Gut dysbiosis, characterized by an imbalance in the composition and function of microbiota, has emerged as a critical factor in the regulation of pain transmission and behavior through the gut–brain axis. When dysbiosis occurs, these regulatory functions are disrupted, leading to systemic inflammation and altered signaling at central nervous system. In this study, we examined the therapeutic potential of palmitoyl glucosamine (PGA), a fatty acid amide with sharp anti-inflammatory properties, in a gut dysbiosis condition induced by antibiotic exposure in male mice. At the intestine level, PGA treatment reduced proinflammatory biomarkers and affects gut microbiota balance. Moreover, it reduced visceral hypersensitivity and pain transmission to the spinal cord by reducing the expression of mechanosensitive ion channel Piezo-2. Finally, PGA counteracted the affective comorbidities of dysbiosis, including depressive-like behavior and impaired cognition. This study sheds light on the function of gut microbiota in the control of pain pathways and provides new insights into the advantages of PGA treatment in managing pathological conditions linked to microbiota-related immune and sensory dysregulation.