Defocused wide-field fluorescence microscopy was used to follow the 3D molecular rotational diffusion of a fluorescent probe molecule in a polymer thin film. The technique allows for visualizing the molecular reorientation both in-plane and out-of-plane. The local environmental change driven by heterogeneous dynamics of the polymer can be probed on a scale of I mu m as demonstrated by parallel imaging of several molecules. A multi-component rotational diffusion decay is observed which might reflect both different relaxation regimes of the polymer as well as rapid changes of the local environment. (c) 2006 Elsevier Ltd. All rights reserved.