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Although loss-of-function mutations in the PARK2 gene, the gene that encodes the protein parkin, cause autosomal recessive juvenile parkinsonism, the responsible molecular mechanisms remain unclear. Evidence suggests that a loss of parkin dysregulates excitatory synapses. Here we show that parkin interacts with the kainate receptor (KAR) GluK2 subunit and regulates KAR function. Loss of parkin function in primary cultured neurons causes GluK2 protein to accumulate in the plasma membrane, potentiates KAR currents and increases KAR-dependent excitotoxicity. Expression in the mouse brain of a parkin mutant causing autosomal recessive juvenile parkinsonism results in GluK2 protein accumulation and excitotoxicity. These findings show that parkin regulates KAR function in vitro and in vivo, and suggest that KAR upregulation may have a pathogenetic role in parkin-related autosomal recessive juvenile parkinsonism.

The solute carrier (SLC) superfamily represents the biggest family of transporters with important roles in health and disease. Despite being attractive and druggable targets, the majority of SLCs remains understudied. One major hurdle in research on SLCs is the lack of tools, such as cell-based assays to investigate their biological role and for drug discovery. Another challenge is the disperse and anecdotal information on assay strategies that are suitable for SLCs. This review provides a comprehensive overview of state-of-the-art cellular assay technologies for SLC research and discusses relevant SLC characteristics enabling the choice of an optimal assay technology. The Innovative Medicines Initiative consortium RESOLUTE intends to accelerate research on SLCs by providing the scientific community with high-quality reagents, assay technologies and data sets, and to ultimately unlock SLCs for drug discovery.

These authors equally contributed to this work Key WordsCigarette smoke • Oxidative stress • Glutathione • Carboxymethylcysteine • Airway epithelial cells Abstract Background: Cigarette smoke extract (CSE), a model for studying the effects of tobacco smoke in vivo and in vitro, induces cell oxidative stress and affects the antioxidative glutathione system. We evaluated the impact of CSE on airway epithelial cells and the possible cytoprotective effect of the mucolitic drug S-carboximethilcysteine lysine salt (S-CMCLys). Methods: Reduced glutathione (GSH) and reactive oxygen species (ROS) intracellular levels were evaluated by fluorimetry in human bronchial epithelial cells (16-HBE) and the expression and activity of enzymes of the GSH metabolic pathway were investigated by RT-PCR, Western blot and colorimetric assays. Results: CSE significantly increased cell mortality in a time and dose dependent manner, via an apoptosis-independent pathway. Short-term (3 hours) CSE exposure induced an increase in ROS levels and a GSH intracellular concentration drop. In parallel, the expression of glutathione peroxidases 2 and 3, glutathione reductase and glutamate-cysteine-ligase was increased. S-CMC-Lys was effective in counteracting these effects. Conclusion: CSE affects ROS levels, GSH concentration and GSH enzymes pathway. These effects can be to some extent reversed by S-CMC-Lys, that could represent a therapeutic tool to counteract CSE induced oxidative cellular injuries.