Calmodulin kinase II interacts with the dopamine transporter C terminus to regulate amphetamine-induced reverse transport

Efflux of dopamine through the dopamine transporter (DAT) is critical for the psychostimulatory properties of amphetamines, but the underlying mechanism is unclear. Here we show that Ca2+/calmodulin-dependent protein kinase II (CaMKII) plays a key role in this efflux. CaMKII alpha bound to the distal C terminus of DAT and colocalized with DAT in dopaminergic neurons. CaMKII alpha stimulated dopamine efflux via DAT in response to amphetamine in heterologous cells and in dopaminergic neurons. CaMKII alpha phosphorylated serines in the distal N terminus of DAT in vitro, and mutation of these serines eliminated the stimulatory effects of CaMKII alpha. A mutation of the DAT C terminus impairing CaMKII alpha binding also impaired amphetamine-induced dopamine efflux. An in vivo role for CaMKII was supported by chronoamperometry measurements showing reduced amphetamine-induced dopamine efflux in response to the CaMKII inhibitor KN93. Our data suggest that CaMKII alpha binding to the DAT C terminus facilitates phosphorylation of the DAT N terminus and mediates amphetamine-induced dopamine efflux.