Rab11 supports amphetamine-stimulated norepinephrine transporter trafficking.

Matthies HJ, Moore JL, Saunders C, Matthies DS, Lapierre LA, Goldenring JR, Blakely RD, Galli A
J Neurosci. 2010 30 (23): 7863-77

PMID: 20534835 · PMCID: PMC2935280 · DOI:10.1523/JNEUROSCI.4574-09.2010

The norepinephrine transporter (NET) is a presynaptic plasma membrane protein that mediates reuptake of synaptically released norepinephrine. NET is also a major target for medications used for the treatment of depression, attention deficit/hyperactivity disorder, narcolepsy, and obesity. NET is regulated by numerous mechanisms, including catalytic activation and membrane trafficking. Amphetamine (AMPH), a psychostimulant and NET substrate, has also been shown to induce NET trafficking. However, neither the molecular basis nor the nature of the relevant membrane compartments of AMPH-modulated NET trafficking has been defined. Indeed, direct visualization of drug-modulated NET trafficking in neurons has yet to be demonstrated. In this study, we used a recently developed NET antibody and the presence of large presynaptic boutons in sympathetic neurons to examine basal and AMPH-modulated NET trafficking. Specifically, we establish a role for Rab11 in AMPH-induced NET trafficking. First, we found that, in cortical slices, AMPH induces a reduction in surface NET. Next, we observed AMPH-induced accumulation and colocalization of NET with Rab11a and Rab4 in presynaptic boutons of cultured neurons. Using tagged proteins, we demonstrated that NET and a truncated Rab11 effector (FIP2DeltaC2) do not redistribute in synchrony, whereas NET and wild-type Rab11a do. Analysis of various Rab11a/b mutants further demonstrates that Rab11 regulates NET trafficking. Expression of the truncated Rab11a effector (FIP2DeltaC2) attenuates endogenous Rab11 function and prevented AMPH-induced NET internalization as does GDP-locked Rab4 S22N. Our data demonstrate that AMPH leads to an increase of NET in endosomes of single boutons and varicosities in a Rab11-dependent manner.

MeSH Terms (21)

Adrenergic Agents Amphetamine Animals Biotinylation Carrier Proteins Cells, Cultured Fluorescent Antibody Technique Ganglia, Sympathetic Membrane Proteins Mice Mice, Inbred C57BL Microscopy, Confocal Mutation Neurons Norepinephrine Plasma Membrane Transport Proteins Protein Transport rab4 GTP-Binding Proteins rab GTP-Binding Proteins Rats Signal Transduction Synaptic Vesicles

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