Ketamine is a short-acting dissociative anesthetic and its hallucinogenic side effects have led to increased illicit use among night clubs and party goers. Clinically, ketamine abuse is associated with severe lower urinary tract dysfunction and reduced bladder capacity and hemorrhagic cystitis with irreversible pathological changes which may develop in some cases of long-term drug abuse. Up to now, the mechanisms causing these severe side-effects are still not clear. Herein, a novel ketamine addiction rat model was used to examine the pathological changes and explore the mechanisms of urinary bladders destruction. Rats were divided into groups of control, ketamine injection 14 days and 28 days. Ketamine injection (25 mg/kg/day) was given intraperitoneally while normal saline was given for control group. In vivo isovolumetric cystometrography studies were performed, bladder stiffness parameters were measured, and the bladder tissues were collected for protein analysis and immunohistochemical staining. Ketamine treatment significantly increased micturition pressure but decreased bladder capacity in rats. Ketamine treatment also significantly decreased bladder compliance and increased bladder non-voiding contraction during storage phase. Immunofluorescence studies showing significantly decreased neurofilament staining after ketamine injection 28 days confirmed the neurotoxicity of ketamine. TUNEL staining also showed multiple degenerating cells diffusely distributed in urothelium, suburothelium, and smooth muscle layers in ketamine injected rats. Western blotting demonstrated ketamine injection increased bladder iNOS, eNOS and COX-2 expression. It is concluded that chronic exposure to low, subanesthetic concentrations of ketamine could affect cell survival and impair neuronal morphology which subsequently led to dysfunction of neural networks and altered bladder micturation reflex.