Co-release of glutamate and GABA from single vesicles in GABAergic neurons exogenously expressing VGLUT3.

Johannes Zimmermann,Christian Rosenmund,Melissa A Herman

doi:10.3389/fnsyn.2015.00016

Johannes Zimmermann, Christian Rosenmund + Show 1 more

Open Access

https://doi.org/10.3389/fnsyn.2015.00016

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Abstract

The identity of the vesicle neurotransmitter transporter expressed by a neuron largely corresponds with the primary neurotransmitter that cell releases. However, the vesicular glutamate transporter subtype 3 (VGLUT3) is mainly expressed in non-glutamatergic neurons, including cholinergic, serotonergic, or GABAergic neurons. Though a functional role for glutamate release from these non-glutamatergic neurons has been demonstrated, the interplay between VGLUT3 and the neuron’s characteristic neurotransmitter transporter, particularly in the case of GABAergic neurons, at the synaptic and vesicular level is less clear. In this study, we explore how exogenous expression of VGLUT3 in striatal GABAergic neurons affects the packaging and release of glutamate and GABA in synaptic vesicles (SVs). We found that VGLUT3 expression in isolated, autaptic GABAergic neurons leads to action potential evoked release of glutamate. Under these conditions, glutamate and GABA could be packaged together in single vesicles release either spontaneously or asynchronously. However, the presence of glutamate in GABAergic vesicles did not affect uptake of GABA itself, suggesting a lack of synergy in vesicle filling for these transmitters. Finally, we found postsynaptic detection of glutamate released from GABAergic terminals difficult when bona fide glutamatergic synapses were present, suggesting that co-released glutamate cannot induce postsynaptic glutamate receptor clustering.

Highlights

In synaptic transmission, postsynaptic responses are determined in large part by the identity of the neurotransmitter released at the presynaptic terminal, which is, in turn, determined by the type of vesicular neurotransmitter transporter expressed by the presynaptic neuron
Addition of the GABA receptor antagonist Bic to the extracellular solution (ECS) revealed a fast excitatory component (Figures 1A,C) that could be blocked by the AMPA receptor antagonist NBQX, which suggests that glutamate could be released from these GABAergic terminals
To analyze the decay kinetics of quantal events for the co-release experiments we used two different methods, both resulting in a similar outcome: (1) After running the recognition template in AxoGraph the decays of detected miniature PSC (mPSC) were fitted in AxoGraph with a single or double exponential using a simplex optimization procedure from the peak until the decay reached baseline

Summary

Introduction

Postsynaptic responses are determined in large part by the identity of the neurotransmitter released at the presynaptic terminal, which is, in turn, determined by the type of vesicular neurotransmitter transporter expressed by the presynaptic neuron. In cholinergic and serotonergic terminals, VGLUT3 stimulates the vesicular uptake of acetylcholine (Ach; Gras et al, 2008; Nelson et al, 2014) and serotonin (5-HT; Amilhon et al, 2010), respectively, to synaptic vesicles (SVs). This is likely because the co-packaging of glutamate, a negatively charged molecule, causes an increase in the pH gradient across the Glutamate/GABA co-release from single vesicles vesicular membrane, which synergistically increases the driving force for uptake of the other neurotransmitters (Hnasko et al, 2010; El Mestikawy et al, 2011). Transport assays suggest evidence for synergistic effects for the uptake of GABA by glutamate (Zander et al, 2010), but these effects have not been investigated in SVs release from individual terminals

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