Characterization of a human point mutation of VGLUT3 (p.A211V) in the rodent brain suggests a non-uniform distribution of the transporter in synaptic vesicles.

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Characterization of a human point mutation of VGLUT3 (p.A211V) in the rodent brain suggests a non-uniform distribution of the transporter in synaptic vesicles.

J Neurosci. 2017 Mar 17;:

Authors: Ramet L, Zimmermann J, Bersot T, Poirel O, De Gois S, Silm K, Yae Sakae D, Mansouri-Guilani N, Bourque MJ, Trudeau LE, Pietrancosta N, Daumas S, Bernard V, Rosenmund C, El Mestikawy S

Abstract
The atypical vesicular glutamate transporter type 3 (VGLUT3) is expressed by sub-populations of neurons using acetylcholine, GABA or serotonin as neurotransmitters. In addition, VGLUT3 is expressed in the inner hair cells of the auditory system. A mutation (p.A211V) in the gene that encodes VGLUT3 is responsible for progressive deafness in two unrelated families. In this study, we investigated the consequences of the p.A211V mutation in cell cultures and in the central nervous system (CNS) of a mutant mouse. The mutation substantially decreased VGLUT3 expression (-70%). We measured VGLUT3-p.A211V activity by vesicular uptake in BON cells, electrophysiological recording of isolated neurons and its ability to stimulate serotonergic accumulation in cortical synaptic vesicles. Despite a marked loss of expression, the activity of the mutated isoform was only minimally altered. Furthermore, mutant mice displayed none of the behavioral alterations that have previously been reported in VGLUT3 knockout mice. Finally, we used stimulated emission depletion microscopy (STED) to analyze how the mutation altered VGLUT3 distribution within the terminals of mice expressing the mutated isoform. The mutation appeared to reduce the expression of the VGLUT3 transporter by simultaneously decreasing the number of VGLUT3-positive synaptic vesicles and the amount of VGLUT3 per synapses. These observations suggested that VGLUT3 global activity is not linearly correlated with VGLUT3 expression. Furthermore, our data unraveled a non-uniform distribution of VGLUT3 in synaptic vesicles. Identifying the mechanisms responsible for this complex vesicular sorting will be critical to understand VGLUT's involvement in normal and pathological conditions.SIGNIFICANCE STATEMENTVGLUT3 is an atypical member of the vesicular glutamate transporter family. A point mutation of VGLUT3 (VGLUT3-p.A211V) responsible for a progressive loss of hearing has been identified in humans. We observed that this mutation dramatically reduces VGLUT3 expression in terminals (approximately 70%) without altering its function. Furthermore, using stimulated emission depletion (STED) microscopy, we found that reducing the expression levels of VGLUT3 diminished the number of VGLUT3-positive vesicles at synapses. These unexpected findings challenge the vision of a uniform distribution of synaptic vesicles at synapses. Therefore, the overall activity of VGLUT3 is not proportional to the level of VGLUT3 expression. These data will be key in interpreting the role of VGLUTs in human pathologies.

PMID: 28314816 [PubMed - as supplied by publisher]

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