Objectives: To investigate the effect of different laser power settings on intra-renal temperature (IRT) under different irrigation conditions during flexible ureteroscopy (FURS) in a live-anesthetized porcine model. Methods: Following ethics approval, 2 female pigs weighing ~ 28 kg were used. Under general anesthesia, a percutaneous access was obtained to fix a K-type thermocouple inside the pelvi-calyceal system for real-time recording of IRT during FURS without UAS, UAS-10/12, UAS-12/14, and UAS-14/16F. A high-power holmium laser was used and the IRT was recorded during laser activation for up to 60 s at a laser power of 20 W, 40 W, and 60 W under gravity irrigation and manual pump irrigation. Results: Under gravity irrigation, FURS without UAS was associated with hazardous IRT at a laser power as low as 20 W for as short as 20 s of laser activation. The IRT was rendered borderline when UAS was used. This UAS buffering effect disappeared with the use of higher laser-power settings (40 W and 60 W) with the maximal IRT exceeding 60 °C. Moreover, laser activation at 60 W was associated with very rapid increase in IRT within few seconds. Under pump irrigation, laser activation at the highest power setting (60 W) for 60 s was associated with a safe IRT, even without the use of UAS. The maximal IRT was below 45 °C. Conclusion: The use of high-power Ho:YAG laser carries potentially harmful thermal effect when used under gravity irrigation, even when large-diameter UAS is used. High-power settings (> 40 W) require high irrigation flow. The use of UAS is advisable to reduce the IRT and balance any intra-renal pressure increase.