Annotated protein:	Excitatory amino acid transporter 2 (GLT-1) (Sodium-dependent glutamate/aspartate transporter 2) (Solute carrier family 1 member 2). Gene symbol: SLC1A2. Taxonomy: Mus musculus (Mouse). Uniprot ID: P43006
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SynGO gene info:	SynGO data @ SLC1A2
Ontology domain:	Biological Process
SynGO term:	neurotransmitter reuptake (GO:0098810)
Synapse type(s):	forebrain, glutamatergic
Annotated paper:	Petr GT, et al. "Conditional deletion of the glutamate transporter GLT-1 reveals that astrocytic GLT-1 protects against fatal epilepsy while neuronal GLT-1 contributes significantly to glutamate uptake into synaptosomes" J Neurosci. 2015 Apr 1;35(13):5187-201 PMID:25834045
Figure(s):	Fig 5B-F.
Annotation description:	Fig 5B-F, Synaptosomal uptake of L-[ 3H]glutamate was significantly decreased in neuronal GLT-1 knock-out but not in astrocytic GLT-1 knock-out. Literal: "The uptake in GFAP/cre- mice (n = 4) was decreased to 25.6 ± 2.2% of control by application of 300 μm DHK. In contrast, neuronal deletion of GLT-1 produced a major loss in synaptosomal uptake of glutamate, which was reduced to 44.3 ± 3.4% (p < 0.05, n = 5) of the controls in these experiments using a single concentration (10 μm) of glutamate (Fig. 5B). Pharmacological inhibition by DHK (300 μm) resulted in decrease in syn/cre- (reduced to 26.9 ± 3.3% of controls, n = 5) and syn/cre+ (reduced to 17.2 ± 2.8% of controls, n = 5) mice. The uptake activities in all genotypes were greatly inhibited, to 15.0 ± 2.8% (GFAP/cre-), 9.0 ± 1.1% (GFAP/cre+), 14.7 ± 3.4% (syn/cre-), and 11.7 ± 1.8% (syn/cre+) of control, respectively, by 30 μm TBOA, which is a broad-spectrum competitive high affinity glutamate transporter antagonist (Fig. 5A,B). Increasing the concentration of TBOA to 300 μm almost completely abolished the glutamate uptake in all the genotypes (reduced to 4.5 ± 1.3%, 3.5 ± 1.2%, 2.8 ± 1.3% and 2.7 ± 0.8% of control in GFAP/cre-, GFAP/cre+, syn/cre-, and syn/cre+ mice, respectively; Fig. 5A,B). Given prior genetic evidence that synaptosomal glutamate uptake is almost exclusively mediated by GLT-1 (Tanaka et al., 1997), the discrepancy between the effectiveness of DHK and TBOA in inhibiting glutamate uptake in synaptosomes from the conditional knock-outs and control animals seemed likely to be due to the relatively low affinity of DHK (Robinson et al., 1993; Chen et al., 2002; Kabakov and Rosenberg, 2009), and the fact that in these assays we used 10 μm glutamate. Indeed, we found that, at a glutamate concentration of 0.5 μm, 300 μm DHK abolished ~85% of the glutamate uptake in crude synaptosomes isolated from syn/cre+ mice and their controls (data not shown). We also performed saturation assay of glutamate uptake into crude synaptosomes from the mutant mice and their littermate controls to determine maximal velocity (Vmax) and glutamate binding affinity (Km) values for glutamate transport (Fig. 5C,D). Vmax values determined using GFAP/cre+ brain synaptosomes were, in two experiments, 11.1 (Fig. 5C) and 19.2 pmol/μg/min, similar to values obtained using GFAP/cre- brain synaptosomes [11.7 (Fig. 5C) and 18.3 pmol/μg/min]. Km values were 9.7 (Fig. 5C) and 10.2 μm for the GFAP/cre- mice and 12.4 (Fig. 5C) and 13.0 μm for the GFAP/cre+ mice. The mean Vmax value of syn/cre+ synaptosomes in four experiments was 7.2 ± 0.9 pmol/μg/min (n = 4), significantly lower (40.5%) than that of syn/cre- synaptosomes (12.1 ± 1.8 pmol/μg/min, n = 4, p < 0.05). There was no difference in Km values between the syn/cre- (9.7 ± 0.6 μm, n = 4) and syn/cre+ mice (11.8 ± 0.9 μm, n = 4, p > 0.05). Pooled data from the four experiments that were performed are shown in Figure 5D. Similar results were observed when d-[3H]aspartate was used instead of l-[3H]glutamate for the measurement of synaptosomal uptake in astrocytic or neuronal GLT-1 knock-out mice (Fig. 5E,F). No significant reduction was found in the d-aspartate uptake in the astrocytic GLT-1 knock-out mice (reduced to 87.5 ± 14.5% of control, n = 3, p = 0.49; Fig. 5E). Neuronal deletion of GLT-1 caused a significant decrease of d-aspartate uptake (reduced to 51.3 ± 5.4% of control, n = 3, p < 0.05; Fig. 5F)."
Evidence tracking, Biological System:	Intact tissue
Evidence tracking, Protein Targeting:	Genetic transformation (eg; knockout, knockin, mutations)
Evidence tracking, Experiment Assay:	Transmembrane transport assay
Annotator(s):	Chiara Verpelli (ORCID:0000-0003-2949-9725) Carlo Sala (ORCID:0000-0003-0662-9523)
Lab:	CNR Neuroscience Institute Milan and Dept. of Biotechnology and Translational Medicine, University of Milan, 20129 Milan, Italy
SynGO annotation ID:	1820
Dataset release (version):	20231201
View annotation as GO-CAM model: