A chloride conductance in VGLUT1 underlies maximal glutamate loading into synaptic vesicles

Abstract

Uptake of glutamate into synaptic vesicles is mediated by vesicular glutamate transporters (VGLUTs). Although glutamate uptake has been shown to depend critically on Cl(-), the precise contribution of this ion to the transport process is unclear. We found that VGLUT1, and not ClC-3 as proposed previously, represents the major Cl(-) permeation pathway in synaptic vesicles. Using reconstituted VGLUT1, we found that the biphasic dependence of glutamate transport on extravesicular Cl(-) is a result of the permeation of this anion through VGLUT1 itself. Moreover, we observed that high luminal Cl(-) concentrations markedly enhanced loading of glutamate by facilitation of membrane potential-driven uptake and discovered a hitherto unrecognized transport mode of VGLUT1. Because a steep Cl(-) gradient across the synaptic vesicle membrane exists in endocytosed synaptic vesicles, our results imply that the transport velocity and the final glutamate content are highly influenced, if not determined, by the extracellular Cl(-) concentration.

Electrophysiological Signature of Homomeric and Heteromeric Glycine Receptor Channels

“…Because the vesicular membrane potential would be much more positive compared with the plasma membrane potential, ranging between ϩ40 and ϩ80 mV (61), voltage-dependent rectification of GlyR-dependent currents can be particularly relevant with regard to the loading of vesicles with neurotransmitters because it would contribute to increasing ⌬pH. The glycine transporter GlyT1 is also expressed at glutamatergic terminals in the vesicular membrane (62), and vesicle loading with glutamate indeed depends on the vesicular chloride gradient (63). Thus, vesicular GlyRs would be permanently desensitized (due to GlyT1-dependent glycine fill) and restrain chloride-facilitated glutamate loading of presynaptic vesicles, due to voltagedependent rectification.…”

Synaptic and Vesicular Coexistence of VGLUT and VGAT in Selected Excitatory and Inhibitory Synapses

“…due to the increased chloride gradient over the vesicular membrane, which favors VGLUT activity (Schenck et al, 2009). Similarly, VMAT2-mediated serotonin transport was also improved by VGLUT activity.…”

“…It is not clear, however, whether the presence of VGLUT on monoamine-or GABA-storing vesicles necessarily means that glutamate is indeed stored in these vesicles. Interestingly, VGLUT also transports chloride (Schenck et al, 2009). Increasing the vesicular chloride content affects ⌬pH in the same way as glutamate uptake.…”

Vesicular Zinc Regulates the Ca2+Sensitivity of a Subpopulation of Presynaptic Vesicles at Hippocampal Mossy Fiber Terminals

“…A positive correlation has been established between the number of transporter molecules per vesicle and quantal size (Wojcik et al, 2004;Wilson et al, 2005). Cl Ϫ content of endocytosed synaptic vesicles could also be a major determinant of glutamate load (Schenck et al, 2009). ZnT3 and VGLUT1 are cotargeted to the same vesicle population and can reciprocally regulate their transport mechanisms.…”