Neuroinflammation, particularly Toll-like receptor 4 (TLR4)-mediated signaling, plays a critical role in the pathophysiology of depression. This study aimed to investigate the mechanisms by which S-ketamine (S-ket) and S-hydroxynorketamine (S-HNK) modulate TLR4 and peroxisome proliferator-activated receptor alpha (PPARα) signaling in the prefrontal cortex (PFC) to exert antidepressant effects. Male C57BL/6 J mice were subjected to chronic social defeat stress (CSDS) to induce depression-like behaviors. Mice received intraperitoneal administration of S-ket or S-HNK. In a subset of mice, adeno-associated virus expressing TLR4 (AAV-TLR4) or a negative control (NC) was stereotaxically injected into the medial prefrontal cortex (mPFC). Behavioral assessments included the sucrose preference test (SPT), forced swim test (FST), and social interaction test (SIT). Levels of monoaminergic neurotransmitters, including dopamine (DA), serotonin (5-hydroxytryptamine, 5-HT), and norepinephrine (NA), pro-inflammatory cytokines (interleukin (IL)-1β, IL-6, IL-8, and tumor necrosis factor-α (TNF-α)), TLR4, PPARα, indoleamine 2,3-dioxygenase 1 (IDO1), and kynurenine (Kyn) in the PFC were analyzed. CSDS induced depression-like behaviors, decreased DA, 5-HT, and NA levels in the PFC, and increased the expression of pro-inflammatory cytokines IL-1β, IL-6, IL-8, and TNF-α. Treatment with S-ket or S-HNK significantly reversed these behavioral deficits, neurotransmitter alterations, and neuroinflammatory responses. Both compounds also reduced plasma and PFC Kyn levels, upregulated PPARα expression, and downregulated TLR4 and IDO1. Notably, TLR4 overexpression partially abolished the antidepressant and anti-inflammatory effects of S-ket and S-HNK. S-ket and its metabolite S-HNK exert antidepressant effects by modulating the TLR4/PPARα signaling pathway, restoring monoaminergic neurotransmission, and attenuating neuroinflammation.
