Perikaryal and synaptic localization of α_2A-adrenergic receptor-like immunoreactivity

Through molecular cloning, the existence of three distinct subtypes of α₂-adrenergic receptors (α₂AR) - A, B and C - has been established and are referred to as α_2AAR, α_2BAR and α_2CAR. Due to limitations in pharmacological tools, it has been difficult to ascribe the role of each subtype to the central functions of α₂AR. In situ hybridization studies have provided valuable information regarding their distribution within brain. However, little is known about their subcellular distribution, and in particular, their pre- versus postsynaptic localization or their relation to noradrenergic neurons in the CNS. We used an antiserum that selectively recognizes the A-subtype of α₂AR to determine: (1) the regional distribution of the receptor within brains of rat and monkey; (2) the subcellular distribution of the receptor in locus coeruleus (LC) of rats and prefrontal cortex of monkeys; and (3) the ultrastructural relation of the receptor to noradrenergic processes in LC. Light microscopic immunocytochemistry revealed prominent immunoreactivity in LC, the brainstem regions modulating the baroreflex, the granule cell layer of the cerebellar cortex, the paraventricular and supraoptic nuclei of the hypothalamus (PVN, SON), the basal ganglia, all thalamic nuclei, the hippocampal formation and throughout cerebral cortical areas. Comparison of results obtained from rat and monkey brains revealed no apparent interspecies-differences in the regional distribution of immunoreactivity. Immunoreactivity occurred as small puncta, less than 1 μm in diameter, that cluster over neuronal perikarya. Besides these puncta, cell bodies, proximal dendrites and fine varicose processes - most likely to be axonal - of the PVN and SON and the hippocampal granule cells also exhibited homogeneously intense distribution of immunoreactivity. Subcellularly, α_2AAR-ir in LC and prefrontal cortex were associated with synaptic and non-synaptic plasma membrane of dendrites and perikarya as well as perikaryal membranous organelles. In addition, cortical tissue, but not LC, exhibited prominent immunoreactivity within spine heads. Rat brainstem tissue immunolabeled dually for α_2AAR and dopamine β-hydroxylase (DβH, the noradrenaline-synthesizing enzyme) revealed that α_2AAR-li occurs in catecholaminergic terminals but is also prevalent within non-catecholaminergic terminals. Terminals exhibiting α_2AAR-li formed symmetric and asymmetric types of synapses onto dendrites with and without DβH-immunoreactivity. These results indicate that: (1) the A-subtype of α₂AR is distributed widely within brain; (2) α_2AAR-li reflects the presence of newly synthesized α_2AAR in perikarya as well as those receptors along the plasma membrane of perikarya, dendritic trunks and spines; and (3) α_2AAR in LC may operate as heteroreceptors on non-catecholaminergic terminals as well as autoreceptors on noradrenergic terminals.