Enzyme-Constrained Metabolic Modeling of the Tripartite Synapse Links Astrocyte-Neuron Lactate Shuttle to Lipid Trafficking and Redox Homeostasis

Neuron-astrocyte metabolic coupling sustains synaptic function through coordinated substrate exchange, redox homeostasis, and lipid handling, yet the systems-level mechanism remains poorly understood. Here, we developed an enzyme-constrained, genome-scale metabolic model of the mouse tripartite synapse to test whether protein allocation can reproduce the astrocyte-neuron lactate shuttle (ANLS). Starting from the iMM1865 mouse metabolic model, we corrected mass and charge imbalances, expanded fatty acid and lipid pathways, and standardized annotations to obtain a high-quality, generic mouse model, improving the MEMOTE score from 46% to 83%. Cell type-specific astrocyte and neuron models were reconstructed using the integrative metabolic analysis tool (iMAT) with cell type-resolved mouse brain proteomics, and were further extended to represent the tripartite synapse. Enzyme-constrained resource allocation models (RAMs) were reconstructed for the astrocyte and neuron models using the GECKO 3.0 pipeline and integrated into a tripartite synapse community model via a modified SteadyCom framework. In the stoichiometry-only model, minimum lactate exchange remained at zero across the entire biomass envelope, indicating no coupling between astrocyte lactate secretion and neuronal uptake arising from the metabolic network itself. Imposing enzyme constraints caused growth-coupled lactate secretion above 90% of maximal growth, with corresponding neuronal lactate uptake, indicating that protein limitation partially drives astrocyte-neuron lactate coupling consistent with ANLS. Incorporation of reactive oxygen species (ROS) and lipid peroxidation pathways demonstrated that the brain-type fatty acid binding protein (FABP7) knockout redistributed flux away from astrocytic detoxification, increased neuronal ROS burden, and produced cell-type-specific suppression of lipid-associated reporter metabolites, including prostaglandin J2, hydroxymethylglutaryl-CoA (HMG-CoA), mevalonate, phosphoinositide/DAG intermediates, fatty-acyl-CoA pools, and ceramide-related sphingolipids. Model predictions, including lactate shuttling and lipid metabolic shifts in FABP7 knockout, align with published experimental studies, supporting the biological relevance of this framework. These findings indicate that ANLS emerges from enzyme limitation in the tripartite synapse model and that FABP7-mediated lipid trafficking is essential for lipid-redox homeostasis at the tripartite synapse. Author SummaryThe brain relies on metabolic cooperation between astrocytes and neurons. Astrocytes convert glucose into lactate, which neurons use for energy. This process is known as the astrocyte-neuron lactate shuttle (ANLS). It is still unclear whether this shuttle is just one possible metabolic pathway or is required due to resource limitations. To explore this, we created two detailed mathematical models of the tripartite synapse, where an astrocyte process extends to meet the synapse. The first model consisted only of the stoichiometry of the metabolic network, whereas the second reflected each cells limited ability to make proteins. When these protein limits were imposed, lactate secretion by astrocytes and uptake by the neurons became mandatory, providing a systems-level mathematics-driven demonstration of ANLS. We also examined the role of Fatty Acid Binding Protein 7 (FABP7), a protein primarily expressed in astrocytes that shuttles fatty acids within astrocyte cells. Modeling FABP7 knockout caused astrocytes to lose their ROS detoxification capacity, increased oxidative stress in neurons, and altered lipid metabolism across cell types. Key affected lipid pathways included prostaglandin signaling, HMG-CoA/ketone body metabolism, cholesterol/mevalonate metabolism, phosphoinositide/DAG signaling, fatty acyl-CoA metabolism, and ceramide-related sphingolipid metabolism. These results show how energy supply, lipid trafficking, and oxidative stress control are linked across cell types and provide a way to identify metabolic vulnerabilities in neurological diseases.