Transurethral instillation can be used to deliver different solutions with active ingredients (e.g., drugs, chemicals, bacteria, and viruses) locally into the urinary bladder to either induce animal models of bladder pathologies or evaluate the effectiveness of intravesical treatments.Most rodent models of lower urinary tract (LUT) pathologies are induced in female mice due to ease of intravesical instillation of the substances via the female urethra.However, due to anatomical differences between the female and male LUT, transurethral instillation in a male mouse has been deemed a very challenging procedure, and it has not been previously described.In this manuscript, we provide a detailed description of how to prepare polyethylene (PE) tubing for subsequent insertion into the urethra of a male mouse.In addition, we discuss the ideal types of PE tubing to be used depending on the desired site of inoculation.Furthermore, we describe point by point how to prepare an animal for a successful transurethral instillation to avoid injury to the urethra and ensure the delivery of the solution to the desired location.The procedure is started by retracting the prepuce and the glans to expose the opening of the urethral meatus.Next, the glans are grasped by blunt non-crushing forceps to stabilize the penis and the PE tubing.The PE tubing is first inserted into the urethral meatus parallel to the animal body, then its angle is adjusted by tilting the catheter to maneuver it to follow the natural curvature of the urethra.This technique can be used to induced murine models of bladder pathologies and/or evaluate the effectiveness of intravesical treatments in male mice. Video LinkThe video component of this article can be found at https://www.jove.com/video/56663/ Preparation of TubingNOTE: To instill the inoculum, a needle attached to the polyethylene (PE) tubing (Figure 1A) or a small gauge (<24 G) angiocatheter can be used (Figure 1G).The use of an angiocatheter requires no preparation other than sterilization with 70% ethanol (EtOH).The inoculum is www.jove.com