Auditory brainstem neurons that are involved in the precise analysis of the temporal pattern of sounds have ionic currents activated near the resting potential to shorten membrane time constants. One of these currents is the hyperpolarization-activated current (Ih). Molecular cloning of the channels underlying Ih revealed four different isoforms (HCN1–4). HCN1 and HCN2, which are widely distributed in the brain, differ in their activation kinetics, voltage dependence and sensitivity to cAMP. We determined the distribution of the HCN1 and HCN2 isoform in the auditory brainstem and midbrain of young rats (P20–30), using standard immunohistochemical techniques. HCN1 antibodies gave rise to punctate staining on the somatic and dendritic membrane. Strong HCN1 staining was present on octopus and bushy cells of the ventral cochlear nucleus, principal neurons of the lateral and medial superior olive, and neurons of the ventral nucleus of the lateral lemniscus. No HCN1 staining was observed in the dorsal cochlear nucleus and the medial nucleus of the trapezoid body (MNTB). In contrast, HCN2 staining was strongest in the MNTB and the dorsal nucleus of the lateral lemniscus. Strong HCN2 antibody labelling was also observed in bushy cells of the ventral cochlear nucleus. In the central nucleus of the inferior colliculus only a subpopulation of neurons showed HCN1 or HCN2 immunolabelling. This differential distribution of HCN1 and HCN2 channels is in agreement with the physiologically observed Ih currents in corresponding neuronal populations and might represent the basis for functional heterogeneity and diverse sensitivity to neuromodulators.