Background and purpose Recently, stem cell therapy has been proposed to allow regeneration of radiation damaged salivary glands. It has been suggested that glial-cell-derived neurotrophic factor (GDNF) promotes survival of mice salivary gland stem cells (mSGSCs). The purpose of this study was to investigate the role of GDNF in the modulation of mSGSC response to irradiation and subsequent salivary gland regeneration. Methods Salivary gland sphere derived cells of Gdnf hypermorphic (Gdnf^wt/hyper) and wild type mice (Gdnf^wt/wt) were irradiated (IR) with γ-rays at 0, 1, 2, 4 and 8 Gy. mSGSC survival and stemness were assessed by calculating surviving fraction measured as post-IR sphere forming potential and population doublings. Flow cytometry was used to determine the CD24^hi/CD29^hi stem cell (SC) population. QPCR and immunofluorescence was used to detect GDNF expression. Results The IR survival responses of mSGSCs were similar albeit resulted in larger spheres and an increased cell number in the Gdnf^wt/hyper compared to Gdnf^wt/wt group. Indeed, mSGSC of Gdnf^wt/hyper mice showed high sphere forming efficiency upon replating. Interestingly, GDNF expression co-localized with receptor tyrosine kinase (RET) and was upregulated after IR in vitro and in vivo, but normalized in vivo after mSGSC transplantation. Conclusion GDNF does not protect mSGSCs against irradiation but seems to promote mSGSCs proliferation through the GDNF-RET signaling pathway. Post-transplantation stimulation of GDNF/RET pathway may enhance the regenerative potential of mSGSCs.