Autoradiograms were developed from free running strands of Purkinje fibers and from left ventricular myocardium of dog hearts exposed to [7-methoxy- 3 H]prazosin or [O-methyl- 3 H]yohimbine in the presence or absence of excessive non-radioactive ligands. Both Purkinje fiber and ventricular myocardium showed high density, tissue-specific binding to [ 3 H]prazosin. In contrast, high density, tissue-specific binding to [ 3 H]yohimbine was present in Purkinje fibers but not in ventricular myocardium. Membrane fractions showed high affinity, saturable, and displaceable binding with [ 3 H]yohimbine in preparations from canine cardiac Purkinje fibers but not those from canine cardiac ventricular myocardium. Scatchard analysis of the canine Purkinje membrane α 2 -adrenergic receptor binding showed a B m a x of 54.9 fmol/mg protein with a K d of 6.25 nM. These results confirm the electrophysiological findings that post-junctional α 2 -adrenergic receptors are present in Purkinje fibers.