Intravital fluorescence microscopy (IVM) is a widely used method to study the microcirculation in several organs. Our aim was to develop a standard rat model to evaluate the microcirculatory characteristics of the urinary bladder under physiological pressure conditions using the most advanced fluorescence videomicroscopic techniques. Spraque-Dawley rats were used after filling their bladders with a constant volume of saline solution. The intravesical pressure was continuously monitored. The bladder was positioned on a specially designed stage and IVM measurements were made at the beginning, the 90th, 120th and 180th min. Arteriolar and venular diameters, functional capillary density, venular red blood cell velocity, arteriolar and venular macromolecular leakage and leukocyte-endothelial cell interactions (observation of rolling and firmly adherent leukocytes) were quantitatively assessed by a computer assisted analysis system. Neither microcirculatory parameters nor the intravesical pressure changed significantly during the observation period of 180 min using constant filling volume. We successfully established a new, well functioning and reproducible model to study the microcirculation of the rat bladder using intravital fluorescent microscopy.