Purpose <p>To quantify bladder wall relaxation times in men without and with lower urinary tract symptoms (LUTS) using three-dimensional (3D) magnetic resonance fingerprinting (MRF).</p> Materials and methods <p>Following informed consent, 51 men (mean age 68.2 years, range 46–80) on active surveillance for prostate cancer underwent multiparametric MRI that included an investigational 3D MRF sequence. Anterior bladder wall T₁ and T₂ relaxation times were measured from the quantitative maps. Anterior bladder wall thickness (BWT) was measured and prostate and transition zone (TZ) volumes were estimated, from the conventional T2w images. Body mass index (BMI), LUTS scores (AUA-SI), risk factors for metabolic syndrome, history of diabetes (DM), and medications prescribed for LUTS were documented from the electronic medical record. The Kruskal-Wallis rank sum test, Wilcoxon rank sum exact test, and Fisher’s exact test were used to assess the associations between clinical variables, categorical imaging data, and mean T₁ and T₂ relaxation times.</p> Results <p>Native bladder wall T₁ relaxation times were longer in men with higher AUA-SI scores [AUA-SI 0–7: 1,496 ms vs. AUA-SI 8–20: 1,787 ms vs. AUA-SI 21–35: 2,063 ms, <i>p</i> = 0.03] while native bladder wall T₂ relaxation times were similar [AUA-SI 0–7: 67 ms vs. AUA-SI 8–20: 78 ms vs. AUA-SI 21–35: 122 ms, <i>p</i> = 0.3]. Native bladder wall T₁ relaxation times were shorter in diabetic men compared to non-diabetic men (1239 ms vs. 1633 ms, <i>p</i> &lt; 0.001), while T₂ relaxation times were similar (86 ms vs. 62 ms, <i>p</i> = 0.4). Prostate and TZ volume, and BWT were similar in men without and with LUTS and DM.</p> Conclusion <p>3D MRF of the bladder is feasible, provides in-vivo tissue characterization of the bladder wall in men with LUTS.</p>