Porous anatase TiO2 spheres have been synthesized by a microwave-assisted hydrothermal reaction of spherical particle precursors followed by annealing in air. The synthesized TiO2 spheres are formed by interconnected nanocrystals with size of 8.7 nm in average and have grain diameters of 250–400 nm. After annealing at 500 °C, the TiO2 samples maintain spherical shape and develop highly mesoporous characteristics with a specific surface area of 151 m2 g−1. The TiO2 samples annealed at 750 °C consist of larger aggregated particles with diameters of 500–900 nm and still retain mesoporous anatase structure, but with a reduced specific surface area of 25.6 m2 g−1. Electrochemical studies reveal that the porous TiO2 spheres annealed at 500 °C own very high and stable lithium ion (Li+) storage capacities of 207, 184, 166, and 119 mA h g−1 at 0.5, 1, 2, and 5 C (850 mA g−1) rates, respectively, owing to their highly porous nanostructures and fine spherical morphology. In contrast, the TiO2 spheres annealed at 700 °C exhibit modest electrochemical performance due to their reduced pore structures and larger crystallite size. The prepared porous TiO2 spherical particles show great promise for use as high performance anode materials for lithium ion batteries (LIBs).