Co-packaging of opposing neurotransmitters in individual synaptic vesicles in the central nervous system
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Abstract
Many mammalian neurons release multiple neurotransmitters to activate diverse classes of postsynaptic ionotropic receptors. Entopeduncular nucleus somatostatin (EP Sst+) projection neurons to the lateral habenula (LHb) release both glutamate and GABA, but it is unclear whether these are packaged into the same or segregated pools of synaptic vesicles. Here, we describe a method combining electrophysiology, spatially patterned optogenetics, and computational modeling designed to analyze the mechanism of glutamate/GABA co-release in mouse brain. We find that the properties of postsynaptic currents elicited in LHb neurons by optogenetically activating EP Sst+ terminals are only consistent with co-packaging of glutamate/GABA into individual vesicles. Furthermore, presynaptic neuromodulators that weaken EP Sst+ to LHb synapses maintain the co-packaging of glutamate/GABA while reducing vesicular release probability. Our approach is applicable to the study of multi-transmitter neurons throughout the brain, and our results constrain the mechanisms of neuromodulation and synaptic integration in LHb.
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The Glut-GABA clusters include four clusters in the isocortex and hippocampus and three clusters in globus pallidus, internal segment, which probably correspond to previously well-characterized glutamate-GABA co-releasing neuronal types in these regions.
“They include four clusters in the isocortex and hippocampus and three clusters in globus pallidus, internal segment (GPi), which probably correspond to previously well-characterized glutamate–GABA co-releasing neuronal types in these regions41,42.”
From A high-resolution transcriptomic and spatial atlas of cell types in the whole mouse brain (2023) · cites this paper as [41]
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Experimental — accuracy not yet measuredIn mammals, cases of co-packaging of fast-acting clear core vesicular transmitters are known, including transmitters of opposing downstream effect.
“In mammals, cases of co-packaging of fast-acting clear core vesicular transmitters are known, including transmitters of opposing downstream effect.172”
From Neurotransmitter classification from electron microscopy images at synaptic sites in Drosophila melanogaster (2024) · cites this paper as [172]
These results support a model in which individual synaptic vesicles express both vGAT and vGluT2 and release GABA and glutamate simultaneously to activate postsynaptic receptors across the cleft.
“These results support a model in which individual synaptic vesicles express both vGAT and vGluT2, and GABA/glutamate are released simultaneously to activate postsynaptic receptors across the synaptic cleft.”
From Synaptic and circuit functions of multitransmitter neurons in the mammalian brain (2023) · cites this paper as [25]
Optical quantal analysis found that unitary responses from EP release sites are consistent only with co-packaging of GABA and glutamate into the same synaptic vesicle.
“Optical quantal analysis determined that the statistical properties of unitary responses resulting from activation of cotransmitting EP release sites were consistent only with copackaging of GABA and glutamate into the same synaptic vesicle 25 (Figure 2).”
From Synaptic and circuit functions of multitransmitter neurons in the mammalian brain (2023) · cites this paper as [25]
The structure, organization, and regulation of postsynaptic receptors and related molecular constituents of GABA/glutamate synapses remain unknown but appear specialized within LHb due to low PSD-95 expression.
“The structure, organization, and regulation of the postsynaptic receptors and related molecular constituents of GABA/glutamate synapses remain unknown but appear specialized within LHb due to low expression of the glutamatergic postsynaptic-terminal-associated MAGUK PSD-95.25”
From Synaptic and circuit functions of multitransmitter neurons in the mammalian brain (2023) · cites this paper as [25]
Individual synaptic vesicles from EP Sst neurons in the LHb contain both GABA and glutamate.
“individual synaptic vesicles from EP Sst neurons in the LHb contain both GABA and glutamate”
From Synaptic and circuit functions of multitransmitter neurons in the mammalian brain (2023) · cites this paper as [25]
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A high-resolution transcriptomic and spatial atlas of cell types in the whole mouse brain
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- Authors
- Kim, S., Wallace, M. L., El-Rifai, M., Knudsen, A. R., & Sabatini, B. L.
- Affiliations
- Howard Hughes Medical Institute (US)
- Harvard University (US)
- Corresponding author
- Sabatini, Bernardo L.
- Venue
- Neuron
- Publisher
- Elsevier BV
- Published
- 2022-04
- Volume
- 110
- Issue
- 8
- Pages
- 1371-1384.e7
- ISSN
- 0896-6273 (ISSN-L)
- PMC
- PMC9056948
- OpenAlex
- W4210791415
- Cited by
- 58
- References
- 47
- Citations by year
- 2026: 11 · 2025: 11 · 2024: 19 · 2023: 10 · 2022: 4 · 2021: 2 · 2020: 1
- FWCI
- 4.86
- Type
- journal article
- Language
- en
- Open access
- bronze
- doi.org
- Licence
- www.elsevier.com/tdm/userlicense/1.0/ (tdm)
- www.elsevier.com/legal/tdmrep-license (tdm)
- www.elsevier.com/open-access/userlicense/1.0/ (vor, 384 d)
- doi.org/10.15223/policy-017 (stm-asf)
- doi.org/10.15223/policy-037 (stm-asf)
- doi.org/10.15223/policy-012 (stm-asf)
- doi.org/10.15223/policy-029 (stm-asf)
- doi.org/10.15223/policy-004 (stm-asf)
- Locations
- Neuron (publishedVersion)
- bioRxiv (Cold Spring Harbor Laboratory) (acceptedVersion)
- PubMed (publishedVersion)
- PubMed Central (acceptedVersion)
- Funders
- National Institutes of Health (P30NS072030, R00NS105883-03, R01NS103226)
- Topics
- Life Sciences › Neuroscience › Cellular and Molecular Neuroscience › Neuroscience and Neuropharmacology Research
- Life Sciences › Biochemistry, Genetics and Molecular Biology › Cell Biology › Cellular transport and secretion
- Life Sciences › Neuroscience › Cellular and Molecular Neuroscience › Neurotransmitter Receptor Influence on Behavior
- Keywords
- entopeduncular nucleus, lateral habenula, release probability, postsynaptic currents, synaptic integration, optogenetic stimulation
- MeSH terms
- Animals, gamma-Aminobutyric Acid, Mammals, Synaptic Vesicles, Neurotransmitter Agents, Glutamic Acid, Habenula, Mice
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