Immunoelectron Microscopy of Trace Amine‐Associate Receptor 1 Intracellular Localization

Abstract

Trace amine‐associated receptor 1 (TAAR1) is a G protein‐coupled receptor (GPCR) in brain regions that are critical to reward and habit formation. TAAR1 is activated by a variety of substituted phenethylamines (e.g. methamphetamine, MA), and is uniquely involved in the mechanism of action of amphetamine‐like psychostimulants. Pharmacological manipulation of TAAR1 alters the spontaneous firing rate of neurons in the ventral tegmental area, and psychiatric disorders characterized by dysregulated dopaminergic neurotransmission, including schizophrenia and substance use disorders, have been associated with altered trace amine levels or TAAR1 polymorphisms.Unlike many GPCRs, TAAR1 is an intracellular receptor; rather than localizing to the plasma membrane, TAAR1 is associated with unidentified membranous compartments within the cell. Previous efforts to identify the intracellular localization of TAAR1 have had limited success due to difficulty with heterologous expression systems and a lack of sufficiently specific antibodies, as well as inherent spatial resolution limits of light‐based microscopic techniques. In order to determine the precise subcellular localization of TAAR1, immunoelectron microscopy was performed in HEK293T cells stably expressing human TAAR1 with a C‐terminal EGFP tag. Preliminary results suggest that TAAR1 localizes to the endoplasmic reticulum and mitochondria. Future experiments using similar techniques will explore TAAR1 localization in primary neurons. Together, these results will further elucidate the molecular mechanisms by which TAAR1 signals within neurons and ultimately alters neurotransmission.