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Single Cell Measurement of Dopamine Release with Simultaneous Voltage-clamp and Amperometry
Published on: November 21, 2012
Rab11 supports amphetamine-stimulated norepinephrine transporter trafficking
Heinrich J G Matthies1, Jessica L Moore, Christine Saunders
1Department of Molecular Physiology and Biophysics, Vanderbilt University School of Medicine, Nashville, Tennessee 37232, USA. heinrich.j.matthies@vanderbilt.edu
Summary
Amphetamine (AMPH) causes the norepinephrine transporter (NET) to move into endosomes within neurons. This trafficking is dependent on Rab11, a protein involved in intracellular transport, and impacts NET regulation.
Area of Science:
- Neuroscience
- Molecular Biology
- Pharmacology
Background:
- The norepinephrine transporter (NET) is crucial for regulating norepinephrine levels and is a therapeutic target for various neurological and psychiatric disorders.
- NET function is modulated by intracellular trafficking, but the mechanisms and specific membrane compartments involved in drug-induced trafficking remain unclear.
- Amphetamine (AMPH), a psychostimulant, is known to affect NET, yet its precise impact on NET trafficking at a molecular level has not been visualized.
Purpose of the Study:
- To directly visualize and define the molecular basis of amphetamine (AMPH)-induced norepinephrine transporter (NET) trafficking in neurons.
- To investigate the role of specific intracellular proteins, particularly Rab GTPases, in regulating AMPH-modulated NET trafficking.
- To elucidate the nature of membrane compartments involved in the dynamic redistribution of NET in response to AMPH.
Main Methods:
- Utilized a novel NET antibody for direct visualization of NET trafficking in sympathetic neurons.
- Employed cultured neurons and cortical slices to examine basal and AMPH-modulated NET localization.
- Investigated the interaction and colocalization of NET with Rab11a, Rab4, and Rab11 effectors using tagged proteins and mutant analyses.
Main Results:
- AMPH treatment significantly reduced surface levels of NET in cortical slices.
- AMPH induced the accumulation and colocalization of NET with Rab11a and Rab4 within presynaptic boutons of cultured neurons.
- NET trafficking dynamics were shown to be dependent on functional Rab11, with specific mutants and effectors altering AMPH-induced NET internalization.
Conclusions:
- Amphetamine (AMPH) induces the internalization of the norepinephrine transporter (NET) into endosomal compartments.
- Rab11-dependent intracellular trafficking pathways are critical for mediating AMPH-induced NET redistribution.
- These findings provide a molecular framework for understanding how psychostimulants modulate NET localization and function in neurons.
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