Three subtypes of vesicular transporters accumulate glutamate into synaptic vesicles to promote its vesicular release. One of the subtypes, VGLUT3, is expressed in neurons, including cholinergic striatal interneurons, that are known to release other classical transmitters. Here we showed that disruption of the Slc17a8 gene (also known as Vglut3) caused an unexpected hypocholinergic striatal phenotype. Vglut3(-/-) mice were more responsive to cocaine and less prone to haloperidol-induced catalepsy than wild-type littermates, and acetylcholine release was decreased in striatum slices lacking VGLUT3. These phenotypes were associated with a colocalization of VGLUT3 and the vesicular acetylcholine transporter (VAChT) in striatal synaptic vesicles and the loss of a synergistic effect of glutamate on vesicular acetylcholine uptake. We propose that this vesicular synergy between two transmitters is the result of the unbalanced bioenergetics of VAChT, which requires anion co-entry for continuing vesicular filling. Our study reveals a previously unknown effect of glutamate on cholinergic synapses with potential functional and pharmacological implications.

Three different subtypes of Hϩ -dependent carriers (named VGLUT1-3) concentrate glutamate into synaptic vesicles before its exocytotic release. Neurons using other neurotransmitter than glutamate (such as cholinergic striatal interneurons and 5-HT neurons) express VGLUT3. It was recently reported that VGLUT3 increases acetylcholine vesicular filling, thereby, stimulating cholinergic transmission. This new regulatory mechanism is herein designated as vesicular-filling synergy (or vesicular synergy). In the present report, we found that deletion of VGLUT3 increased several anxiety-related behaviors in adult and in newborn mice as early as 8 d after birth. This precocious involvement of a vesicular glutamate transporter in anxiety led us to examine the underlying functional implications of VGLUT3 in 5-HT neurons. On one hand, VGLUT3 deletion caused a significant decrease of 5-HT 1A -mediated neurotransmission in raphe nuclei. On the other hand, VGLUT3 positively modulated 5-HT transmission of a specific subset of 5-HT terminals from the hippocampus and the cerebral cortex. VGLUT3-and VMAT2-positive serotonergic fibers show little or no 5-HT reuptake transporter. These results unravel the existence of a novel subset of 5-HT terminals in limbic areas that might play a crucial role in anxiety-like behaviors. In summary, VGLUT3 accelerates 5-HT transmission at the level of specific 5-HT terminals and can exert an inhibitory control at the raphe level. Furthermore, our results suggest that the loss of VGLUT3 expression leads to anxiety-associated behaviors and should be considered as a potential new target for the treatment of this disorder.

Both acute and chronic dehydration can have important implications for human behaviour and health. Young children, non-autonomous individuals and the elderly are at a greater risk of dehydration. Mild hypertonic dehydration could be related to less efficient cognitive and physical performance and has been reported to be associated with frequently occurring pathological conditions, especially nephrolithiasis. The assessment of hydration status in a large sample appears to be of interest for conducting epidemiological and large clinical studies aimed at improving preventive and curative care. Especially in large-population studies, methods that are used have to be accurate, cheap, quick and require no technical expertise. Body weight change is widely used to determine acute hydration changes, but seems to be insufficiently accurate in longitudinal studies. Bioimpedance analysis methods enable the assessment of total body water content, but their use is still under debate. Because plasma osmolality directly reflects intracellular osmolality, it constitutes a good marker to assess acute hydration changes, but not chronic hydration status because it changes constantly. Moreover, venepuncture is considered to be invasive and is not suitable for a large-sample study, especially in children. Urinary markers appear to be good alternatives for assessing hydration status in large populations. Collection of urine samples is non-invasive and cheap. High technical expertise is not required to perform urinary marker measurements and these measurements can be carried out quickly. Thus, methods based on urinary markers are very well suited for field studies. Urine colour is probably the least sensitive marker despite its high specificity. Urine osmolality and especially urine specific gravity could be easily used for determining hydration status in large-sample studies.