Androgen Receptors in the Middle Aged Male Rat Brain: Influence of Testosterone Deprivation on Expression




Contreras, Jo
Smith, Charity B.S.
Cunningham, Rebecca
Fort*, Callie
Cuellar*, Elric
Lopez, Gladys
Metzger, Daniel
Oppong-Gyebi, Anthony
Schreihofer, Derek


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Purpose: 1) To determine whether long-term testosterone deprivation (LTTD) alters the levels and/or distribution of androgen receptors in the middle-aged male rat brain and 2) to determine whether testosterone replacement after LTTD influences androgen receptor levels. Methods: Twelve-month old male Fischer 344 rats were left intact or castrated for 2 weeks and replaced with subcutaneous implants containing testosterone (STTD). Additional groups were castrated for 10 weeks before being treated with testosterone (LTTD+T) or cholesterol (LTTD). Four weeks later, rats were euthanized and brains were collected for immunoblotting and immunohistochemistry (IHC) for androgen receptors (AR) using antibodies targeting the N-terminus or C-terminus of the protein. The cerebral cortex, hippocampus, thalamus, hypothalamus were examined, and testes were used as positive control tissue. Results: Contrary to expectations, the full-length AR (116 kDa) was barely detectable in the hippocampus and cerebral cortex by immunoblotting with antibodies directed to either end of the AR. Rather, smaller fragments were readily detected. Examination of the size of these fragments (~30, 37, 50, and 80 kDa) and consultation of the literature for the human AR, suggested that they represented calpain-dependent cleavage fragments. A series of control experiments was performed in an attempt to extract the full-length AR using rat testes as a positive control tissue. Protease inhibitors, EDTA, and the AR agonist dihydrotestosterone failed to reduce the appearance of fragments. Interestingly, the pattern of fragments from the hippocampus (80 [greater than] 37 kDa) differed from that from the cortex (37 [greater than] 80 kDa) suggesting differential processing. IHC of coronal brain sections though the forebrain revealed nuclear AR staining consistent with full-length AR in regions of high expression, including the hypothalamus. In agreement with immunoblotting AR staining in the cortex and hippocampus appeared to be cytoplasmic, rather than nuclear. No significant differences were observed between treatment groups. Conclusions: These data suggest that AR protein in some areas of the middle-aged male rat brain is rapidly degraded into fragments with altered localization and potential for transcriptional activity and/or signaling functions. Although no differences in expression were apparent between treatment groups, the differential processing of AR in the rat brain is a novel finding warranting further investigation.