The efficacy of synaptic transmission depends on the availability of ionotropic and metabotropic neurotransmitter receptors at the plasma membrane, but the contribution of the endocytic and recycling pathways in the regulation of ␥-aminobutyric acid type B (GABA B ) receptors remains controversial. To understand the mechanisms that regulate the abundance of GABA B receptors, we have studied their turnover combining surface biotin labeling and a microscopic immunoendocytosis assay in hippocampal and cortical neurons. We report that internalization of GABA B receptors is agonist-independent. We also demonstrate that receptors endocytose in the cell body and dendrites but not in axons. Additionally, we show that GABA B receptors endocytose as heterodimers via clathrin-and dynamin-1-dependent mechanisms and that they recycle to the plasma membrane after endocytosis. More importantly, we show that glutamate decreases the levels of cell surface receptors in a manner dependent on an intact proteasome pathway. These observations indicate that glutamate and not GABA controls the abundance of surface GABA B receptors in central neurons, consistent with their enrichment at glutamatergic synapses.The efficacy of synaptic transmission depends on the mechanisms of intracellular trafficking, which modify the availability of neurotransmitter receptors (1). At the molecular level, the trafficking of glutamate receptors and ionotropic GABA 5 receptors has been extensively studied (2, 3). For example, differential trafficking of ␣-amino-3-hydroxy-5-methyl-4-isoxazole propionate receptors modifies synaptic strength and influences experience-dependent plasticity in vivo (4). However, the molecular mechanisms that govern the trafficking of metabotropic GABA B receptors (GABA B Rs) remain less clear.GABA B Rs mediate the slow component of synaptic inhibition by acting on pre-and postsynaptic targets (5, 6). They have been implicated in epilepsy, anxiety, stress, sleep disorders, nociception, depression, and cognition (7). They also represent attractive targets for the treatment of withdrawal symptoms from drugs of addiction such as cocaine (8). GABA B Rs are heteromers composed of two subunits, namely GABA B R1 and GABA B R2. GAB-A B R1 contains an endoplasmic reticulum retention motif in the intracellular C-terminal domain (9, 10). The endoplasmic reticulum retention sequence is masked upon assembly with GABA B R2, which results in the appearance of the functional receptor at the plasma membrane. GABA B R1 contains the ligand-binding site, whereas G protein signaling is exclusively mediated by GABA B R2 (11). GABA B Rs are located in dendrites and axons, but it has been difficult to establish what determines their pre-versus postsynaptic localization. Recent evidence suggests that the extracellular domains of the GABA B R1a and -1b isoforms, which differ in two sushi domains (12), may specify axonal/dendritic targeting (13). Unexpectedly, postsynaptic GABA B Rs are enriched at glutamatergic synapses, frequently adjacent to the postsy...

Understanding the mechanisms that control synaptic efficacy through the availability of neurotransmitter receptors depends on uncovering their specific intracellular trafficking routes. ␥-Aminobutyric acid type B (GABA B ) receptors (GABA B Rs) are obligatory heteromers present at dendritic excitatory and inhibitory postsynaptic sites. It is unknown whether synthesis and assembly of GABA B Rs occur in the somatic endoplasmic reticulum (ER) followed by vesicular transport to dendrites or whether somatic synthesis is followed by independent transport of the subunits for assembly and ER export throughout the somatodendritic compartment. To discriminate between these possibilities we studied the association of GABA B R subunits in dendrites of hippocampal neurons combining live fluorescence microscopy, biochemistry, quantitative colocalization, and bimolecular fluorescent complementation. We demonstrate that GABA B R subunits are segregated and differentially mobile in dendritic intracellular compartments and that a high proportion of non-associated intracellular subunits exist in the brain. Assembled heteromers are preferentially located at the plasma membrane, but blockade of ER exit results in their intracellular accumulation in the cell body and dendrites. We propose that GABA B R subunits assemble in the ER and are exported from the ER throughout the neuron prior to insertion at the plasma membrane. Our results are consistent with a bulk flow of segregated subunits through the ER and rule out a post-Golgi vesicular transport of preassembled GABA B Rs.

In this study, we targeted the N-terminal domain (NTD) of transactive response (TAR) DNA binding protein (TDP-43), which is implicated in several neurodegenerative diseases. In silico docking of 50K compounds to the NTD domain of TDP-43 identified a small molecule (nTRD22) that is bound to the N-terminal domain. Interestingly, nTRD22 caused allosteric modulation of the RNA binding domain (RRM) of TDP-43, resulting in decreased binding to RNA in vitro. Moreover, incubation of primary motor neurons with nTRD22 induced a reduction of TDP-43 protein levels, similar to TDP-43 RNA binding-deficient mutants and supporting a disruption of TDP-43 binding to RNA. Finally, nTRD22 mitigated motor impairment in a Drosophila model of amyotrophic lateral sclerosis. Our findings provide an exciting way of allosteric modulation of the RNA-binding region of TDP-43 through the N-terminal domain.

Neurodegenerative diseases represent a formidable challenge to global health. As advances in other areas of medicine grant healthy living into later decades of life, aging diseases such as Alzheimer's disease (AD) and other neurodegenerative disorders can diminish the quality of these additional years, owed largely to the lack of efficacious treatments and the absence of durable cures. Alzheimer's disease prevalence is predicted to more than double in the next 30 years, affecting nearly 15 million Americans, with AD-associated costs exceeding $1 billion by 2050. Delaying onset of AD and other neurodegenerative diseases is critical to improving the quality of life for patients and reducing the burden of disease on caregivers and healthcare systems. Significant progress has been made to model disease pathogenesis and identify points of therapeutic intervention. While some researchers have contributed to our understanding of the proteins and pathways that drive biological dysfunction in disease using in vitro and in vivo models, others have provided mathematical, biophysical, and computational technologies to identify potential therapeutic compounds using in silico modeling. The most exciting phase of the drug discovery process is now: by applying a target-directed approach that leverages the strengths of multiple techniques and validates lead hits using Drosophila as an animal model of disease, we are on the fast-track to identifying novel therapeutics to restore health to those impacted by neurodegenerative disease.