It has previously been demonstrated that 1,25‐dihydroxyvitamin D₃ [1,25(OH)₂D₃] administration, whether in cell cultures or in vivo to rats, increases glial cell line‐derived neurotrophic factor (GDNF) expression levels, suggesting that this hormone may have beneficial effects in neurodegenerative disorders. This study was carried out to explore the effects of 1,25(OH)₂D₃ administration in a 6‐OHDA‐lesioned rat model of Parkinson's disease on GDNF and tyrosine hydroxylase (TH) expression in substantia nigra (SN) and striatum. Two groups of animals received 1,25(OH)₂D₃ intraperitoneally, the first group 7 days before the unilateral injection of 6‐OHDA into the medial forebrain bundle (MFB) and the second group 21 days (days 21–28) after the unilateral injection of 6‐OHDA. Animals of both groups were sacrificed on day 28. In addition, two other groups received a unilateral injection of either saline or 6‐OHDA into the MFB. Rats were killed, and the SN and striatum were then removed for GDNF and TH determination. Striatal GDNF protein expression was increased on the ipsilateral with respect to the contralateral side after 6‐OHDA injection alone as well as in 1,25(OH)₂D₃‐treated rats before or after 6‐OHDA administration. As expected, 6‐OHDA injection induced an ipsilateral decrease in TH‐immunopositive neuronal cell bodies and axonal terminals in the SN and striatum. However, treatment with 1,25(OH)₂D₃ before and after 6‐OHDA injection partially restored TH expression in SN. These data suggest that 1,25(OH)₂D₃ may help to prevent dopaminergic neuron damage. © 2008 Wiley‐Liss, Inc.