Background: Alzheimer's disease (AD) is a type of age-related disease that affects hippocampus tissue. The AD is a type of amnesiac disorder with dysfunction of the brain in which the patient's mental ability is gradually dissipated. Objectives:The aim of the present study was to investigate the effect of running on positive and negative slopes on serotonin in the hippocampus tissue of rats with AD. Methods: In this experimental study, 18 rats were injected with 8 mg/kg trimethyltin chloride (TMT) intra-peritoneally and after being assured of AD, they were divided into three groups of 6 rats, including (1) control, ( 2) training on positive slope, and (3) training on negative slope. To investigate the effects of AD induction on the serotonin levels, 6 rats were assigned to the healthy control group. Positive training group (at a speed of 16 m/min on positive upward slope) and negative training group (at a speed of 16 m/min on negative downhill slope) ran on the treadmill for eight weeks, five sessions per week and 60 minutes per session. The Shapiro-Wilk, one-way ANOVA with Tukey's post hoc tests were used to analyze the data (P ≤ 0.05). Results:The induction of AD significantly decreased serotonin gene expression levels (P = 0.04); nevertheless, running on positive (P = 0.01) and negative slopes (P = 0.001) significantly increased serotonin gene expression levels in rats with AD. Conclusions: Running on positive and negative slopes seems to improve serotonin gene expression levels in the hippocampus tissue of rats with AD.