Major depressive disorder (MDD) and anxiety are linked to brain deficits in the bioenergetic molecules creatine (CR) and acetyl-L carnitine (ALC), nutritional supplements which show potential as antidepressants. MDD and anxiety are often comorbid, and affect 20% of US adults. Moreover, cognitive dysfunction contributes markedly to MDD-linked functional disability. Classical antidepressants such as selective serotonin reuptake inhibitors (SSRIs) are ineffective in ~50% of patients, and mostly do not improve cognitive symptoms. Unlike classical antidepressants, bioenergetic compounds show promise for improving multiple aspects of MDD symptomology, including cognitive function and comorbid anxiety. We now plan to explore the bioenergetic compounds CR, cyclocreatine (CyCR, a CR analog) and ALC as therapeutics in a model for treatment resistant depression (TRD). We established a sex-based animal model to study etiology of the high rates of MDD and anxiety in people at altitude (hypobaric hypoxia). At moderate altitude (4500ft), rats of both sexes exhibit symptoms of depression, anxiety and cognitive dysfunction vs. those at sea level, and a sustained lack of response to most SSRIs. Hypo- baric hypoxia induces brain bioenergetic deficits, and CR, CyCR and ALC can reduce the deficit to show sex- based antidepressant-like or anxiolytic-like efficacy in this model. Brain bioenergetic deficits are directly linked to MDD, but can further promote onset of MDD by causing inflammation and oxidative stress. Hypoxia-inducible transcription factors (HIFs) interact with the oxidative stress system to mediate cellular response to hypoxia, and CR can alter HIF activation. Our central hypothesis is that bioenergetic compounds will improve status of de- pression, anxiety and cognitive function by reducing inflammation and oxidative stress at altitude. In Aim 1, we will identify impact of CR, CyCR and ALC vs. SSRI on behavioral symptoms. Efficacy in reducing symptoms of depression (in forced swim, sucrose preference tests), anxiety (e.g., marble burying, elevated T-maze tests) and cognitive dysfunction (e.g., novelty object, radial arm tests) will be tested. In Aim 2, we will study the mode of action by which bioenergetic compounds reduce MDD-linked biomarkers. The impact of CR, CyCR and ALC on biomarkers of inflammation (e.g., the nonspecific inflammatory marker CRF, inflammatory cytokines TNFα, IL6, IL1β), HIF1a activation and oxidative stress (e.g., reactive oxygen species, antioxidants) will be assessed. In Aim 3, we will test molecular mechanisms via which bioenergetic compounds protect against TRD. In our model, bioenergetic drugs may protect against TRD by activating HIFs. CR and ALC can also activate mTOR, PI3K or NfκB pathways. We will now use inhibitors of the signaling pathways likely involved (HIF1a, mTOR, PI3K, NfκB) to block behavioral and biomarker impacts of bioenergetic drugs. Data from studies in this model will further be confirmed using the chronic ...