Per and polyfluoroalkyl substances (PFAS) are synthetic compounds extensively utilized in industrial applications and consumer products. Long-chain PFAS has been linked to negative health impacts, prompting the introduction of shorter-chain alternatives like perfluorobutane sulfonate (PFBS). While long-chain PFAS are known to induce oxidative stress, neuroinflammation, and neuronal apoptosis, the neurotoxic potential of short-chain PFAS like PFBS was not well studied. This study aims to evaluate the neurotoxic effect and bioaccumulation of PFBS on adult zebrafish. In this study, adult zebrafish were exposed to PFBS at concentrations of 0.14, 1.4, and 14 μM for 28 days. PFBS accumulation in zebrafish brain tissue was confirmed by specific mass spectrum peaks. Behavioral assays revealed significant anxiety-like behavior, with PFBS (14 μM) exposed zebrafish spending more time in the bottom zone of the novel tank diving test (179.33 ± 1.03 s) and in the light and dark preference results showed increased time spent in the dark zone (165.17 ± 10.89 s). Learning and memory deficits were evident in the T-maze test, where PFBS-exposed zebrafish spent less time in the favorable zone (0.67 ± 1.15 s). Biochemical analysis showed significant inhibition of acetylcholinesterase (AChE) activity in the male and female brains (0.06 μmol/min and 0.09 μmol/min). Antioxidant enzyme levels were reduced, with superoxide dismutase (SOD) 5.45 U/mg protein in the male brain and 4.06 U/mg protein in the female brain, leading to increased oxidative stress biomarkers like lipid peroxidation and nitric oxide levels in male (0.99 μmol/mg/ml and 8.85 μM) and female brain (1.83 μmol/mg/ml and 8.74 μM), respectively. Gene expression analysis demonstrated the downregulation of SOD, CAT, GSR, and GPx, indicating impaired antioxidant defense mechanisms. Histopathological analysis of PFBS exposure groups revealed vacuolation and increased pyknotic neurons in the optic tectum region of the brain. Our study suggests that PFBS exposure leads to bioaccumulation in the brain, causing histopathological changes and cognitive impairment. In conclusion, PFBS induces neurotoxicity which can be a potential risk as they are incorporated into a range of consumer products.