Neurite outgrowth and elongation of neural cells is the most important subject that is considered in nerve tissue engineering. In this regard, aligned nanofibers have taken much attention in terms of providing guidance for newly outgrown neurites. The main objective of this study was to fabricate aligned polyurethane nanofibers by electrospinning process and decorate them with gold nanoparticles to further investigate the synergistic effects of nanotopography, biological nerve growth factor (NGF) and electrical stimulations on neurite outgrowth and elongation of pheochromocytoma (PC-12) model cells. In this regard, smooth and uniform aligned polyurethane nanofibers with the average diameter of 519 ± 56 nm were fabricated and decorated with the gold nanoparticles with the average diameter of ∼50 nm. PC-12 cells were cultured on the various nanofiber surfaces inside the bio-mimetic bioreactor system and exposed either to NGF alone or combination of NGF and electrical stimulation. It was found that 50 ng/mL NGF concentration is an optimal value for the stimulation of neurite outgrowth. After 4 days of culture under 100 mV, 10 ms electrical stimulation in 1 h/day period it was found that the gold nanoparticle decorated aligned polyurethane nanofibers increased the neurite outgrowth and elongation more with the combinational NGF and electrical stimulation. © 2018 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 106A: 1604-1613, 2018.