The measurement and control of the temperature in microscopic systems, which are increasingly required in diverse applications, are fundamentally important. Yet, the measurement of the three-dimensional (3D) temperature distribution in microscopic systems has not been demonstrated. We propose and experimentally demonstrate the measurement of the 3D temperature distribution by exploiting the temperature dependency of the refractive index (RI). Measurement of the RI distribution of water makes it possible to quantitatively obtain its 3D temperature distribution above a glass substrate coated with gold nanorods with submicrometer resolution in a temperature range of 100 °C and with a sensitivity of 2.88 °C. The 3D temperature distributions that are obtained enable various thermodynamic properties including the maximum temperature, heat flux, and thermal conductivity to be extracted and analyzed quantitatively.