A primary goal of the proposed studies is the identification of androgen regulated gene products which are essential for the neuromodulation of male sexual behavior (MSB). This project will address two key aspects of androgen action. First, we will challenge the hypothesis that gradual changes in protein synthesis occurring in certain androgen-concentrating brain nuclei underlie the long lag time required for the postcastration decline of MSB as well as the long testosterone (T) exposure required for the restoration of behavior. We have shown that implants of the protein synthesis inhibitor, anisomycin (ANI), into the medial preoptic area (MPOA) prevent restoration of MSB.
Specific Aim 1 will extend this finding to establish whether ANI implants into other androgen- concentrating areas, the hypothalamic ventromedial nucleus (VMN), medial amygdala (AME) and lateral septum (SEPT) also prevent restoration of MSB.
In Specific Aim 2 we will implant ANI into sites where restoration of MSB was inhibited to determine if maintenance of MSB is also inhibited.
Specific Aim 3 will use 2-dimensional gel electrophoresis (2DE) to extend our finding that 5 MPOA proteins are increased by T, and establish whether these same proteins are induced by T in VMN, AME and SEPT.
Specific Aim 4 will use 2DE to determine if these proteins gradually disappear after castration and reappear following T exposure.
Specific Aim 5 is to identify the cellular and subcellular localization of these proteins by immunocytochemistry. The results of these experiments will: 1) identify brain nuclei essential for mediating the genomic effects of T on the restoration and maintenance of male sexual behavior, 2) single out a small population of proteins uniquely expressed by T and demonstrate if there is site specificity in their induction, and if their disappearance/reappearance coincides with the decline and restoration of MSB, and 3) identify potential functional sites of action for these proteins. The second basic question is how specific are the efects of T? Specific Aim 6 will use Northern blots and in situ hybridization to establish whether androgen receptor (AR) and estrogen receptor (ER) mRNA are colocalized in the same neurons, and whether T, and estradiol (E2) are similar or different in inducing the message for these receptors in MPOA, VMN, AME and SEPT.
Specific Aim 7 will employ 2DE of micropunched MPOA, VMN, AME and SEPT to detect local similarities and differences in the profiles of proteins uniquely expressed by T, E2 and DHT. These results should answer whether aromatization of T to E2 is in fact an essential step in evoking MSB, and point to salient differences in the actions of T vs its metabolites that could account for differences in their behavioral effects. The results of these studies will make it possible to devise a molecular approach to understanding reproductive behavior and yield insight into the treatment of hormone dependent tumors, neurologic diseases and injuries influenced by androgens, androgen resistance syndromes, and sexual dysfunctions such as impotence.
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