1/2 or n for Ca²⁺-dependent activation of either hIK1 or rSK2. Rather, CZ increased channel activity at all Ca²⁺ concentrations (V _max). Event-duration analysis revealed hIK1 was minimally described by two open and three closed times. Activation by 1-EBIO had no effect on τ_o1, τ_o2, or τ_c1, whereas τ_c2 and τ_c3 were reduced from 9.0 and 92.6 ms to 5.0 and 44.1 ms, respectively. In conclusion, we define 1-EBIO, CZ, and ZOX as the first known activators of hIK1 and rSK2. Openers of IK and SK channels may be therapeutically beneficial in cystic fibrosis and vascular diseases.