Objective To study the cellular changes of colonic smooth muscle cells in rats with spleen-deficiency functional diarrhea treated with Pixu Sihao Fang (PXSHF, No.4 Formula for Spleen Deficiency) dosed serum.Methods Colonic smooth muscle cells from functional diarrhea spleen-deficiency rats were extracted for primary culture and then divided into the following six groups: normal control group, model group, PXSHF low-dose, medium-dose, and high-dose groups, and montmorillonite powder (Mnt) group.All the dosed serum groups were treated with 5% drug serum for 48 h.CCK-8 test for detecting the cell proliferation activity and QRT-PCR test for checking the expression of brain-gut petide mRNA were performed.Results OD value of the model group was significantly lower than that of the control group (P<0.01) in CCK-8 test while the OD value of the drug serum groups were significantly higher than that of the model group (P<0.01).QRT-PCR test showed that the CCK/VIP/SS mRNA expression of the model group was significant lower than that of the control group (P<0.05).Compared with the model group, the CCK/VIP/SS mRNA expression in each drug serum group presented with an increase tendency.CCK mRNA expression of the group treated with low-dose PXSHF dosed serum was higher than of the model group (P<0.05).VIP mRNA expression of the groups treated with PXSHF dosed serum were significantly higher than that of the model group (P<0.01).Conclusion PXSHF dosed serum can effectively promote the proliferation activity of the colonic smooth muscle cells.Changes in the expression of CCK, VIP and SS mRNA might be indicative of the role of brain-gut peptide in the development of functional diarrhea with spleen deficiency pattern.