Titanium dioxide (TiO2) has a number of uses in catalysis, photochemistry, and sensing that are linked to the reducibility of the oxide. Usually, bridging oxygen (Obr) vacancies are assumed to cause the Ti3d defect state in the band gap of rutile TiO2(110). From high-resolution scanning tunneling microscopy and photoelectron spectroscopy measurements, we propose that Ti interstitials in the near-surface region may be largely responsible for the defect state in the band gap. We argue that these donor-specific sites play a key role in and may dictate the ensuing surface chemistry, such as providing the electronic charge required for O2 adsorption and dissociation. Specifically, we identified a second O2 dissociation channel that occurs within the Ti troughs in addition to the O2 dissociation channel in O(br) vacancies. Comprehensive density functional theory calculations support these experimental observations.