Restoring hippocampal glucose metabolism rescues cognition across Alzheimer disease pathologies

Impaired cerebral glucose metabolism is a pathologic feature of Alzheimer Disease (AD) and recent proteomic studies highlight a disruption of glial carbohydrate metabolism with disease progression.  Here, we report that inhibition of indoleamine-2,3-dioxygenase 1 (IDO1), which metabolizes tryptophan to kynurenine (KYN) in the first step of the kynurenine pathway, rescues hippocampal memory function and plasticity in preclinical models of amyloid and tau pathology by restoring astrocytic metabolic support of neurons.  Activation of IDO1 in astrocytes by amyloid-beta and tau oligomers, two major pathologic effectors in AD, increases KYN and suppresses glycolysis in an AhR-dependent manner. Conversely, pharmacological IDO1 inhibition restores glycolysis and lactate production. In amyloid-producing APPSwe-PS1∆E9 and 5XFAD mice and in tau-producing P301S mice, IDO1 inhibition restores spatial memory and improves hippocampal glucose metabolism by metabolomic and MALDI-MS analyses. IDO1 blockade also rescues hippocampal long-term potentiation in a monocarboxylate transporter (MCT)-dependent manner, suggesting that IDO1 activity disrupts astrocytic metabolic support of neurons. Indeed, in vitro mass-labeling of human astrocytes demonstrates that IDO1 regulates astrocyte generation of lactate that is then taken up by human neurons.  In co-cultures of astrocytes and neurons derived from AD subjects, deficient astrocyte lactate production and transfer to neurons was corrected by IDO1 inhibition, resulting in improved neuronal glucose metabolism.  Thus, IDO1 activity disrupts astrocytic metabolic support of neurons across both amyloid and tau pathologies and in a model of AD iPSC-derived neurons. These findings also suggest that IDO1 inhibitors developed for adjunctive therapy in cancer could be repurposed for treatment of amyloid- and tau-mediated neurodegenerative diseases.

Methods
Metabolites were extracted from isolated astrocytes and neurons in a -80˚C 80:20 methanol:water solution in a volume of 1.5 ml per 1x106 cells, vortexed, incubated on dry ice for 10 min, and centrifuged at 16,000 g for 20 min, and the supernatant was assayed by LC-MS analysis. Extracts were analyzed within 24 hr by liquid chromatography coupled to a mass spectrometer (LC-MS). The LC-MS method involved hydrophilic interaction chromatography (HILIC) coupled to the Q Exactive PLUS mass spectrometer (Thermo Scientific) 7 (17). The LC separation was performed on an XBridge BEH Amide column (150 mm 3 2.1 mm, 2.5 mm particle size, Waters, Milford, MA). Solvent A was 95%: 5% H2O: acetonitrile with 20 mM ammonium bicarbonate, and solvent B was acetonitrile. The gradient was 0 min, 85% B; 2 min, 85% B; 3 min, 80% B; 5 min, 80% B; 6 min, 75% B; 7 min, 75% B; 8 min, 70% B; 9 min, 70% B; 10 min, 50% B; 12 min, 50% B; 13 min, 25% B; 16 min, 25% B; 18 min, 0% B; 23 min, 0% B; 24 min, 85% B; and 30 min, 85% B. Other LC parameters are: flow rate 150 ml/min, column temperature 25°C, and injection volume 10 mL and autosampler temperature was 5°C. The mass spectrometer was operated in both negative and positive ion mode for the detection of metabolites. Other MS parameters were: resolution of 140,000 at m/z 200, automatic gain control (AGC) target at 3e6, maximum injection time of 30 ms and scan range of m/z 75- 1000. Raw LC/MS data were converted to mzXML format using the command line “msconvert” utility (18). Data were obtained with MAVEN software (19, 20). For identification of hexose phosphates and glycolytic intermediates, capillary electrophoresis mass spectroscopy was used as previously described (21). In brief, adherent cells on dishes were washed with 5% mannitol aqueous solution at room temperature. The cells were immersed in 400 μl methanol for 30 s, and 275 μl of the Internal Standard Solution (10 μM, Solution ID: H3304-1002, Human Metabolome Technologies) for 30 s. The extraction liquid was centrifuged at 2,300g for 5 min at 4 °C. The supernatant (400 μl) was centrifugally filtered at 9,100g for 4 h at 4°C through a 5-kDa cut-off filter (Millipore) to remove proteins, and then the filtrate was lyophilized and suspended in 25 μl Milli-Q water. The metabolite suspension was analysed by capillary electrophoresis time of flight mass spectrometry (CE-TOF/MS) using an Agilent capillary electrophoresis (CE) system equipped with an Agilent 6210 TOFMS, an 1100 isocratic high-performance