Introduction: Spinal cord injuries are accompanied with significant demyelination of axons and subsequent locomotor dysfunction. To identify the extent of damage following electrolytic lesion of ventrolateral white matter, essential area for initiation of locomotor activity, we assessed demyelination as well as alteration in motor performance. Moreover, the protective effect of estradiol as a candidate treatment for preservation of myelin and locomotor activity after injury was examined due to its anti-apoptotic and anti-inflammatory activities. Methods: A unilateral electrolytic lesion positioned in the right ventrolateral funiculus (VLF) was applied following laminectomy at T8-T9. In the estradiol-treated injury group, animals received a pharmacological single dose of estradiol valerate (4 mg/kg) at 30min post injury. Locomotor function was assessed using rotarod and open field tasks during 4 weeks after injury. Results: Obtained results showed significant demyelination at the site of injury and caudal areas following lesion as well as altered motor performance. Post-spinal cord injury administration of estradiol enhanced white matter maintenance at the site of lesion, restored the level of myelin basic protein (MBP), decreased TUNEL positive cells and improved functional recovery. Conclusion: Taken together, these results indicate that demyelination after lesion in VLF may be a contributing factor to limited motor performance, and suggest that pharmacological doses of estradiol may have an early protective effect through sparing of white matter.