biostudies-literatureWarlow SMBLRD VANIDA NIH HHSNIMH NIH HHSNational Institutes of HealthU.S. Department of Veterans Affairs488-499.e5https://www.ebi.ac.uk/biostudies/studies/S-EPMC10922836biostudies-literatureUnknown112(3)Ventral tegmental area (VTA) projections to the nucleus accumbens (NAc) drive reward-related motivation. Although dopamine neurons are predominant, a substantial glutamatergic projection is also present, and a subset of these co-release both dopamine and glutamate. Optogenetic stimulation of VTA glutamate neurons not only supports self-stimulation but can also induce avoidance behavior, even in the same assay. Here, we parsed the selective contribution of glutamate or dopamine co-release from VTA glutamate neurons to reinforcement and avoidance. We expressed channelrhodopsin-2 (ChR2) in mouse VTA glutamate neurons in combination with CRISPR-Cas9 to disrupt either the gene encoding vesicular glutamate transporter 2 (VGLUT2) or tyrosine hydroxylase (Th). Selective disruption of VGLUT2 abolished optogenetic self-stimulation but left real-time place avoidance intact, whereas CRISPR-Cas9 deletion of Th preserved self-stimulation but abolished place avoidance. Our results demonstrate that glutamate release from VTA glutamate neurons is positively reinforcing but that dopamine release from VTA glutamate neurons can induce avoidance behavior.NeuronMesoaccumbal glutamate neurons drive reward via glutamate release but aversion via dopamine co-release.PMC10922836F32MH122192I01BX005782R01DA036612P30DA048736R01 DA036612K99 MH130688I01 BX005782F32 MH122192I01 BX003759K99MH130688P30 DA048736Dowlat DSSinghal SMFaget LHnasko TSWarlow SMZell VHunker ACZweifel LSHollon NGfalseMesoaccumbal glutamate neurons drive reward via glutamate release but aversion via dopamine co-release.Ventral tegmental area (VTA) projections to the nucleus accumbens (NAc) drive reward-related motivation. Although dopamine neurons are predominant, a substantial glutamatergic projection is also present, and a subset of these co-release both dopamine and glutamate. Optogenetic stimulation of VTA glutamate neurons not only supports self-stimulation but can also induce avoidance behavior, even in the same assay. Here, we parsed the selective contribution of glutamate or dopamine co-release from VTA glutamate neurons to reinforcement and avoidance. We expressed channelrhodopsin-2 (ChR2) in mouse VTA glutamate neurons in combination with CRISPR-Cas9 to disrupt either the gene encoding vesicular glutamate transporter 2 (VGLUT2) or tyrosine hydroxylase (Th). Selective disruption of VGLUT2 abolished optogenetic self-stimulation but left real-time place avoidance intact, whereas CRISPR-Cas9 deletion of Th preserved self-stimulation but abolished place avoidance. Our results demonstrate that glutamate release from VTA glutamate neurons is positively reinforcing but that dopamine release from VTA glutamate neurons can induce avoidance behavior.2024-01-01T00:00:00Z2024 Feb2026-05-29T14:46:31.796Z2025-04-04T00:52:03.115ZS-EPMC109228363808637410.1016/j.neuron.2023.11.002