phytoestrogens and 2-3-bis(4-hydroxyphenyl)-propionitrile

The organization of the developing male rodent brain is profoundly influenced by endogenous steroids, most notably estrogen. This process may be disrupted by estrogenic endocrine disrupting compounds (EDCs) resulting in altered sex behavior and the capacity to attract a mate in adulthood. To better understand the relative role each estrogen receptor (ER) subtype (ERα and ERβ) plays in mediating these effects, we exposed male Long Evans rats to estradiol benzoate (EB, 10 μg), vehicle, or agonists specific for ERβ (DPN, 1 mg/kg) or ERα (PPT, 1 mg/kg) daily for the first four days of life, and then assessed adult male reproductive behavior and attractiveness via a partner preference paradigm. DPN had a greater adverse impact than PPT on reproductive behavior, suggesting a functional role for ERβ in the organization of these male-specific behaviors. Therefore the impact of neonatal ERβ agonism was further investigated by repeating the experiment using vehicle, EB and additional DPN doses (0.5 mg/kg, 1 mg/kg, and 2 mg/kg bw). Exposure to DPN suppressed male reproductive behavior and attractiveness in a dose dependent manner. Finally, males were exposed to EB or an environmentally relevant dose of genistein (GEN, 10 mg/kg), a naturally occurring xenoestrogen, which has a higher relative binding affinity for ERβ than ERα. Sexual performance was impaired by GEN but not attractiveness. In addition to suppressing reproductive behavior and attractiveness, EB exposure significantly lowered the testis to body weight ratio, and circulating testosterone levels. DPN and GEN exposure only impaired behavior, suggesting that disrupted androgen secretion does not underlie the impairment.

Topics: Animals; Dose-Response Relationship, Drug; Estradiol; Estrogen Receptor alpha; Estrogen Receptor beta; Estrogens; Genistein; Male; Mating Preference, Animal; Nitriles; Phenols; Phytoestrogens; Propionates; Pyrazoles; Rats; Rats, Long-Evans; Sexual Behavior, Animal

We have scrutinized the effects of the phytoestrogen genistein and three synthetic estrogen receptor agonists, 17 alpha-ethynylestradiol (EE), propylpyrazole-triol (PPT) and diarylpropionitrile (DPN) in the completely estrogen-free background of aromatase knockout (ArKO) mice by means of two routes of substance administration: oral via diet (per os; po) or subcutaneous injection (sc) with the intention to evaluate the ArKO mice as sensitive model organism for uterotrophic assays. Additionally, we were aiming to qualitatively analyze effects resulting from oral administration path, in particular for PPT and DPN. Therefore, we analyzed the resulting uterine wet weights (UWW) and epithelial heights as physiological endpoints of function as well as the gonadotropin levels. Moreover, the gene expression profiles of estrogen receptors as well as important uterine and ovarian estrogen-response genes were investigated by real-time PCR. The uterus of ArKO mice responded very sensitive upon the substitution with EE (sc 5 microg/kg BW; po 50 microg/kg BW) in a proliferative manner. This was evaluated inter alia by increased UWW and by up-regulation of the expression of proliferation-associated and estrogen-response genes. It is important to note, that ER alpha and ER beta-agonist, PPT and DPN respectively (po 5mg/kg BW and sc 0.5mg/kg BW), have only been used for sc applications so far. Here, effects resulting from oral application were qualitatively described and evaluated for their applicability. The UWW and expression of proliferation-associated genes were increased following both po and sc treatment with PPT. In contrast, DPN did not exert an increase of the UWW, but a significant decrease of proliferation-associated gene and protein expression. Additionally, a substantial hypoplasia was detectable in the uterine cross-sections of DPN-treated mice. On the other hand, the phytoestrogen genistein (sc 10mg/kg BW; po 70 mg/kg BW) did not cause detectable uterotrophic responses or large changes of uterine and ovarian gene expression profiles under the applied experimental conditions, but significantly reduced the elevated gonadotropin levels of ArKO mice. In summary, we showed the utility of ArKO mice to detect ER-specific effects, in particular those of PPT and DPN also when applied orally.

Topics: Animals; Aromatase; Estrogen Receptor alpha; Estrogen Receptor beta; Ethinyl Estradiol; Female; Follicle Stimulating Hormone; Gene Expression Profiling; Genistein; Luteinizing Hormone; Mice; Mice, Inbred C57BL; Mice, Knockout; Nitriles; Ovary; Phenols; Phytoestrogens; Proliferating Cell Nuclear Antigen; Propionates; Pyrazoles; Receptors, LH; Uterus

The development of highly selective agonists for the two major subforms of the estrogen receptor (ERalpha and ERbeta) has produced new experimental methodologies for delineating the distinct functional role each plays in neurobehavioral biology. It has also been suggested that these compounds might have the potential to treat estrogen influenced behavioral disorders, such as anxiety and depression. Prior work has established that the ERbeta agonist, diarylpropionitrile (DPN) is anxiolytic in gonadectomized animals of both sexes, but whether or not this effect persists in gonadally intact individuals is unknown. Isoflavone phytoestrogens, also potent but less selective ERbeta agonists, have also been shown to influence anxiety in multiple species and are becoming more readily available to humans as health supplements. Here we determined the effects of 0.5, 1 or 2 mg/kg DPN, 1 mg/kg of the ERalpha agonist propyl-pyrazole-triol (PPT), 3 or 20 mg/kg of the isoflavone equol (EQ) and 3 or 20 mg/kg of the isoflavone polyphenol resveratrol (RES) on anxiety behavior in the gonadally intact male rat using the light/dark box and the elevated plus maze. We first determined that DPN can be successfully administered either orally or by subcutaneous injection, although plasma DPN levels are significantly lower if given orally. Once injected, plasma levels peak rapidly and then decline to baseline levels within 3 h of administration. For the behavioral studies, all compounds were injected and the animals were tested within 3 h of treatment. None of the compounds, at any of the doses, significantly altered anxiety-related behavior. Plasma testosterone levels were also not significantly altered suggesting that these compounds do not interfere with endogenous androgen levels. The results suggest that the efficacy of ERbeta agonists may depend on gonadal status. Therefore the therapeutic potential of ERbeta selective agonists to treat mood disorders may be limited.

Topics: Analysis of Variance; Animals; Anti-Anxiety Agents; Anxiety; Behavior, Animal; Equol; Estrogen Receptor alpha; Isoflavones; Male; Nitriles; Phenols; Phytoestrogens; Propionates; Pyrazoles; Rats; Rats, Long-Evans; Rats, Wistar; Resveratrol; Stilbenes; Testosterone