Objective To investigate the change in the fibrosis degree of cultured rat arachnoid cells in vitro after being stimulated by thrombin, and to explore its relationship with the development of chronic hydrocephalus after intraventricular hemorrhage (IVH) in rats.Methods We used different concentrations of thrombin to stimulate the arachnoid cells of Sprague-Dawley (SD) rats, simulating in vitro the pathophysiological process of fibrosis of arachnoid cells in rats with chronic hydrocephalus after IVH. Immunocytochemistry was used to identify the expression of the markers of subcultured SD rat arachnoid cells, i.e., cytokeratin (CK 8/18) and desmoplakin. The mRNA and protein expression of fibrosis factors (Col-I, Col-III, TGF-β1, and α-SMA) in arachnoid cells was measured by quantitative real-time PCR and Western blot, respectively. Those factors were measured for indicating fibrosis degree, and they were compared between different groups.Results (1) Cultured arachnoid cells showed positive expression of the markers cytokeratin and desmoplakin. (2) Thrombin induced fibrosis of arachnoid cells, with the most significant fibrosis degree at 50 u/ml. (3) The experimental group had a higher fibrosis degree than the control group.Conclusions Rat arachnoid cells can be stably cultured in vitro. Thrombin-stimulated arachnoid cells have significantly higher levels of various fibrosis indicators than the control cells, suggesting the successful establishment of fibrosis model. This arachnoid cell fibrosis model can help to further investigate the mechanism of chronic hydrocephalus after IVH.