The Excitatory Amino Acid Transporter 2 (EAAT2) accounts for 80% of brain glutamate clearance and is mainly expressed in astrocytic perisynaptic processes. EAAT2 function is finely regulated by endocytic events, recycling to the plasma membrane and degradation. Noteworthy, deficits in EAAT2 have been associated with neuronal excitotoxicity and neurodegeneration. In this study, we show that EAAT2 trafficking is impaired by the leucine-rich repeat kinase 2 (LRRK2) pathogenic variant G2019S, a common cause of late-onset familial Parkinson’s disease (PD). In LRRK2 G2019S human brains and experimental animal models, EAAT2 protein levels are significantly decreased, which is associated with elevated gliosis. The decreased expression of the transporter correlates with its reduced functionality in mouse LRRK2 G2019S purified astrocytic terminals and in Xenopus laevis oocytes expressing human LRRK2 G2019S. In LRRK2 G2019S knock-in mouse brain, the correct surface localization of the endogenous transporter is impaired, resulting in its interaction with a plethora of endo-vesicular proteins. Mechanistically, we report that pathogenic LRRK2 kinase activity delays the recycling of the transporter to the plasma membrane via Rabs inactivation, causing its intracellular re-localization and degradation. Taken together, our results demonstrate that pathogenic LRRK2 interferes with the physiology of EAAT2, pointing to extracellular glutamate overload as a possible contributor to neurodegeneration in PD.
The role of betaine in the liver and kidney has been well documented, even from the cellular and molecular point of view. Despite literature reporting positive effects of betaine supplementation in Alzheimer's, Parkinson's and schizophrenia, the role and function of betaine in the brain are little studied and reviewed. Beneficial effects of betaine in neurodegeneration, excitatory and inhibitory imbalance and against oxidative stress in the central nervous system (CNS) have been collected and analysed to understand the main role of betaine in the brain. There are many ‘dark’ aspects needed to complete the picture. The understanding of how this osmolyte is transported across neuron and glial cells is also controversial, as the expression levels and functioning of the known protein capable to transport betaine expressed in the brain, betaine‐GABA transporter 1 (BGT‐1), is itself not well clarified. The reported actions of betaine beyond BGT‐1 related to neuronal degeneration and memory impairment are the focus of this work. With this review, we underline the scarcity of detailed molecular and cellular information about betaine action. Consequently, the requirement of detailed focus on and study of the interaction of this molecule with CNS components to sustain the therapeutic use of betaine.
After 50 years, the heterologous expression of proteins in Xenopus laevis oocytes is still essential in many research fields. New approaches and revised protocols, but also classical methods, such as the two-electrode voltage clamp, are applied in studying membrane transporters. New and old methods for investigating the activity and the expression of Solute Carriers (SLC) are reviewed, and the kinds of experiment that are still useful to perform with this kind of cell are reported. Xenopus laevis oocytes at the full-grown stage have a highly efficient biosynthetic apparatus that correctly targets functional proteins at the defined compartment. This small protein factory can produce, fold, and localize almost any kind of wild-type or recombinant protein; some tricks are required to obtain high expression and to verify the functionality. The methodologies examined here are mainly related to research in the field of membrane transporters. This work is certainly not exhaustive; it has been carried out to be helpful to researchers who want to quickly find suggestions and detailed indications when investigating the functionality and expression of the different members of the solute carrier families.
The role of betaine in the liver and kidney has been well documented, even from the cellular and molecular point of view. Despite literature reporting positive effects of betaine supplementation in Alzheimer’s, Parkinson’s, and Schizophrenia, the role and function of betaine in the brain are little studied and reviewed. Beneficial effects of betaine in neurodegeneration, excitatory and Inhibitory imbalance, and oxidative stress in the central nervous system have been collected and analyzed with the aim of understanding the main role of betaine in the brain. There are many “dark” aspects needed to complete the picture. The understanding of how this osmolyte is transported across neuron and glial cells is also controversial, as the expression levels and functioning of the known protein capable to transport betaine expressed in the brain, betaine-GABA transporter 1 BGT-1, is itself not well clarified. The reported actions of betaine beyond BGT-1 related to neuronal degeneration and memory impairment are the focus of this work. With this review, we underline the scarcity of detailed molecular and cellular information about betaine action. Consequently, the requirement of detailed focus on and study of the interaction of this molecule with CNS components to sustain the therapeutic use of betaine.
The Excitatory Amino Acid Transporter 2 (EAAT2) accounts for 80% of brain glutamate clearance and is mainly expressed in astrocytic perisynaptic processes. EAAT2 function is finely regulated by endocytic events, recycling to the plasma membrane and degradation. Noteworthy, deficits in EAAT2 have been associated with neuronal excitotoxicity and neurodegeneration. In this study, we show that EAAT2 trafficking is impaired by the leucine-rich repeat kinase 2 (LRRK2) pathogenic variant G2019S, a common cause of late-onset familial Parkinson's disease (PD). In LRRK2 G2019S human brains and experimental animal models, EAAT2 protein levels are significantly decreased, which is associated with elevated gliosis. The decreased expression of the transporter correlates with its reduced functionality in mouse LRRK2 G2019S purified astrocytic terminals and in Xenopus laevis oocytes expressing human LRRK2 G2019S. In Lrrk2 G2019S knockin mouse brain, the correct surface localization of the endogenous transporter is impaired, resulting in its interaction with a plethora of endo-vesicular proteins. Mechanistically, we report that pathogenic LRRK2 kinase activity delays the recycling of the transporter to the plasma membrane, causing its intracellular relocalization and degradation. Taken together, our results demonstrate that pathogenic LRRK2 interferes with the physiology of EAAT2, pointing to extracellular glutamate overload as a possible contributor to neurodegeneration in PD.
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