Elucidating the presynaptic roles of the α2δ family of proteins

Synapses are the fundamental unit of information transfer in the central nervous system (CNS) and are composed of highly complex molecular machinery that tightly regulates synaptic transmission and neuronal circuit output. The α2δ proteins (α2δ1-4) are extracellular proteins initially identified as auxiliary subunits of voltage-gated Ca2+ (CaV) channel complexes. Mutations in α2δ1-4 are linked to a wide range of disorders including neuropathic pain and epilepsy, and α2δ1-2 are targeted by drugs used to treat these disorders. Multiple roles have been described for the α2δ isoforms in independently regulating synaptic function and CaV channel complexes. Many CNS synapses contain a mixture of α2δ1-3, which are found in both the pre and postsynaptic compartments, with α2δ4 mainly restricted to the retina. Currently, studies of α2δ are mostly done using global mutant/knock out (KO) models or in cell culture. While powerful, these studies prevent conclusions on the presynaptic functions of α2δ in an in vivo circuit. To elucidate the presynaptic regulatory roles of α2δ1-3 in a native neuronal circuit, we developed a novel α2δ1-3 conditional KO mouse model and ablated α2δ1-3 at the calyx of Held, a large glutamatergic axosomatic synapse in the lower auditory brainstem. Using this animal model, we ablated all three isoforms using a Helper-Dependent Adenovirus that expresses Cre recombinase at postnatal day (P)1. Subsequently, using a multidisciplinary approach, we analyzed how loss of α2δ1-3 at the adult stage (P18 onwards) calyx of Held impacted synaptic function. The data presented herein defines the roles of presynaptic α2δ-1, 2, and 3 and proposes a new role regulating Munc13 abundance. It has also laid the foundation for several avenues of exploration that have the potential to answer some of the defining questions in this subfield.