Tension-tension fatigue tests at 750 °C were performed to state on fatigue structure-properties of Ni-based superalloy in long-life regime under high temperature by means of ultra-depth-of-field overlay three-dimensional, scanning electron microscopy, energy dispersive spectrometer, electron back scattered diffraction, focused ion beam and transmission electron microscope. Results show that interior crystallographic plane-fisheye failures are divided into the single plane & multiple planes cracking. The formation of plane with micro-crack initiation and growth is mainly under Mode Ⅱ cracking from the Copper grain with large size and the highest Schmid factor or the Goss grain with small size and high Schmid factor. Moreover, owing to high thermal activation energy, grain-related interior cracking is mainly induced by climbing and bypassing mechanisms with bowing of dislocations and zig-zag dislocations, and assisted by shearing mechanism with tangled dislocations. Finally, the interior plane-fisheye cracking behavior with microstructure in high-temperature and vacuum environment is elucidated.