Estrogen receptor alpha (ERα) mediates estrogen (E 2 ) actions in the brain and is critical for normal reproductive function and behavior. In the classical pathway, ERα binds to estrogen response elements (EREs) to regulate gene transcription. ERα can also participate in several non-classical pathways, including ERE-independent gene transcription via protein-protein interactions with transcription factors and rapid, non-genotropic pathways. To distinguish between ERE-dependent and ERE-independent mechanisms of E 2 action in vivo, we have created ERα null mice that possess an ER knock-in mutation (E207A/G208A; "AA"), in which the mutant ERα cannot bind to DNA but retains activity in ERE-independent pathways (ERα −/AA mice). Understanding the molecular mechanisms of ERα action will be helpful in developing pharmacological therapies that differentiate between ERE-dependent and -independent processes. This review focuses on how the ERα −/AA model has contributed to our knowledge of ERα signaling mechanisms in estrogen regulation of the reproductive axis and sexual behavior. Keywordsestrogen receptor alpha; estrogen response element; non-classical signaling; negative feedback; sexual behaviorThe biological effects of estrogens are mediated through at least two distinct nuclear receptors, ERα and ERβ, which belong to the nuclear hormone receptor superfamily (Mangelsdorf et al., 1995). In the classical pathway of estrogen action, E 2 binds to ER, inducing conformational changes within the receptor that promote dimerization and interaction with coactivator and corepressor molecules. The ligand-receptor complex binds with high affinity to specific estrogen response elements (EREs) in the regulatory regions of target genes to either activate or repress gene expression (Glass, 1994;McKenna et al

Androgens exert significant organizational and activational effects on the nervous system and behavior. Despite the fact that female mammals generally produce low levels of androgens, relative to the male of the same species, increasing evidence suggests that androgens can exert profound effects on the normal physiology and behavior of females during fetal, neonatal, and adult stages of life. This review examines the effects of exposure to androgens at three stages of development--as an adult, during early postnatal life and as a fetus, on reproductive hormone secretions in female rats. We examine the effects of androgen exposure both as a model of neuroendocrine sexual differentiation and with respect to the role androgens play in the normal female. We then discuss the hypothesis that androgens may cause epigenetic modification of estrogen target genes in the brain. Finally we consider the clinical consequences of excess androgen exposure in women.

Estrogen receptor (ER)-alpha mediates estradiol (E(2)) actions in the male gonads and brain and is critical for normal male reproductive function. In the classical pathway, ERalpha binds to estrogen response elements (EREs) to regulate gene transcription. ERalpha can also regulate gene transcription independently of EREs via protein-protein interactions with transcription factors and additionally signal via rapid, nongenomic pathways originating at the cell membrane. This study assessed the degree to which ERE-independent ERalpha signaling can rescue the disrupted masculine sexual behaviors and elevated serum testosterone (T) levels that have been shown to result from ERalpha gene deletion. We utilized male ERalpha null mice that possess a ER knock-in mutation (E207A/G208A; AA), in which the mutant ERalpha is incapable of binding to DNA and can signal only through ERE-independent pathways (ERalpha(-/AA) mice). We found that sexual behavior, including mounting, is virtually absent in ERalpha(-/-) and ERalpha(-/AA) males, suggesting that ERE-independent signaling is insufficient to maintain any degree of normal sexual behavior in the absence of ERE binding. By contrast, ERE-independent signaling in the ERalpha(-/AA) mouse is sufficient to restore serum T levels to values observed in wild-type males. These data indicate that binding of ERs to EREs mediates most if not all of E(2)'s effects on male sexual behavior, whereas ERE-independent ERalpha signaling may mediate E(2)'s inhibitory effects on T production.

Nonclassical estrogen receptor ␣ (ER␣) signaling can mediate E2 negative feedback actions in the reproductive axis; however, downstream pathways conveying these effects remain unclear. These studies tested the hypothesis that p21-activated kinase 1 (PAK1), a serine/threonine kinase rapidly activated by E 2 in nonneural cells, functions as a downstream node for E 2 signaling pathways in cells of the preoptic area, and it may thereby mediate E 2 negative feedback effects. Treatment of ovariectomized (OVX) rats with estradiol benzoate (EB) caused rapid and transient induction of phosphorylated PAK1 immunoreactivity in the medial preoptic nucleus (MPN) but not the arcuate nucleus. To determine whether rapid induction of PAK phosphorylation by E 2 is mediated by nonclassical [estrogen response element (ERE)-independent] ER␣ signaling, we used female ER␣ null (ER␣ ؊/؊ ) mice possessing an ER knock-in mutation (E207A/G208A; AA), in which the mutant ER␣ is incapable of binding DNA and can signal only through membrane-initiated or ERE-independent genotropic pathways (ER␣ ؊/AA mice). After 1-h EB treatment, the number of pPAK1-immunoreactive cells in the MPN was increased in both wild-type (ER␣ ؉/؉ ) and ER␣ ؊/AA mice but was unchanged in ER␣ ؊/؊ mice. Serum luteinizing hormone (LH) was likewise suppressed within 1 h after EB treatment in ER␣ ؉/؉ and ER␣ ؊/AA but not ER␣ ؊/ ؊ mice. In OVX rats, 5-min intracerebroventricular infusion of a PAK inhibitor peptide but not control peptide blocked rapid EB suppression of LH secretion. Taken together, our findings implicate PAK1 activation subsequent to nonclassical ER␣ signaling as an important component of the negative feedback actions of E 2 in the brain.GnRH ͉ LH ͉ estrogen receptor ␣ O varian estradiol-17␤ (E 2 ) conveys negative feedback actions within the reproductive axis that include inhibition of gonadotropin-releasing hormone (GnRH) neurosecretion and suppression of gonadotrope responsiveness to GnRH stimulation. Both actions can be sustained by E 2 treatment regimens that maintain serum E 2 levels in low physiological ranges (1) and they can also be manifested rapidly, within minutes after E 2 injection (2). Studies of estrogen receptor ␣ (ER␣), ER␤, and ER␣/␤ null mutant mice have clearly implicated ER␣ as the isoform essential for E 2 negative feedback regulation in vivo (3, 4).Cell signaling pathways that transduce ER␣-mediated negative feedback are not well understood. In classical ER␣ signaling mechanisms, E 2 binds nuclear ERs and recruits coactivators to consensus palindromic estrogen response elements (EREs). Direct binding of ERs to EREs thereby mediates alterations in transcription of target genes. Nonclassical ER␣ signaling mechanisms operate independently of ER␣ binding directly to EREs and include protein-protein interactions with transcription factors, such as AP1, SP1, and NF-B (5), which in turn mediate transcriptional regulation at their cognate response elements. Nonclassical ER␣ signaling also includes membrane-associated receptor activation coupled...

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