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Exercise is one of dementia-s modifiable factors. Exercise could delay neurodegeneration process by increase Klotho, endogen antioxidant and known as anti-aging gene in human. Klotho in hippocampus have certain function such as neuroprotective against oxidative damage induced by β amyloid and glutamate , regulate hippocampal synaptic plasticity, and enhance GluN2B concentration. However, there-s no study about optimal duration of exercise to enhance Klotho and GluN2B expression in hippocampus. This study conducted to compare acute and chronic exercise effect on hippocampal klotho and GluN2B in Rat This study aims to analyse the effect of acute and chronic treadmill exercise on Klotho and GluN2B gene expression in rat-s hippocampus. This study was done using semi-quantitative and animal experimental study. This study used 36 wistar rat from Biofarma were divided into 4 group, 2 control group (acute control and chronic control) and 2 exercise group (Acute and chronic). All group received moderate intensity(20m/min) in 30min/day for 5x/week. After treatment was given, rats were terminated under anaesthesia, hippocampus was extracted from brain, and expression of klotho and GluN2B were studied using PCR and Electrophoresis. Gel band of each gene was visualized using BioDocAnalyze and band thickness quantified using ImageJ. Statistical analysis were done using IBM SPSS version 20.0 software. Data were served as means ± standard error of means. Data were statistically analyzed with Multivariate Analysis of Variance (MANOVA) followed by Post Hoc comparison using Tukey and Dunett. P-Values ˂ 0.05 were considered to be significant.