17β‐ and 17α‐estradiol are non‐competitive inhibitors of dopamine uptake: implications for Parkinson's disease models and therapeutics

Despite compelling literature documenting the cyto‐ and neuro‐protective activities of various estrogens in a wide variety of cell and animal models, several large clinical trials of hormone replacement therapies have failed to detect such beneficial responses in humans. To some degree, this failure stems from profound hormonal responses that counterbalance the protective effects. This limitation could be circumvented by the development of less hormonally active estrogen analogs that retain the cytoprotective activities of the parental hormones. For example, 17α‐estradiol (17α‐E2) is at least 200‐fold less active than 17β‐estradiol (17β‐E2) as a transactivating hormone, yet it is equipotent as a cytoprotectant in many cell types exposed to a variety of stressors, such as amyloid toxicity, serum withdrawal, oxidative stress, and glutamate excitotoxicity. Both estradiol isomers stabilize mitochondrial function under pathogenic conditions, and both have shown efficacy in animal models of stroke. The results for models of Parkinson's disease differ, and 17β‐E2, but not 17α‐E2, has been reported previously to provide neuroprotection against MPTP toxicity. However, such findings must be regarded in light of reports that 17β‐E2 impedes cellular uptake of MPTP by the dopamine transporter (DAT), thereby forestalling development of the lesion rather than providing authentic cytoprotection. We report here that both 17α‐E2 and 17β‐E2 show classical, non‐competitive, inhibition of dopamine uptake by the human DAT (hDAT) transfected into L tk − mouse fibroblasts, HEK293 human embryonic kidney cells, and SH‐SY5Y human neuroblastoma cells. IC 50 values vary slightly with cell type, but are 400 and 70 nM for 17α‐E2 and 17β‐E2, respectively, in SH‐SY5Y cells. We also evaluated 17α‐E2 in the 6‐OH‐dopamine (6‐OHDA) model of PD, where toxin uptake occurs via the DAT. In these studies, treatment with estrogen was delayed for 6 hr after unilateral intrastriatal 6‐OHDA injection to allow for toxin uptake. After 6 hr, a loading dose of 17α‐E2 (100 µg/kg in sesame oil) was injected s.c., accompanied by implanting of a sustained release silastic device. After 21 days, animals treated with 17α‐E2 showed significant improvements in both gait asymmetry and apo‐morphine induced rotational defects. These positive results encourage evaluation of 17α‐E2 in a host of neurodegenerative diseases. Drug Dev. Res. 66:160–171, 2006. © 2006 Wiley‐Liss, Inc.