Two layer thermal barrier coatings (TBCs) were prepared by EB-PVD (Electron Beam-Physical Vapor Deposition) at different substrate temperatures in the range of 823 to 1123K, and their microstructure was investigated with SEM and AC impedance as a function of substrate temperature and thermal cycling time. YSZ layer of all TBCs samples is in column structure, but the grain size and growth orientation are different with substrate. In this research, impedance spectra (IS) was measured as a function of thermal cycling between 1323K and 298K for these thermal barrier coatings. Grain boundary and bulk can be distinguished from analysis of AC impedance spectroa to provide information about the relation between microstructure and electric properties.The change in IS until failure was found to be related with the thickness, microcracks and macrocracks of TGO and the change in the interfacial of TGO/YSZ.