HIV-1 infected macrophages play a significant role in the neuropathogenesis of
AIDS. HIV-1
viral protein R (Vpr) not only facilitates HIV-1
infection but also contribute to long-lived persistence in macrophages. Our previous studies using SILAC-based proteomic analysis showed that the expression of critical metabolic
enzymes in the glycolytic pathway and
tricarboxylic acid (TCA) cycle were altered in response to Vpr expression in macrophages. We hypothesized that Vpr-induced modulation of glycolysis and TCA cycle regulates
glutamate metabolism and release in HIV-1 infected macrophages. We assessed the amount of specific metabolites induced by Vpr and HIV-1 in macrophages at the intracellular and extracellular level in a time-dependent manner utilizing multiple reaction monitoring (MRM) targeted metabolomics. In addition, stable
isotope-labeled
glucose and an MRM targeted metabolomics assay were used to evaluate the de novo synthesis and release of
glutamate in Vpr overexpressing macrophages and HIV-1 infected macrophages, throughout the metabolic flux of glycolytic pathway and TCA cycle activation. The metabolic flux studies demonstrated an increase in
glucose uptake,
glutamate release and accumulation of α-ketoglutarate (α-KG) and
glutamine in the extracellular milieu in Vpr expressing and HIV-1 infected macrophages. Interestingly,
glutamate pools and other intracellular intermediates (
glucose-6-phosphate (G6P),
fructose-6-
phosphate (F6P),
citrate,
malate, α-KG, and
glutamine) showed a decreased trend except for
fumarate, in contrast to the
glutamine accumulation observed in the extracellular space in Vpr overexpressing macrophages. Our studies demonstrate that dysregulation of mitochondrial
glutamate metabolism induced by Vpr in HIV-1 infected macrophages commonly seen, may contribute to neurodegeneration via excitotoxic mechanisms in the context of NeuroAIDS.