Glutamate (Glu) is a major excitatory neurotransmitter in the brain, essential for synaptic plasticity, neuronal activity, and memory formation. However, its dysregulation leads to excitotoxicity, implicated in neurodegenerative diseases and brain ischemia. Vesicular glutamate transporters (VGLUTs) regulate Glu loading into synaptic vesicles, crucial for maintaining optimal extracellular Glu levels. This study investigates the neuroprotective effects of VGLUT1 inhibition in HT22 cells overexpressing VGLUT1 under oxygen glucose deprivation (OGD) conditions. HT22 cells, a hippocampal neuron model, were transduced with lentiviral vectors to overexpress VGLUT1. Cells were subjected to OGD, with pre-incubation of Chicago Sky Blue 6B (CSB6B), an unspecific VGLUT inhibitor. Cell viability, lactate dehydrogenase (LDH) release, mitochondrial membrane potential, and hypoxia-related protein markers (PARP1, AIF, NLRP3) were assessed. Results indicated that VGLUT1 overexpression increased vulnerability to OGD, evidenced by higher LDH release and reduced cell viability. CSB6B treatment improved cell viability and reduced LDH release in OGD conditions, particularly at 0.1 μM and 1.0 μM concentrations. Moreover, CSB6B preserved mitochondrial membrane potential and decreased levels of PARP1, AIF, and NLRP3 proteins, suggesting neuroprotective effects through mitigating excitotoxicity. This study demonstrates that VGLUT1 inhibition could be a promising therapeutic strategy for ischemic brain injury, warranting further investigation into selective VGLUT1 inhibitors.

Alzheimer\'s disease (AD) is a progressive neurodegenerative disorder for which there are very limited treatment options. Dysfunction of the excitatory neurotransmitter system is thought to play a major role in the pathogenesis of this condition. Vesicular glutamate transporters (VGLUTs) are key to controlling the quantal release of glutamate. Thus, expressional changes in disease can have implications for aberrant neuronal activity, raising the possibility of a therapeutic target. There is no information regarding the expression of VGLUTs in the human medial temporal lobe in AD, one of the earliest and most severely affected brain regions. This study aimed to quantify and compare the layer-specific expression of VGLUT1 and VGLUT2 between control and AD cases in the hippocampus, subiculum, entorhinal cortex, and superior temporal gyrus. Free-floating fluorescent immunohistochemistry was used to label VGLUT1 and VGLUT2 in the hippocampus, subiculum, entorhinal cortex, and superior temporal gyrus. Sections were imaged using laser-scanning confocal microscopy and transporter densitometric analysis was performed. VGLUT1 density was not significantly different in AD tissue, except lower staining density observed in the dentate gyrus stratum moleculare (p = 0.0051). VGLUT2 expression was not altered in the hippocampus and entorhinal cortex of AD cases but was significantly lower in the subiculum (p = 0.015) and superior temporal gyrus (p = 0.0023). This study indicates a regionally specific vulnerability of VGLUT1 and VGLUT2 expression in the medial temporal lobe and superior temporal gyrus in AD. However, the causes and functional consequences of these disturbances need to be further explored to assess VGLUT1 and VGLUT2 as viable therapeutic targets.

[This corrects the article DOI: 10.3389/fnana.2023.1242839.].

The thalamus is a central link between cortical and subcortical brain motor systems. Axons from the deep nuclei of the cerebellum (DCN), or the output nuclei of the basal ganglia system (substantia nigra reticulata, SNr; and internal pallidum GPi/ENT) monosynaptically innervate the thalamus, prominently some nuclei of the ventral nuclear group. In turn, axons from these ventral nuclei innervate the motor and premotor areas of the cortex, where their input is critical for planning, execution and learning of rapid and precise movements. Mice have in recent years become a widely used model in motor system research. However, information on the distribution of cerebellar and basal ganglia inputs in the rodent thalamus remains poorly defined. Here, we mapped the distribution of inputs from DCN, SNr, and GPi/ENT to the ventral nuclei of the mouse thalamus. Immunolabeling for glutamatergic and GABAergic neurotransmission markers delineated two distinct main territories, characterized each by the presence of large vesicular glutamate transporter type 2 (vGLUT2) puncta or vesicular GABA transporter (vGAT) puncta. Anterograde labeling of axons from DCN revealed that they reach virtually all parts of the ventral nuclei, albeit its axonal varicosities (putative boutons) in the vGAT-rich sector are consistently smaller than those in the vGLUT2-rich sector. In contrast, the SNr axons innervate the whole vGAT-rich sector, but not the vGLUT2-rich sector. The GPi/ENT axons were found to innervate only a small zone of the vGAT-rich sector which is also targeted by the other two input systems. Because inputs fundamentally define thalamic cell functioning, we propose a new delineation of the mouse ventral motor nuclei that is consistent with the distribution of DCN, SNr and GPi/ENT inputs and resembles the general layout of the ventral motor nuclei in primates.

Parkinson\'s disease (PD) is a common neurodegenerative disease implicated in multiple interacting neurotransmitter pathways. Glutamate is the central excitatory neurotransmitter in the brain and plays critical influence in the control of neuronal activity. Impaired Glutamate homeostasis has been shown to be closely associated with PD. Glutamate is synthesized in the cytoplasm and stored in synaptic vesicles by vesicular glutamate transporters (VGLUTs). Following its exocytotic release, Glutamate activates Glutamate receptors (GluRs) and mediates excitatory neurotransmission. While Glutamate is quickly removed by excitatory amino acid transporters (EAATs) to maintain its relatively low extracellular concentration and prevent excitotoxicity. The involvement of GluRs and EAATs in the pathophysiology of PD has been widely studied, but little is known about the role of VGLUTs in the PD. In this review, we highlight the role of VGLUTs in neurotransmitter and synaptic communication, as well as the massive alterations in Glutamate transmission and VGLUTs levels in PD. Among them, adaptive changes in the expression level and function of VGLUTs may exert a crucial role in excitatory damage in PD, and VGLUTs are considered as novel potential therapeutic targets for PD.

Brain ischemia is one of the leading causes of death and long-term disability in the world. Interruption of the blood supply to the brain is a direct stimulus for many pathological events. The massive vesicular release of glutamate (Glu) after ischemia onset induces excitotoxicity, which is a potent stress on neurons. Loading of presynaptic vesicles with Glu is the first step of glutamatergic neurotransmission. Vesicular glutamate transporters 1, 2, and 3 (VGLUT1, 2, and 3) are the main players involved in filling presynaptic vesicles with Glu. VGLUT1 and VGLUT2 are expressed mainly in glutamatergic neurons. Therefore, the possibility of pharmacological modulation to prevent ischemia-related brain damage is attractive. In this study, we aimed to determine the effect of focal cerebral ischemia on the spatiotemporal expression of VGLUT1 and VGLUT2 in rats. Next, we investigated the influence of VGLUT inhibition with Chicago Sky Blue 6B (CSB6B) on Glu release and stroke outcome. The effect of CSB6B pretreatment on infarct volume and neurological deficit was compared with a reference model of ischemic preconditioning. The results of this study indicate that ischemia upregulated the expression of VGLUT1 in the cerebral cortex and in the dorsal striatum 3 days after ischemia onset. The expression of VGLUT2 was elevated in the dorsal striatum and in the cerebral cortex 24 h and 3 days after ischemia, respectively. Microdialysis revealed that pretreatment with CSB6B significantly reduced the extracellular Glu concentration. Altogether, this study shows that inhibition of VGLUTs might be a promising therapeutic strategy for the future.

Glutamate acts as the main excitatory neurotransmitter in the brain and plays a vital role in physiological and pathological neuronal functions. In mammals, glutamate can cause detrimental excitotoxic effects under anoxic conditions. In contrast, Trachemys scripta, a freshwater turtle, is one of the most anoxia-tolerant animals, being able to survive up to months without oxygen. Therefore, turtles have been investigated to assess the molecular mechanisms of neuroprotective strategies used by them in anoxic conditions, such as maintaining low levels of glutamate, increasing adenosine and GABA, upregulating heat shock proteins, and downregulating K ATP channels. These mechanisms of anoxia tolerance of the turtle brain may be applied to finding therapeutics for human glutamatergic neurological disorders such as brain injury or cerebral stroke due to ischemia. Despite the importance of glutamate as a neurotransmitter and of the turtle as an ideal research model, the glutamatergic circuits in the turtle brain remain less described whereas they have been well studied in mammalian and avian brains. In reptiles, particularly in the turtle brain, glutamatergic neurons have been identified by examining the expression of vesicular glutamate transporters (VGLUTs). In certain areas of the brain, some ionotropic glutamate receptors (GluRs) have been immunohistochemically studied, implying that there are glutamatergic target areas. Based on the expression patterns of these glutamate-related molecules and fiber connection data of the turtle brain that is available in the literature, many candidate glutamatergic circuits could be clarified, such as the olfactory circuit, hippocampal-septal pathway, corticostriatal pathway, visual pathway, auditory pathway, and granule cell-Purkinje cell pathway. This review summarizes the probable glutamatergic pathways and the distribution of glutamatergic neurons in the pallium of the turtle brain and compares them with those of avian and mammalian brains. The integrated knowledge of glutamatergic pathways serves as the fundamental basis for further functional studies in the turtle brain, which would provide insights on physiological and pathological mechanisms of glutamate regulation as well as neural circuits in different species.

A high-fat diet with appropriate protein and low carbohydrate content, widely known as the ketogenic diet (KD), is considered as an effective non-pharmacotherapeutic treatment option for certain types of epilepsies. Several preclinical and clinical studies have been carried out to elucidate its mechanism of antiepileptic action. Ketone bodies produced after KD\'s breakdown interact with cellular excito-inhibitory processes and inhibit abnormal neuronal firing. The generated ketone bodies decrease glutamate release by inhibiting the vesicular glutamate transporter 1 and alter the transmembrane potential by hyperpolarization. Apart from their effect on the well-known pathogenic mechanisms of epilepsy, some recent studies have shown the interaction of KD metabolites with novel neuronal targets, particularly adenosine receptors, adenosine triphosphate-sensitive potassium channel, mammalian target of rapamycin, histone deacetylase, hydroxycarboxylic acid receptors, and the NLR family pyrin domain containing 3 inflammasomes to suppress seizures. The role of KD in augmenting gut microbiota as a potential mechanism for epileptic seizure suppression has been established. Furthermore, some recent findings also support the beneficial effect of KD against epilepsy- associated comorbidities. Despite several advantages of the KD in epilepsy management, its use is also associated with a wide range of side effects. Hypoglycemia, excessive ketosis, acidosis, renal stones, cardiomyopathies, and other metabolic disturbances are the primary adverse effects observed with the use of KD. However, in some recent studies, modified KD has been tested with lesser side effects and better tolerability. The present review discusses the molecular mechanism of KD and its role in managing epilepsy and its associated comorbidities.

Glutamate (Glu) is the primary excitatory transmitter in the mammalian brain. But, we know little about the evolutionary history of this adaptation, including the selection of l-glutamate as a signaling molecule in the first place. Here, we used comparative metabolomics and genomic data to reconstruct the genealogy of glutamatergic signaling. The origin of Glu-mediated communications might be traced to primordial nitrogen and carbon metabolic pathways. The versatile chemistry of L-Glu placed this molecule at the crossroad of cellular biochemistry as one of the most abundant metabolites. From there, innovations multiplied. Many stress factors or injuries could increase extracellular glutamate concentration, which led to the development of modular molecular systems for its rapid sensing in bacteria and archaea. More than 20 evolutionarily distinct families of ionotropic glutamate receptors (iGluRs) have been identified in eukaryotes. The domain compositions of iGluRs correlate with the origins of multicellularity in eukaryotes. Although L-Glu was recruited as a neuro-muscular transmitter in the early-branching metazoans, it was predominantly a non-neuronal messenger, with a possibility that glutamatergic synapses evolved more than once. Furthermore, the molecular secretory complexity of glutamatergic synapses in invertebrates (e.g., Aplysia) can exceed their vertebrate counterparts. Comparative genomics also revealed 15+ subfamilies of iGluRs across Metazoa. However, most of this ancestral diversity had been lost in the vertebrate lineage, preserving AMPA, Kainate, Delta, and NMDA receptors. The widespread expansion of glutamate synapses in the cortical areas might be associated with the enhanced metabolic demands of the complex brain and compartmentalization of Glu signaling within modular neuronal ensembles.

Parkinson\'s disease (PD) is a common neurodegenerative disease in middle-aged and elderly individuals. At present, no effective drug has been developed to treat PD. Although a variety of drugs exist for the symptomatic treatment of PD, they all have strong side effects. Most studies on PD mainly focus on dopaminergic neurons. This review highlights the function of glutamic acid transporters (GLTs), including excitatory amino acid transporters (EAATs) and vesicular glutamate transporters (VGLUTs), during the development of PD. In addition, using bioinformatics, we compared the expression of different types of glutamate transporter genes in the cingulate gyrus of PD patients and healthy controls. More importantly, we suggest that the functional roles of glutamate transporters may prove beneficial in the treatment of PD. In summary, VGLUTs and EAATs may be potential targets in the treatment of PD. VGLUTs and EAATs can be used as clinical drug targets to achieve better efficacy. Through this review article, we hope to enable future researchers to improve the condition of PD patients.