Hydrogels are widely used as cell-culture platforms for various biomedical applications. With the biocompatible polymers as building blocks for hydrogels, it is often difficult to provide various physical properties to tailor to specific needs. In this study, bioactive and electrically conductive nanofibers consisting of conductive polymer (PEDOT:PSS) and biopolymer (photocrosslinkable gelatin) are prepared via electrospinning and further processed to generate short, diffusible nanofibers (< 10 um). These short nanofibers are incorporated into gelatin-based hydrogels to improve their mechanical properties as well as electrical conductivity which is otherwise generally non-existent in polymer-based materials. Their properties could be tuned in a wide range by controlling their physical parameters (e.g. concentrations, molar ratio, size, etc.). The nanofiber-infused hydrogel was used as a scaffold for cardiomyocytes to not only improve their viability, but also their electrophysiological functions, which was aided by the presence of conductive nanofibers. Overall, the nanofiber-based composite hydrogel systems presented in this study could provide unique and yet practical 3D cell culture platforms for biomedical applications.