Neurotropic action of androgens: principles, mechanisms and novel targets

Experimental Gerontology 39 (2004) 1651–1660 www.elsevier.com/locate/expgero

Review article

Neurotropic action of androgens: principles, mechanisms and novel targets Vladimir K. Patcheva,*, Jens Schroedera, Franziska Goetzb, Wolfgang Rohdeb, Alexandre V. Patchevb a

Male Health Care 2, Corporate Research Gynecology and Andrology, Schering AG/Jenapharm, Otto-Schott-Str. 15, D-07745 Jena, Germany b Institute of Experimental Endocrinology, School of Medicine Charite´, Humboldt University, Berlin, Germany Received 27 May 2004; accepted 7 July 2004 Available online 7 October 2004

Abstract The importance of androgen signaling is well recognized for numerous aspects of central nervous system (CNS) function, ranging from sex-specific organization of neuroendocrine and behavioral circuits to adaptive capacity, resistance and repair. Nonetheless, concepts for the therapeutic use of androgens in neurological and mental disorders are far from being established. This review outlines some critical issues which interfere with decisions on the suitability of androgens as therapeutic agents for CNS conditions. Among these, sex-specific organization of neural substrates and resulting differential responsiveness to endogenous gonadal steroids, convergence of steroid hormone actions on common molecular targets, co-presence of different sex steroid receptors in target neuronal populations, and in situ biotransformation of natural androgens apparently pose the principal obstacles for the characterization of specific neurotropic effects of androgens. Additional important, albeit less explored aspects consist in insufficient knowledge about molecular targets in the CNS which are under exclusive or predominant androgen control. Own experimental data illustrate the variability of pharmacological effects of natural and synthetic androgens on CNS functions of adaptive relevance, such as sexual behavior, anxiety and endocrine responsiveness to stress. Finally, we present results from an analysis of the consequences of aging for the rat brain transcriptome and examination of the influence of androgens on differentially expressed genes with presumable significance in neuropathology. q 2004 Elsevier Inc. All rights reserved. Keywords: Androgen; Brain; Behavior; Stress response; Target genes; Aging

Androgens significantly contribute to several aspects of central nervous system (CNS) development, circuit formation and function, and resistance against noxious impact. Their effects on the establishment and operation of neural components of endocrine regulation and manifestation of behavioral patterns associated with reproduction are well described. More recent research has suggested a role for androgens in CNS physiology and pathology beyond reproduction, such as cognitive performance, neurogenesis, resistance and survival. Dissemination of novel knowledge on neurotropic actions of androgens has prompted ideas about their therapeutic use in medical conditions, especially

* Corresponding author. Tel.: C49 3641 646211; fax: C49 3641 646357. E-mail address: [email protected] (V.K. Patchev). 0531-5565/$ - see front matter q 2004 Elsevier Inc. All rights reserved. doi:10.1016/j.exger.2004.07.011

those which may be directly or circumstantially related to aging in both, the endocrine and central nervous systems. Although a critical involvement of androgens in CNS function has been long recognized, identification of pure neural androgenic effects has been difficult. This is probably due to three inherent issues which are specifically associated with: (i) regional distribution of androgen receptors in the CNS and co-occurrence of confounding targets of steroid hormone action in the identified neuronal populations; (ii) cross-talk between molecular pathways of steroid hormone signaling, and (iii) the chemical nature and biotransformation of androgens in the CNS. Undoubtedly, the androgen receptor (AR) is the principal mediator of androgen action in the nerve cell, with signal transmission being based on nuclear translocation of the ligand-bound AR and transcriptional regulation of target

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genes endowed with appropriate DNA sequences (also known as response elements). Thus, densities of ARencoding mRNA and/or receptor protein in individual brain structures may serve as a reliable tool for the prediction and verification of functional consequences of androgen signaling in the brain. Indeed, regional distribution maps of AR in the mammalian brain (Simerly et al., 1990; Abdelgadir et al., 1999) could convincingly explain androgen-induced changes in several aspects of CNS function. Likewise, localization of AR in the brain is indicative of their involvement in the control of reproductive function and sexual behavior, and also endocrine responsiveness to stress, mood, cognition and nociception. The relatively high abundance of AR in substantia nigra (Simerly et al., 1990; Ravizza et al., 2003) is of note, although its possible significance in health and disease is only just being recognized (Okun, et al., 2002a,b). However, past anatomical studies have revealed that AR-expressing brain areas largely overlap with structures which are richly endowed with estrogen receptors (ER) (Simerly et al., 1990; Shughrue et al., 1997). The juxtaposition of these distribution maps (Fig. 1) reveals high co-localization of androgen- and estrogen-concentrating neuronal population. This anatomic background, together with in loco-biotransformation of androgens to agonists of the ER pose certain difficulties in assigning neurotropic effects of the major natural androgen, testosterone, to activation of either AR or ER. Interaction of nuclear receptors with distinct DNA sequences in the promoter region of target genes is a prerequisite for their influence on gene transcription. However, there are few CNS-specific genes whose transcriptional regulation depends on the exclusive presence of androgen response elements (ARE). Thus, identification of distinct downstream targets for AR in the brain remains a challenging issue. However, expectations for a major breakthrough are rather modest, due to extensive similarity between DNA binding domains of the so-called class I steroid hormone receptors, encompassing those for androgens, progestesterone, glucocorticoids and mineralocorticoids (Glass, 1994; Zilliacus et al., 1995). This homology opens numerous possibilities for promiscuous interactions and cross-regulation of target genes by more than one steroid hormone receptor. Furthermore, DNA response elements are embedded within larger enhancers which integrate complex signals transmitted by other transcription factors. Expression of several steroid receptors in the CNS is subject to homologous down-regulation by their cognate ligand (DonCarlos and Handa, 1994; Patchev et al., 1994). In this context, the AR shows a different behavior, as its presence has been shown to increase following endogenous release or exogenous administration of androgens (McAbee and DonCarlos, 1998, 1999; Lu et al., 1998; Xiao and Jordan, 2002). Thus, it seems that physiological androgen levels create target densities needed for the full

Fig. 1. Schematic diagram of the regional distribution of androgen (AR, blue) and estrogen receptors (ERa, red; ERb, green) in virtual rostro-caudal sections of the rat brain, and their possible involvement in CNS function. The scheme is based on data from Shughrue et al. (1997) and Simerly et al. (1990). Templates for coronal section levels were extracted from L.W. Swanson (1998/1999) Brain Maps: Structure of the Rat Brain, 2nd Edition, Elsevier, Amsterdam.

manifestation of neurotropic androgen effects. This phenomenon is best exemplified by the androgen-mediated stimulation of aromatase expression and activity in the brain. Aromatase regulation in the CNS is AR-dependent and, in the male brain, the expression of this enzyme correlates with both, systemic androgen levels and AR density (Roselli and Resko, 1993; Abdelgadir et al., 1994). The major endogenous androgen, testosterone, is transformed in the CNS by 5a-reductase and aromatase to the pure AR-agonist dihydrotestosterone (DHT) and estradiol, respectively. These testosterone-metabolizing enzymes are present in several sex-hormone-sensitive brain areas (Lauber and Lichtensteiger, 1994; Roselli et al., 1998; Melcangi et al., 1998) and are subject to control by androgens, albeit to a different extent (Roselli and Resko, 1984; Celotti et al., 1997; Torres and Ortega, 2003).

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The latter implies that generation of both, DHT and estrogens and, subsequent degree of AR- and ER-activation ultimately depends on systemic androgen levels and, thus, individual contribution of each receptor class cannot be readily discerned. Control of male sexual behavior is probably the best studied and unanimously recognized neurotropic action of androgens. Extensive research during the past decades led to the postulate that activation of ER by testosteronederived estrogens plays a critical role for the perinatal organization of neural circuits which account for the manifestation of male sexual behavior (Arnold and Gorski, 1984; Pilgrim and Hutchison, 1994). This view has been disputed in the past (Toran-Allerand, 1984; Bloch and Mills, 1995) and more recently (Cooke et al., 1998), with novel molecular species and mechanisms being taken into consideration (Blaustein, 2004; Arnold et al., 2004; De Vries, 2004; Toran-Allerand, 2004; Garcia-Segura and McCarthy, 2004) for re-evaluation of the role of individual sex steroids in brain organization during early ontogeny. Importantly, data accumulated from observations in animals with targeted disruption of AR or ER (Wersinger et al., 1997; Sato et al., 2004) cannot fully explain the contribution of individual hormone receptors to the behavioral phenotype. However, without question, the importance of hormone signaling during pre- and perinatal brain development for the formation of sex-specific behavioral patterns is unchallenged. Disruption of this process in early ontogeny, as it occurs in conventional transgenics, results in faulty organization of relevant neural circuits. Thus, behavioral induction in adult individuals, who have experienced physiological sex-specific brain differentiation, remains a primary tool for the examination of neurotropic effects of sex hormones. Aleviation of CNS symptoms of sex hormone deficiency, resulting from age-related decline in gonadal endocrine activity is a major challenge in hormone therapy. Despite frequently addressed safety issues, it is obvious that sudden (in the female) or insidious (in the male) decreases in sex steroid levels are associated with symptoms of withdrawal which can be managed by hormone therapy. However, strategy for the therapeutic application of androgens in the aging male is still far from being firmly established. While causal relationship between androgen deficiency and neurological/mental symptoms requires confirmation, clinical evidence of such an interaction is steadily accumulating. Drug discovery and development, on the other hand, still await answers to questions of critical importance for the future; e.g. (1) Are there medical conditions in neurology and psychiatry for which decisive benefit from androgen therapy can be expected? (2) Which pharmacological properties of androgens deserve emphasis, in order to achieve therapeutic effects beyond those of mere supplementation? and (3) Is it possible to identify target molecules in the aging brain which are controlled by

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androgens and plausibly involved in the pathogenesis of CNS diseases?

1. Androgen effects on behavior and neuroendocrine regulation are mediated by different sets of sex steroid receptors in the brain Alterations of sexual activity, anxiety and magnitude of the endocrine response to stress are considered reliable descriptors of neuropharmacological effects of steroid hormones which also permit evaluation the organism’s adaptive capacity. Medical implications can also be easily deduced from these descriptors: decreased sexual activity is a pathognomonic symptom of hypogonadism, whereas anxiety and poor control of pituitary–adrenal responsiveness to stress are considered hallmarks of affective disorders (e.g. major depression) (Sapolsky and Plotsky, 1990). Age-related androgen deficiency has been associated with affective disorders, and androgens have been sporadically used in combination with other drugs as treatment for this condition (Seidman et al., 2001; Carnahan and Perry, 2004; Epperson et al., 1999). In the following study we compared the effects of three androgens with different pharmacological profiles on the above-listed parameters with the aim to elucidate whether (i) biotransformation to estrogens and (ii) pronounced anabolic properties differentially contribute to behavioral and neuroendocrine actions of androgens. We used the natural occurring androgens testosterone (an AR agonist which can be aromatized to estradiol or acted upon by 5a-reductase to yield DHT) and dihydrotestosterone (the cognate endogenous ligand of the AR which cannot be aromatized), as well as the synthetic steroid anadrol (oxymetholone), a 5a-reduced androgen with pronounced anabolic properties. Anadrol cannot be aromatized. Based on its anabolic profile, anadrol is approved by the FDA for the treatment of hypoplastic anemias, and has been an investigational drug for several other conditions (e.g. HIV-associated wasting, antithrombin III deficiency, growth impairment and chronic cardiomyopathy) (Pavlatos et al., 2001). However, there are no consistent reports on clinical experience with this compound in neurological and mental conditions. These steroids were administered s.c. to gonadectomized adult male rats for 3 months at daily doses of 200 and 600 mg/kg. In order to minimize bias, the animals were repeatedly screened for manifestation of sexual competence before castration, and lack of male sexual behavior was documented before initiation of treatment. During the last 14 days of treatment, male sexual activity was examined in three independent sessions of exposure to an estrogenprimed female for 5 min. The number of complete mounts (i.e. with pelvic thrusts) within the observation period was recorded as a measure of male sexual activity. Influence of test compounds on anxiety was investigated by monitoring the animals’ behavior in the elevated

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plus-maze, a widely used paradigm for demonstration of anxiogenic/anxiolytic drug effects (File et al., 1988). Video records during observation periods of 3 min for each individual were evaluated by an independent observer, and time spent in the closed compartment of the apparatus was scored as a measure of anxiety. Number of entries (transitions) between the closed and open compartments were used to monitor possible compound effects on general locomotor activity. Effects of test compounds on basal and stress-induced activity of the pituitary–adrenal axis were documented by measurement of serum corticosterone levels at the diurnal zenith of adrenocortical output (i.e. in the rat, at the onset of the circadian dark phase) and 30 min after exposure to acute emotional stress (i.e. intermittent air-blows for 2 min delivered in a non-familiar environment), respectively. Blood samples were collected by microincisions of the tail vein, and serum corticosterone levels were determined by a specific radioimmunoassay. Experimental procedures were in compliance with NIH guidelines and national regulations on animal welfare. As shown in Fig. 2, chronic administration of the anabolic androgen anadrol failed to induce male sexual activity in castrated animals which have previously reproducibly demonstrated sexual competence. The potent, however non-aromatizable, AR-agonist DHT showed a non-significant trend toward induction of sexual behavior at the highest dose tested, while at this dose, testosterone was able to fully restore mounting activity to the level seen in intact rats. These findings largely confirm data from previous investigations (Vagell and McGinnis, 1998; Scordalakes et al., 2002) suggesting that activation of the ER is a necessary, albeit not sufficient, prerequisite for induction/restoration

Fig. 2. Induction of sexual behavior in gonadectomized (GDX) adult male rats following 120 days of treatment with testosterone (T), dihydrotestosterone (DHT) and anadrol (ANA) at daily s.c. doses of 200 and 600 mg/kg. Data depict meanGSEM of three independent sessions; each treatment group consists of 8–9 individuals; asterisks denote significant differences (p!0.05 and less) as compared to vehicle-treated controls.

of mating behavior in the male. Also, in accordance with earlier studies (Baum and Vreeburg, 1973; Cooke et al., 2003), our observations in DHT-treated animals suggest a possible contribution of pure AR-mediated effect which, however, may require a supporting estrogen signal for the full manifestation of male sexual activity. According to studies using a different experimental protocol (Clark and Harrold, 1997; Clark et al., 1997), anabolic androgens were at best inefficient with regard to induction of male sexual activity. In the case of anadrol, however, their effects were described as detrimental. While our studies could not demonstrate behavioral inhibition, we could show that anabolic steroids fail to induce male sexual activity in the absence of endogenous sex steroids. Thus, if stimulation of male sexual behavior is considered as a reliable sign of neurotropic action of androgens, the conclusion can be drawn that testosteronelike pharmacological features, such as liability to aromatization, a role as substrate in this reaction, and sufficient agonistic activity at the AR, may outline the profile of compounds capable of stimulation of sexual activity (Roselli, 1998). The anabolic androgen anadrol displayed significant anxiolytic effects at both doses examined, whereas testosterone was effective only in the higher dose (Fig. 3). On the other hand, DHT failed to produce measurable anxiolysis, as defined by the time spent in exploration of the open compartment of the plus-maze. All androgens tested showed a dose-dependent trend toward increasing locomotor activity; however, the pre-set level of significance could not be achieved in any treatment group. At the first glance, these data fail to provide a clear answer of whether a defined pharmacological profile of an androgen may specifically account for its pronounced anxiolytic action, as both, the aromatizable natural androgen testosterone and the anabolic steroid anadrol showed almost indistinguishable anxiolytic effects. While anxiolytic and anti-conflict potential of testosterone has been extensively documented in the past (Bitran et al., 1993; Bing et al., 1998; Aikey et al., 2002; Frye and Seliga, 2001), experimental studies with anabolic steroids have suggested that these compounds may actually display anxiogenic action (Minkin et al., 1993), thus in opposition to the present data. It is certainly debatable whether the relatively simple conflict between exploratory drive in a novel environment and the rodent-specific natural preference for safe dark and closed spaces exploited in the plus-maze paradigm may reflect the entire complexity of mechanisms involved in the control of anxiety. Natural and synthetic androgens may interact with several aspects of neurotransmission in different spatial and temporal modes, thus influencing the entire chain of behavioral responsiveness to aversive stimuli, from perception to assessment and reaction adequacy. Although these issues have been addressed in several excellent reviews (Clark and Henderson, 2003; Mong and Pfaff, 2003), two interdependent conclusions can be drawn from the present investigation: (i) activation of AR alone by DHT is not sufficient

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Fig. 3. Anxiolytic effects of long-term treatment with 200 and 600 mg/kg of testosterone (T), dihydrotestosterone (DHT) and anadrol (ANA) in gonadectomized (GDX) male rats. The left panel shows time spent in the open arms of the plus maze (meanGSEM of 7–9 individuals) during an observation period of 3 min; the right panel describes general locomotion, as revealed by the number of entries into all maze compartments. Asterisks indicate significant differences to vehicle-treated GDX animals.

for measurable anxiolysis upon long-term treatment and, consequently, (ii) the anxiolytic effects of the anabolic steroid anadrol cannot be solely ascribed to its interaction with the AR. As effects of anadrol have not been previously examined in a similar setting, speculations on potential mechanisms of its anxiolytic action are certainly premature. Yet, it is worth mentioning that androgens, including compounds with anabolic properties, may target neurochemical mechanisms which operate through steroidmediated interactions with GABAA-ergic transmission (Frye and Reed, 1998; Jorge-Rivera et al., 2000). Also, it is prudent to consider that the outcome of anxiogenic/ anxiolytic action of synthetic androgens may be biased by their capacity to interfere with glucocorticoid signaling,

which significantly contributes to control of anxiety (Korte, 2001). Long-term administration of androgens did not result in significant changes in adrenocortical secretions at the time of diurnal physiological activation of the pituitary–adrenal axis, although a stimulatory effects of anabolic androgens has been previously reported (Schlussman et al., 2000). Stressinduced corticosterone secretion soon after exposure to emotional stimuli was significantly suppressed in all treatment groups (Fig. 4). Among the compounds used, this effect was most pronounced under testosterone treatment. The gradual difference between the suppressive efficacy of testosterone and DHT or anadrol is suggestive of a possible additional involvement of non-AR-operating mechanisms.

Fig. 4. Serum corticosterone levels in gonadectomized (GDX) rats which have received s.c. injections of testosterone (T), dihydrotestosterone (DHT) and anadrol (ANA) at daily doses of 200 and 600 mg/kg for 120 days. The left panel shows corticosterone secretions measured under quiescent conditions at the time of circadian zenith of adrenal activity; the right panel monitors corticosterone levels in samples collected 30 min after exposure to brief emotional stress. Data are presented as meanGSEM; each treatment group consists of 7–10 animals; asterisks indicate significant differences to vehicle-treated GDX rats (p!0.05).

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The discussion of these differences evolves into two major directions: (i) interactions of sex steroids with neuronal populations which account for the limbic-hypothalamic drive on the pituitary–adrenal axis and (ii) capacity of androgens to restrain this neuroendocrine axis in a glucocorticoid-like fashion. Besides earlier studies on these interactions (Handa et al., 1994; Patchev and Almeida, 1998), later studies dissected of androgen- and glucocorticoid-dependent mechanisms which regulate the expression of two principal hypothalamic activators of pituitary–adrenal secretions, CRH and vasopressin (Viau et al., 1999; Viau, 2002). These reports convincingly demonstrated that androgens and glucocorticoids jointly contribute to the control of pituitary– adrenal secretions under both, basal and stress-related conditions, and also influence the discrimination threshold of central mechanisms which account for the efficacy of glucocorticoid-mediated feedback in the pituitary–adrenal axis. The observation that testosterone exerts a stronger restraint on the secretory response to stress is, to certain extent, surprising, and cannot be readily explained by add-on effects of testosterone-derived estrogens. At present, we speculate that testosterone-mediated suppression may result from either altered secretory capacity of pituitary corticotrophs, or changes in the responsiveness/function of mechanisms involved in the perception and assessment of aversive stimuli. Taken together, the results of this comparative examination of pure AR-agonists (DHT), aromatizable androgens (testosterone) and androgenic-anabolic steroids (anadrol) are indicative of differential neurotropic profiles and, consecutively, applicability to defined neurological symptoms (e.g. sexual dysfunction, anxiety or inadequate responsiveness to emotional stress). It should be mentioned that the neural circuits presumably involved in the generation/maintenance of these symptoms display different degrees of functional specialization; according to recent observations, sex steroid actions in narrowly specialized neuronal populations may experience amplification through steroid receptor co-regulators which with distinct regional presence and operational efficacy (Mitev et al., 2003). However, the search of brain-specific androgen-regulated genes remains a key challenge and, probably, the most appropriate approach for the precise description of the role of androgen signaling in brain physiology and pathology.

2. Differential transcriptome analysis of the aging and androgen-treated rat brain The term transcriptome describes the pool of mRNAs in a given cell or organ. Comparative transcriptome analysis of tissues derived from organisms exposed to physiological or pathological stimuli enables qualitative and quantitative characterization of changes in the mRNA pool. Thus, this technology provides opportunities for the identification

Table 1 Age-associated changes in gene transcripts in the brain of young (2-monthold) and aged (24-month-old) rats and their probable implication in CNS function BLAST homologue

Fold change old vs. young

Probable relevance

Myelin proteolipid protein Myelin basic protein Casein kinase 1a

9.4Y

Major myelin component

3.1Y 2.7Y

Major myelin component Tau and presenilin phosphorylation Cholesterol uptake; steroid synthesis

Sterol regulatory element binding protein 2 High affinity IgE receptor

4.5Y

RAS-related GTP binding protein Inositol triphosphate receptor

5.7Y

3.8[

7.6Y

Linkage between pathologyrelated IgE and cellular response Regulated by growth factors and synaptic activity Coupling of stimuli to Ca2C signaling

of genes whose transcription is specifically affected by the condition or treatment of interest. In the following study we compared the transcriptome of brains from young, sexually mature (2-month-old) and aged gonad-intact male rats (24-month-old) with the aim to reveal differences in the presence of transcripts which encode proteins with probable implication in CNS function and pathology. Using differential display of RT-PCR products followed by sequencing and confirmation by Northern blot analysis, nucleotides of interest were tested for homology to known genomic entities by BLAST search. This procedure ultimately revealed 21 genes whose expression was decreased in the aging brain, whereas 7 were up-regulated in association with aging (Schroeder and Patchev, 2000). Table 1 illustrates a selection of differentially expressed genes with high probability of involvement in CNS function. Even a superficial analysis of these results suggests that aging-related changes in the rat brain transcriptome can be linked to the pathogenesis of numerous medical conditions, from impaired signal processing and cognitive decline to neurodegeneration and reduced repair capacity (Breteler et al., 1994; Sandell and Peters, 2003; Ong et al., 2000; O’Kane et al., 2003; Kuret et al., 1997; Berridge, 1998). However, while age-associated changes in gene transcription are neither uncommon, nor surprising, it was particularly intriguing in the context of this study to examine the responsiveness of the above-listed target transcripts to androgen treatment. In these experiments, microarrays were hybridized with RNA isolated from the cortex, hippocampus and striatum of gonad-intact, castrated and testosterone- or DHT-treated male rats and interrogated for treatment-induced changes in the expression of the previously selected genes with significant aging liability. Among the transcripts examined, only those coding for myelin basic protein and myelin proteolipid protein

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Fig. 5. Changes in the presence of transcripts encoding myelin basic protein and myelin proteolipid protein in the cerebral cortex and hippocampus of rats following gonadectomy (GDX) and s.c. supplementation with testosterone (T) at daily doses of 0.3 and 3 mg for 7 days. Data are presented as arbitrary units derived from fluorescence read-outs of Affymetrix microarrays; for each treatment group and brain area, pooled RNA samples from five animals were used for hybridization.

displayed measurable changes following alterations in the gonadal endocrine status (Fig. 5). Pronounced changes were documented to the cerebral cortex and, to a lesser extent, hippocampus, whereas gonadectomy and androgen treatment failed to influence the transcription of genes of interest in the striatum (data not shown). However, there were regional differences in transcript responsiveness to endocrine alterations. In the cerebral cortex, the expression of both myelin-associated proteins was decreased by sex steroid deprivation and dose-dependently reconstituted by testosterone administration, whereas in the hippocampus, transcription of myelin proteolipid protein was up-regulated by gonadectomy and returned to control levels upon androgen treatment. Furthermore, individual androgens differ in their ability to normalize gonadectomy-induced changes in gene transcription: unlike testosterone, DHT failed to demonstrate measurable effects on the transcription of genes of interest in any of the brain structures examined. Although meticulous scrutiny of these data at cellular and subcellular level is required to gain insight into mechanistic principles of androgen action, certain preliminary inferences can be drawn from the present observations. Compelling evidence exists for aberrant and insufficient remyelination with increasing age, and this phenomenon has been implicated in the pathogenesis and progression of neurological and mental disorders (Sim et al., 2002; Benes, 2004; Wang et al., 2004). The issue of the capacity of sex steroids to counteract myelin damage has been extensively addressed in the past (Ibanez et al., 2003), and convincing

therapeutic effects were demonstrated in models of peripheral neuropathy (Melcangi et al., 2000, 2001, 2003). However, in most of the above-mentioned studies, effects of androgens were rather modest and ascribed to DHT, but not testosterone (Magnaghi et al., 1999). On the basis of those observations, it was postulated that 5a-reduction seems to be a necessary pre-requisite for the neurotropic effects of sex steroids on peripheral nerve myelin formation and/or biochemical composition (Azcoitia et al., 2003). Hence, progesterone and its ring-A-reduced derivatives seem to be major mediators of sex steroid control of myelin formation, with 5a-reduced androgens serving either as ancillary substrate pool or entering promiscuous interactions with progesterone-mediated signaling (Melcangi et al., 2001). Although it remains to be clarified whether these hypotheses may apply to myelin biosynthesis in the CNS, our data neither lend support for a distinct role of 5a-reduced androgens in this process. Our treatment paradigm and genomic endpoints may reveal changes resulting from persistently altered transcriptional regulation. Hence, we favor the interpretation that activation of the AR is not the principal mediator of testosterone action on gene transcription of myelin-associated proteins. In view of accumulating evidence for oligodendrocytes as major targets of neurotropic actions of estrogens (Zhang et al., 2004; Takao et al., 2004; Arvanitis et al., 2004), it appears plausible that aromatization of androgens in situ and subsequent activation of ER in this cell population may represent the pathway through which sex steroid influence myelin biosynthesis in the male brain.

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This overview reveals that neurotropic actions of androgens are still less well understood which precludes elaboration of guidelines for their therapeutic use in neurological and psychiatric conditions. Furthermore, whether androgens will become first-line therapeutic agents for defined nosological entities remains to be resolved. On the other hand, there is little doubt that androgens can evolve as useful supplements to CNS therapeutics, by priming and enhancing the effects of present day therapy. To achieve this outcome would require comprehensive knowledge of the neuropharmacological profile of individual androgen classes. As experimental and clinical data on specific responsiveness of neural targets and processes to androgens accumulates, research protocols designed beyond traditional disease models can accelerate the potential therapeutic use of androgens in CNS disorders.

Acknowledgements The contribution of Sven Ring (Medicinal Chemistry, Schering/Jenapharm) and the expert assistance of Anja Fischbach (Institute of Experimental Endocrinology, School of Medicine Charite´, Berlin) and Simone Thalheim (Schering/Jenapharm) is gratefully acknowledged.

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