Marilyn Y. McGinnis - US grants

The ultimate objective of this research program is to understand how androgens modulate brain function and regulate the expression of male sexual behavior (MSB). Our approach is to employ biochemical and molecular techniques in combination with behavioral studies to address questions on androgen action in male rat brain. We have expanded our behavioral investigations to include androgen-dependent measures such as partner preference, vocalization and anogenital investigation. All of these behaviors, along with copulation itself, are influential in the expression of MSB and will be referred to herein as androgen-dependent sexual behaviors. To achieve our goals, we will investigate three questions. Question l focusses on the neural sites of androgen action in regulating androgen-dependent sexual behaviors. The androgen receptor (AR) blocker, hydroxyflutamide (OHF), will be used to assess the role of AR in mediating androgen-dependent sexual behaviors in two previously unstudied androgen- concentrating brain sites: the ventromedial hypothalamic nucleus (VMN) and cranial nerve motor nuclei (n. ambiguous, hypoglossal n., facial motor n.). Question 2 focusses on the controversy over whether MSB is mediated by testosterone (T) acting via AR, or by aromatization to estradiol (E2). We will extend our recent results showing that both T and E2 are essential for MSB, by employing a newER blocker, RU58668, to assess the role of estrogen receptors (ER) in mediating androgen-dependent sexual behaviors. The site specificity of E2 action will be determined by implanting RU58668 directly into the medial preoptic area (MPOA), VMN or medial amygdala. Question 3 focusses on the identification of androgen-dependent genes in the hypothalamus-preoptic area (HYP-POA). We will use a subtractively hybridized probe to screen our HYP-POA cDNA library from T-treated adult male rats. RNAse protection assays will be used to verify that the isolated clones are androgen-dependent. Confirmed androgen- dependent clones will be sequenced to establish their identity. The answers to the three questions posed above are central to determining the consequences of T exposure in critical brain sites necessary for MSB, to resolving the controversy over the specificity of T action in the expression of androgen-dependent sexual behaviors, and will provide new insights into the cellular effects of T exposure. The results will also provide important knowledge on the effects of AR and ER blockers on behavior, which has clinical relevance for treating patients suffering from hormone-dependent tumors or from hormonal or sexual dysfunction.

DESCRIPTION: (Applicant's Abstract) The overall goals of this project are to assess the behavioral effects of anabolic androgenic steroids (AAS) and to understand the underlying mechanisms for these changes. An intact male rat model will be employed in order to provide empirical data on the behavioral and neuroanatomical consequences of AAS abuse. Three different classes of AAS are typically abused: testosterone esters, 19-nortestosterone esters, and 17-alpha-alkylated androgens. AAS users typically 'stack' these drugs (i.e., use several together). However, since little is known in rats about the behavioral effects of AAS other than testosterone, we propose to test the of one AAS from each category individually. The testosterone ester will be testosterone propionate (TP), the 19-nortetsosterone ester will be nandrolone (ND) and the 17-alpha-alkylated androgen will be stanozolol (ST). All are commonly abused AAS. The first two aims will investigate two frequently reported behavioral disturbances in human users: aggression and circadian rhythms. The first aim will determine the role of contextual cues in stimulating aggression during exposure to TP, ND or ST and whether these same cues are effective during withdrawal from AAS. The second aim will test the effects of AAS on circadian rhythms by measuring wheel running activity in 12:12 LD, phase reversal and constant dark during AAS exposure and during AAS withdrawal. Because AAS abuse is on the rise in adolescent males, the experiments proposed in the third aim will assess the effects of AAS on circadian rhythms, aggression, and reproductive function in peripubertal male rats. We will also examine whether the behavioral effects of AAS exposure during this critical period of development are reversible or permanent. To assess whether the ability of these AAS to bind to androgen receptors is related to their behavioral effects, the fourth aim will assess the binding characteristics of TP, ND and ST on androgen receptors in brain. The fifth aim will follow up our novel finding that chronic AAS exposure results in a loss of detectable cells in specific brain nuclei. We will determine if the loss of detect ability in cells of TP-treated males occurs in neurons or glia, and if the affected cells contain androgen receptors. Double label immunofluorescence and confocal microscopy will be used in conjunction with volume/size analyses to investigate the cellular characteristics of affected cells. These studies will aid in understanding the effects of AAS abuse on behavior in bot adults and juveniles, and will delineate possible mechanisms underlying AAS effects on brain.

DESCRIPTION (provided by applicant): The prevalence of anabolic androgenic steroid (AAS) use has risen in teenagers and is now a major health problem. AAS use during adolescence is particularly alarming because puberty is a hormonally sensitive period during which adult behavioral patterns develop, and one consistent effect of AAS use is increased aggression. The goal of this research is to determine the factors underlying aggressive and impulsive behaviors in adolescent AAS users, employing rats as the animal model. The proposed studies will continue our work on AAS-exposed pubertal males. Aim I will identify early experiential factors that predict whether aggression will be potentiated in AAS-treated males. The effects of early exposure to low serotonin and to social subjugation will be investigated. The effects of exposure to social subjugation on serotonin will be assessed. Aim II will investigate the role of impulsivity in AAS-induced aggression. The first aim will employ a test battery we have developed for measuring impulsivity in a sociosexual encounter. An advantage of this test is that it is biologically relevant and does not require extensive training or teaming. In the second experiment, the role of AAS in mediating impulsivity will be determined using the androgen receptor antagonist, hydroxyflutamide. In the third experiment, the role of serotonin in mediating impulsivity will be tested in AAS and gonadally intact rats using the SSRI, fluoxetine. Aim III will investigate the long term effects of pubertal AAS exposure. The first experiment will identify long term effects of pubertal exposure to low brain serotonin in AAS-exposed males. The second experiment will determine whether increased aggressive behavior in socially subjugated males persists into adulthood. In Aim IV we will assess the neuroanatomical effects of pubertal AAS exposure. Since gonadal hormones influence dendritic spine density, we will determine whether spine density is affected by AAS exposure during puberty. Spine densities in the medial amygdala and dorsal hippocampus will be quantified using the fluorescent dye, Dil. Disruption of the endogenous hormonal milieu during puberty may lead to irreversible behavioral and neurochemical consequences, and have a long-term negative impact on adult behavioral patterns. By superimposing AAS exposure on the developing nervous system, these studies will answer important questions regarding the effects of AAS use on the maturation of adolescent brain and aggressive behavior.