Background--Inhibition of cardiac sympathetic tone represents an important strategy for treatment of cardiovascular disease, including arrhythmia, coronary heart disease, and chronic heart failure. Activation of presynaptic ₂-adrenoceptors is the most widely accepted mechanism of action of the antisympathetic drug clonidine; however, other target proteins have been postulated to contribute to the in vivo actions of clonidine.

Methods and Results--To test whether clonidine elicits pharmacological effects independent of ₂-adrenoceptors, we have generated mice with a targeted deletion of all 3 ₂-adrenoceptor subtypes (_2ABC^-/-). _2ABC^-/- mice were completely unresponsive to the analgesic and hypnotic effects of clonidine; however, clonidine significantly lowered heart rate in _2ABC^-/- mice by up to 150 bpm. Clonidine-induced bradycardia in conscious _2ABC^-/- mice was 32.3% (10 µg/kg) and 26.6% (100 µg/kg) of the effect in wild-type mice. A similar bradycardic effect of clonidine was observed in isolated spontaneously beating right atria from _2ABC-knockout and wild-type mice. Clonidine inhibited the native pacemaker current (I_f) in isolated sinoatrial node pacemaker cells and the I_f-generating hyperpolarization-activated cyclic nucleotide-gated (HCN) 2 and HCN4 channels in transfected HEK293 cells. As a consequence of blocking I_f, clonidine reduced the slope of the diastolic depolarization and the frequency of pacemaker potentials in sinoatrial node cells from wild-type and _2ABC-knockout mice.

Conclusions--Direct inhibition of cardiac HCN pacemaker channels contributes to the bradycardic effects of clonidine gene-targeted mice in vivo, and thus, clonidine-like drugs represent novel structures for future HCN channel inhibitors.