ErbB4 signaling in dopaminergic axonal projections increases extracellular dopamine levels and regulates spatial/working memory behaviors.

Genetic variants of Neuregulin 1 (NRG1) and its neuronal tyrosine kinase receptor ErbB4 are associated with risk for schizophrenia, a neurodevelopmental disorder characterized by excitatory/inhibitory imbalance and dopamine (DA) dysfunction. To date, most ErbB4 studies have focused on GABAergic interneurons in the hippocampus and neocortex, particularly fast-spiking parvalbumin-positive (PV+) basket cells. However, NRG has also been shown to modulate DA levels, suggesting a role for ErbB4 signaling in dopaminergic neuron function. Here we report that ErbB4 in midbrain DAergic axonal projections regulates extracellular DA levels and relevant behaviors. Mice lacking ErbB4 in tyrosine hydroxylase-positive (TH+) neurons, but not in PV+ GABAergic interneurons, exhibit different regional imbalances of basal DA levels and fail to increase DA in response to local NRG1 infusion into the dorsal hippocampus, medial prefrontal cortex and dorsal striatum measured by reverse microdialysis. Using Lund Human Mesencephalic (LUHMES) cells, we show that NRG/ErbB signaling increases extracellular DA levels, at least in part, by reducing DA transporter (DAT)-dependent uptake. Interestingly, TH-Cre;ErbB4f/f mice manifest deficits in learning, spatial and working memory-related behaviors, but not in numerous other behaviors altered in PV-Cre;ErbB4f/f mice. Importantly, microinjection of a Cre-inducible ErbB4 virus (AAV-ErbB4.DIO) into the mesencephalon of TH-Cre;ErbB4f/f mice, which selectively restores ErbB4 expression in DAergic neurons, rescues DA dysfunction and ameliorates behavioral deficits. Our results indicate that direct NRG/ErbB4 signaling in DAergic axonal projections modulates DA homeostasis, and that NRG/ErbB4 signaling in both GABAergic interneurons and DA neurons contribute to the modulation of behaviors relevant to psychiatric disorders.

Gßγ subunit activation promotes dopamine efflux through the dopamine transporter.

The dopamine transporter (DAT) is an important regulator of brain dopamine (DA) homeostasis, controlling the intensity and duration of DA signaling. DAT is the target for psychostimulants-like cocaine and amphetamine-and plays an important role in neuropsychiatric disorders, including attention-deficit hyperactivity disorder and drug addiction. Thus, a thorough understanding of the mechanisms that regulate DAT function is necessary for the development of clinical interventions to treat DA-related brain disorders. Previous studies have revealed a plethora of protein-protein interactions influencing DAT cellular localization and activity, suggesting that the fine-tuning of DA homeostasis involves multiple mechanisms. We recently reported that G-protein beta-gamma (Gßγ) subunits bind directly to DAT and decrease DA clearance. Here we show that Gßγ induces the release of DA through DAT. Specifically, a Gßγ-binding/activating peptide, mSIRK, increases DA efflux through DAT in heterologous cells and primary dopaminergic neurons in culture. Addition of the Gßγ inhibitor gallein or DAT inhibitors prevents this effect. Residues 582 to 596 in the DAT carboxy terminus were identified as the primary binding site of Gßγ. A TAT peptide containing the Gßγ-interacting domain of DAT blocked the ability of mSIRK to induce DA efflux, consistent with a direct interaction of Gßγ with the transporter. Finally, activation of a G-protein-coupled receptor, the muscarinic M5R, results in DAT-mediated DA efflux through a Gßγ-dependent mechanism. Collectively, our data show that Gßγ interacts with DAT to promote DA efflux. This novel mechanism may have important implications in the regulation of brain DA homeostasis.

G protein ßγ subunits play a critical role in the actions of amphetamine.

Abnormal levels of dopamine (DA) are thought to contribute to several neurological and psychiatric disorders including drug addiction. Extracellular DA levels are regulated primarily via reuptake by the DA transporter (DAT). Amphetamine, a potent psychostimulant, increases extracellular DA by inducing efflux through DAT. Recently, we discovered that G protein ßγ subunits (Gßγ) interact with DAT, and that in vitro activation of Gßγ promotes DAT-mediated efflux. Here, we investigated the role of Gßγ in the actions of amphetamine in DA neurons in culture, ex vivo nucleus accumbens (NAc), and freely moving rats. Activation of Gßγ with the peptide myr-Ser-Ile-Arg-Lys-Ala-Leu-Asn-Ile-Leu-Gly-Tyr-Pro-Asp-Tyr-Asp (mSIRK) in the NAc potentiated amphetamine-induced hyperlocomotion, but not cocaine-induced hyperlocomotion, and systemic or intra-accumbal administration of the Gßγ inhibitor gallein attenuated amphetamine-induced, but not cocaine-induced hyperlocomotion. Infusion into the NAc of a TAT-fused peptide that targets the Gßγ-binding site on DAT (TAT-DATct1) also attenuated amphetamine-induced but not cocaine-induced hyperlocomotion. In DA neurons in culture, inhibition of Gßγ with gallein or blockade of the Gßγ-DAT interaction with the TAT-DATct1 peptide decreased amphetamine-induced DA efflux. Furthermore, activation of Gßγ with mSIRK potentiated and inhibition of Gßγ with gallein reduced amphetamine-induced increases of extracellular DA in the NAc in vitro and in freely moving rats. Finally, systemic or intra-accumbal inhibition of Gßγ with gallein blocked the development of amphetamine-induced, but not cocaine-induced place preference. Collectively, these results suggest that interaction between Gßγ and DAT plays a critical role in the actions of amphetamine and presents a novel target for modulating the actions of amphetamine in vivo.