Genetic and Regulatory Disruptions Impacting Dopamine Transporter Function Contribute to Neuropsychiatric Disease Phenotypes
Dopamine (DA) signaling plays an important role in the central nervous system by regulating a variety of functions including attention, motivation, reward, movement and courtship. Alterations in DA signaling have been implicated in numerous neuropsychiatric disorders. Several synaptic proteins fine-tune central DA homeostasis of which the DA transporter (DAT), a presynaptic Na+/Cl- dependent transporter of the solute carrier 6 gene family, plays a key role. The DAT is capable of numerous structural and functional modalities that are important to its role in maintaining DA homeostasis at the synapse. Of these, the ability of DAT to clear extracellular DA following vesicular release through an uptake process has been widely studied in multiple cell-based and in vivo systems. Additionally, DAT-mediated DA efflux allows for the movement of cytosolic DA to the synaptic space, leading to accumulation of extracellular DA. This accumulation occurs under cellular conditions like membrane depolarization and kinase activation, as well as in response to psychostimulant drugs like amphetamine (AMPH). In recent years, we and others have started to unveil how DAT functional modalities are regulated, in an effort to reverse adverse phenotypes promoted by pathological alterations in DAT function. We investigated a rare, autism-associated deletion variant of the human DAT (hDAT) at Asn336 (∆N336). We discovered a previously uncovered conformation of the transporter (a half-open, inward facing state) that completely deprives DAT of uptake function, resulting in dysregulated DA homeostasis. Next, we sought insights into the mechanisms underlying DAT-mediated DA efflux. We found that DA efflux involves the action of key kinases phosphorylating the DAT and its associated protein syntaxin 1. Lastly, we demonstrated that the cathinone 3,4-methylenedioxypyrovalerone (MDPV) elevates synaptic DA levels by mimicking actions of both cocaine and AMPH. Across all our studies, we describe both the mechanisms underlying elevations in synaptic DA and also the behavioral consequences of enhanced DA signaling in Drosophila. Expression in flies of hDAT ∆N336, pseudophosphorylated hDAT, as well as administration of AMPH or MDPV resulted in aberrant behaviors such as hyperactivity, enhanced grooming and fear-response, impaired social interactions and heightened sexual impetus that we describe as mediated by increases in synaptic DA levels.