Despite the application of nerve grafts and considerable microsurgical innovations, the functional recovery across a long peripheral nerve gap is generally partial and unsatisfactory. Thus, additional strategies are required to improve nerve regeneration across long nerve gaps. Hydrogen possesses antioxidant and anti-apoptotic properties, which could be neuroprotective in the treatment of peripheral nerve injury; however, such a possibility has not been experimentally tested in vivo. The aim of the present study was to investigate the effectiveness of hydrogen-rich saline in promoting nerve regeneration after 10-mm sciatic nerve autografting in rats. The rats were randomly divided into two groups and intraperitoneally administered a daily regimen of 5 ml/kg hydrogen-rich or normal saline. Axonal regeneration and functional recovery were assessed through a combination of behavioral analyses, electrophysiological evaluations, Fluoro-Gold™ retrograde tracings and histomorphological observations. The data showed that rats receiving hydrogen-rich saline achieved better axonal regeneration and functional recovery than those receiving normal saline. These findings indicated that hydrogen-rich saline promotes nerve regeneration across long gaps, suggesting that hydrogen-rich saline could be used as a neuroprotective agent for peripheral nerve injury therapy.