Immunostaining of rat brain, spinal cord, sensory neurons and skeletal muscle for calcium channel alpha2-delta (α2-δ) type 1 protein

Alpha2-delta (α2-δ) is a membrane-spanning auxiliary protein subunit of voltage-gated calcium channels found in muscle and brain. Of the four subtypes of α2-δ, only α2-δ types 1 and 2 (α2-δ-1 and α2-δ-2) bind the drugs gabapentin (Neurontin®) and pregabalin (Lyrica®). Although recent findings indicate that drug binding to α2-δ-1 is required for pharmacology of pregabalin and gabapentin, previous work has not addressed the location of α2-δ-1 protein within nervous tissues. A monoclonal antibody to α2-δ-1 revealed intense immunostaining in certain areas of rat brain, spinal cord, dorsal root ganglia, and skeletal muscle, with weaker staining in heart muscle, gut and liver. Little immunostaining was seen in spleen, kidney, thymus and lung. Staining was dense in some regions of the CNS including spinal dorsal horn, anterior olfactory nucleus, anterior amygdala, basolateral (ventral) amygdala and cortical amygdala, and the piriform, perirhinal, insular and entorhinal cortices. In hippocampus, staining was heterogeneous with greater density in areas of glutamate terminals (mossy fiber endings on CA3 pyramidal cells and perforant path endings on granule cells and CA1 stratum radiatum). Moderate staining occurred in the lateral posterior nucleus of the thalamus, superficial layers of neocortex, periaqueductal gray, substantia nigra, stria terminalis, nucleus accumbens shell and tegmental nucleus. We propose that areas of dense α2-δ-1 staining in brain and spinal cord are likely sites of action for the analgesic, anticonvulsant and anxiolytic-like actions of pregabalin and gabapentin in animal models.