Stable rat models in constipation with depression and anxiety: evaluation and validation

Purpose The study aimed to establish a model of functional constipation (FC) with anxiety and depression in rats using the compound diphenoxylate combined with chronic unpredictable mild stress (CUMS). It further investigated constipation and behavioral indicators in different groups of various compounds of diphenoxylate with CUMS durations to determine the optimal time frame for successful model establishment. Method Thirty-two female rats were randomly assigned to control, FC before CUMS, FC after CUMS, and FC throughout CUMS groups. Control: Same amount of regular saline solution, FC before CUMS: Compound diphenoxylate 15 mg/kg 2 weeks + CUMS 5 weeks, FC after CUMS: CUMS 5 weeks + Compound diphenoxylate 15 mg/kg 2 weeks, FC throughout CUMS: CUMS 5 weeks with compound diphenoxylate 10 mg/kg for 4 weeks. After model establishment, constipation and behavioral indicators were detected. HE staining was used to observe pathological changes in the brain and colon tissue, while AB-PAS was used to observe goblet cells in the colon tissue, and Nissl staining was used to observe neuron cells in the hippocampus. The protein expression of BDNF and Bax in the hippocampus and PGP9.5 and iNOS in the colon was detected by WB. Results In the FC throughout group, synaptic ultrastructure significantly decreased, aligning with pathological changes, indicators of FC with anxiety and depression. The protein expression levels of BDNF and PGP9.5 were markedly decreased, and the levels of Bax and iNOS were elevated. This study provided a reliable reference for determining the optimal establishment way of FC with anxiety and depression models and for in vivo FC with anxiety and depression animal experiments.