Developmental and physiological effects of estrogen receptor gene disruption in mice.

Disruption of the estrogen receptor (ER) gene in mice causes infertility in both sexes. Infertility in female ER knockout (ERKO) mice results from altered development of accessory sex structures, disrupted endocrine physiology, and disrupted gametogenesis. Male accessory sex structures appear relatively normal, with infertility stemming from altered sexual behaviors and disrupted gametogenesis. These findings provide significant insights into the biological importance of the ER and suggest further areas for examining the impact of estrogens on reproductive biology.

The mouse androgen receptor is suppressed by the 5'-untranslated region of the gene.

The androgen receptor (AR) mediates the biological functions of androgens and is essential for normal growth and differentiation of urogenital organs as well as initiation and maintenance of spermatogenesis. Withdrawal of androgens by castration or other methods has been shown to cause a marked, although often temporary, regression of many prostate cancers. In order to gain a better understanding of the transcriptional regulation of the AR, a series of truncation mutants derived from the 5'-region of the mouse AR (mAR) were inserted into the promoter-less plasmid pBLCAT3 and transiently expressed in the mouse alpha T3-1 and GT1-7 cell lines. The results of these experiments indicate the presence of a negative regulatory element in the 5'-untranslated region of the gene, which is able to reduce chloramphenicol acetyltransferase (CAT) activity by 77-89%. We have named this element the mAR suppressor (mARS). DNase-I protection assays of the 5'-untranslated region disclosed a protected domain. Gel mobility assays using the mARS revealed the presence of three protein-DNA complexes that could specifically bind to this protected domain. Insertion of the mARS into the thymidine kinase promoter containing pBLCAT2 vector resulted in a 2- to 10-fold decrease in CAT activity, but only if the insert was 3' to the start of transcription initiation. Finally, point mutations within the mARS were able to increase transcription of the AR promoter by 2.3-fold. The results of these experiments indicate that the mAR 5'-untranslated region contains a suppressor element.(ABSTRACT TRUNCATED AT 250 WORDS)

Regulation of the 5'-flanking region of the mouse androgen receptor gene by cAMP and androgen.

Both androgens and cAMP-mediated hormones are known to regulate expression of the androgen receptor (AR) gene. In order to determine whether these effects occur at the transcriptional level, transfection studies were conducted with a 1.5-kilobase fragment of the 5'-flanking region of the mouse AR gene coupled to a chloramphenicol acetyltransferase (CAT) reporter gene. We demonstrated that the cAMP pathway regulates expression of the mouse AR (mAR) 5'-CAT construct in mouse pituitary (alpha T3-1), rat pituitary (GC), and quail fibroblast (QT6) cell lines. Deletional analysis indicated that several areas of this clone, including a region containing a putative cAMP response element (CRE), are involved in mediating cAMP regulation of the AR gene. Oligonucleotides containing a putative CRE, linked to the thymidine kinase promoter of pBLCAT2, conferred increased forskolin induction of CAT activity. Furthermore, overexpression of a CRE binding protein up-regulated expression of the mAR 5'-CAT constructs. Bandshift data demonstrated that the putative CRE forms specific, competable complexes with nuclear proteins from alpha T3-1 and QT6 cells. Additional experiments indicated that AR can modulate both basal and forskolin-induced CAT activity in an androgen-dependent fashion. These data provide evidence that the 5'-flanking region of the mAR gene contains sequences which mediate the effects of both cAMP and androgens.

Role of estrogen receptor-alpha in the anterior pituitary gland.

Targeted insertional disruption of the mouse estrogen receptor-alpha (ER alpha) gene has provided a genetic model in which to test hypotheses that estrogens exert important effects in development and homeostatic functions of the anterior pituitary gland, particularly in the lactotroph and gonadotroph cell types. Analysis of ER alpha gene-disrupted mice reveals a marked reduction in PRL mRNA and a decrease in lactotroph cell number, but normal specification of lactotroph cell phenotype. Gonadotropin mRNA levels in ER alpha gene-disrupted female mice are elevated, consistent with previously described transcriptional suppression of gonadotropin subunit gene expression in response to sustained administration of estrogen in wild type mice. These results provide genetic evidence that ER alpha plays a critical role in PRL and gonadotropin gene transcription and is involved in lactotroph cell growth, but is not required for specification of lactotroph cell phenotype.

Analysis of transcription and estrogen insensitivity in the female mouse after targeted disruption of the estrogen receptor gene.

We employed homologous recombination in mouse embryonic stem cells to disrupt the estrogen receptor (ER) gene. Subsequently generated mice that are homozygous for the gene disruption, termed ERKO, possess no demonstrable wild-type ER by Western blot analysis. However, the presence of residual high affinity binding, as detected by [3H]estradiol binding assays and sucrose gradients in uterine extracts from ERKO females prompted further investigation of transcription and translation products from the disrupted ER gene. Analysis of ERKO uterine messenger RNA (mRNA) by reverse transcriptase-polymerase chain reaction demonstrated that although no full-length wild-type ER mRNA was present, two smaller transcripts, labeled E1 and E2, were identified and partially sequenced. Both ERKO transcripts are splicing variants that result in the disrupting NEO sequence being partially or completely removed from the mRNA. In the ERKO-E2 variant, this results in a frame shift and the creation of at least two stop codons downstream. In the ERKO-E1 variant, the ER reading frame is preserved and encodes for a smaller mutant ER that could be the source of the residual estradiol binding. When this mutant form is overexpressed and characterized in vitro, it results in a smaller protein of the predicted size that possesses significantly reduced estrogen-dependent transcriptional activity compared with that of the wild-type ER. Despite residual amounts of an impaired ER variant, estrogen insensitivity in the female ERKOs was confirmed by the failure of estrogen treatment to induce known uterine markers of estrogen action, such as increased DNA synthesis, and transcription of the progesterone receptor, lactoferrin, and glucose-6-phosphate dehydrogenase genes. Furthermore, serum levels of estradiol in the ERKO female are more than 10-fold higher than those in the wild type, consistent with a syndrome of hormone insensitivity.

Calcium regulation of androgen receptor expression in the human prostate cancer cell line LNCaP.

Elevation of intracellular calcium levels in the presence of normal androgen levels has been implicated in apoptotic prostate cell death. Since the androgen receptor (AR) plays a critical role in the regulation of growth and differentiation of the prostate, it was of interest to determine whether Ca2+ would affect the expression of androgen receptor messenger RNA (mRNA) and protein, thus affecting the ability of androgens to control prostate function. AR-positive human prostate cancer cells, LNCaP, were incubated with either the calcium ionophore A23187 or the intracellular endoplasmic reticulum Ca(2+)-ATPase inhibitor thapsigargin. Subsequently, AR mRNA and protein levels were assessed by Northern and Western blot analysis. Both A23187 and thapsigargin were found to down-regulate steady state AR mRNA levels in a time- and dose-dependent manner. AR mRNA began to decrease after 6-8 h of incubation with 10(-6) M A23187 or 10(-7) M thapsigargin, reaching a nadir at 16 and 10 h of incubation, respectively. In contrast, control mRNA (glyceraldehyde 3-phosphate dehydrogenase) did not change significantly during the treatments with either A23187 or thapsigargin. AR protein levels were found to be decreased after 12 h of incubation with either 10(-6) M A23187 or 10(-7) M thapsigargin. The decrease in AR mRNA and protein seemed to precede apoptosis, since neither A23187 (24 h) nor thapsigargin (30 h) was found to alter cell morphology within the treatment time. Cycloheximide and actinomycin D were unable to change the calcium-mediated decrease in AR mRNA, ruling out the necessity for de novo protein synthesis or a change in mRNA stability. Moreover, the decrease in AR mRNA induced by calcium does not seem to involve protein kinase C- or calmodulin-dependent pathways, since inhibitors of these cellular components had no effect. Nuclear run-on assays demonstrated little or no effects of either A23187 or thapsigargin treatment on AR gene transcription (8 h and 10 h). In conclusion, these studies show that intracellular calcium seems to be a potent regulator of AR gene expression in LNCaP cells.

Tissue distribution and quantitative analysis of estrogen receptor-alpha (ERalpha) and estrogen receptor-beta (ERbeta) messenger ribonucleic acid in the wild-type and ERalpha-knockout mouse.

Until recently, only a single type of estrogen receptor (ER) was thought to exist and mediate the genomic effects of the hormone 17beta-estradiol in mammalian tissues. However, the cloning of a gene encoding a second type of ER, termed ERbeta, from the mouse, rat, and human has prompted a reevaluation of the estrogen signaling system. Based on in vitro studies, the ERbeta protein binds estradiol with an affinity similar to that of the classical ER (now referred to as ERalpha) and is able to mediate the effects of estradiol in transfected mammalian cell lines. Essential to further investigations of the possible physiological roles of ERbeta, and its possible interactions with ERalpha, are data on the tissue distribution of the two ER types. Herein, we have described the optimization and use of an RNase protection assay able to detect and distinguish messenger RNA (mRNA) transcripts from both the ERalpha and ERbeta genes in the mouse. Because this assay is directly quantitative, a comparison of the levels of expression within various tissues was possible. In addition, the effect of disruption of the ERalpha gene on the expression of the ERbeta gene was also investigated using the ERalpha-knockout (ERKO) mouse. Transcripts encoding ERalpha were detected in all the wild-type tissues assayed from both sexes. In the female reproductive tract, the highest expression of ERbeta mRNA was observed in the ovary and showed great variation among individual animals; detectable levels were observed in the uterus and oviduct, whereas mammary tissue was negative. In the male reproductive tract, significant expression of ERbeta was seen in the prostate and epididymis, whereas the testes were negative. In other tissues of both sexes, the hypothalamus and lung were clearly positive for both ERalpha and ERbeta mRNA. The ERKO mice demonstrated slightly reduced levels of ERbeta mRNA in the ovary, prostate, and epididymis. These data, in combination with the several described phenotypes in both sexes of the ERKO mouse, suggest that the biological functions of the ERbeta protein may be dependent on the presence of ERalpha in certain cell types and tissues. Further characterization of the physiological phenotypes in the ERKO mice may elucidate possible ERbeta specific actions.

Effects of castration and chronic steroid treatments on hypothalamic gonadotropin-releasing hormone content and pituitary gonadotropins in male wild-type and estrogen receptor-alpha knockout mice.

Testicular androgens are integral components of the hormonal feedback loops that regulate circulating levels of LHbeta and FSH. The sites of feedback include hypothalamic areas regulating GnRH neurons and pituitary gonadotropes. To better define the roles of androgen receptor (AR), estrogen receptor-alpha (ERalpha), and estrogen receptor-beta (ERbeta) in mediating feedback effects of sex steroids on reproductive neuroendocrine function, we have determined the effects of castration and steroid replacement therapy on hypothalamic GnRH content, pituitary LHbeta and FSHbeta messenger RNA (mRNA) levels, and serum gonadotropins in male wild-type (WT) and estrogen receptor-alpha knockout (ERKO) mice. Hypothalami from intact WT and ERKO males contained similar amounts of GnRH, whereas castration significantly reduced GnRH contents in both genotypes. Replacement therapy with estradiol (E2), testosterone (T), or dihydrotestosterone (DHT) restored hypothalamic GnRH content in castrated (CAST) WT mice; only the androgens were effective in CAST ERKOs. Analyses of pituitary function revealed that LHbeta mRNA and serum LHbeta levels in intact ERKOs were 2-fold higher than those in intact WT males. Castration increased levels of LHbeta mRNA (1.5- to 2-fold) and serum LHbeta (4- to 5-fold) in both genotypes. Both E2 and T treatments significantly suppressed LHbeta mRNA and serum LH levels in CAST WT males. However, E2 was completely ineffective, and T was only partially effective in suppressing these two indexes in the CAST ERKO males. DHT treatments stimulated a 50% increase in LHbeta mRNA and serum LH levels in WT males, whereas serum LH was significantly suppressed in DHT-treated ERKO males. Although the pituitaries from intact ERKO males contained similar amounts of FSHbeta mRNA, serum FSH levels were 20% higher than those in the intact WT males. Castration increased FSHbeta mRNA levels only in WT males, but significantly increased serum FSH levels in both genotypes. Both E2 and T treatments significantly suppressed serum FSH in CAST WT males, whereas only E2 suppressed FSHbeta mRNA. DHT treatments of CAST WT mice stimulated a small increase in serum FSH, but failed to alter FSHbeta mRNA levels. None of the steroid treatments exerted any significant effect on FSHbeta mRNA or serum FSH levels in CAST ERKOs. These data suggest that hypothalamic GnRH contents can be maintained solely through AR signaling pathways. However, normal regulation of gonadotrope function requires aromatization of T and activation of ERalpha signaling pathways in the gonadotrope. In addition, serum FSH levels in male ERKOs appear to be regulated largely by nonsteroidal testicular factors such as inhibin. Finally, these data suggest that hypothalamic ERbeta may not be involved in mediating the negative feedback effects of T on serum LH and FSH in male mice.

Prevention of the polycystic ovarian phenotype and characterization of ovulatory capacity in the estrogen receptor-alpha knockout mouse.

Ovarian-derived estradiol plays a critical endocrine role in the regulation of gonadotropin synthesis and secretion from the hypothalamic-pituitary axis. In turn, several para/autocrine effects of estrogen within the ovary are known, including increased ovarian weight, stimulation of granulosa cell growth, augmentation of FSH action, and attenuation of apoptosis. The estrogen receptor-alpha (ERalpha) is present in all three components of the hypothalamic-pituitary-ovarian axis of the mouse. In contrast, estrogen receptor-beta (ERbeta) is easily detectable in ovarian granulosa cells but is low to absent in the pituitary of the adult mouse. This distinct expression pattern for the two ERs suggests the presence of separate roles for each in the regulation of ovarian function. Herein, we definitively show that a lack of ERalpha in the hypothalamic-pituitary axis of the ERalpha-knockout (alphaERKO) mouse results in chronic elevation of serum LH and is the primary cause of the ovarian phenotype of polycystic follicles and anovulation. Prolonged treatment with a GnRH antagonist reduced serum LH levels and prevented the alphaERKO cystic ovarian phenotype. To investigate a direct role for ERalpha within the ovary, immature alphaERKO females were stimulated to ovulate with exogenous gonadotropins. Ovulatory capacity in the immature alphaERKO female was reduced compared with age-matched wild-type (14.5+/-2.9 vs. 40.6+/-2.6 oocytes/animal, respectively); however, oocytes collected from the alphaERKO were able to undergo successful in vitro fertilization. A similar discrepancy in oocyte yield was observed after superovulation of peripubertal (42 days) wild-type and alphaERKO females. In addition, ovaries from immature superovulated alphaERKO females possessed several ovulatory but unruptured follicles. Investigations of the possible reasons for the reduced number of ovulations in the alphaERKO included ribonuclease protection assays to assess the mRNA levels of several markers of follicular maturation and ovulation, including ERbeta, LH-receptor, cyclin-D2, P450-side chain cleavage enzyme, prostaglandin synthase-2, and progesterone receptor. No marked differences in the expression pattern for these mRNAs during the superovulation regimen were observed in the immature alphaERKO ovary compared with that of the wild-type. Serum progesterone levels just before ovulation were slightly lower in the alphaERKO compared with wild-type. These studies indicate that treatment of alphaERKO females with a GnRH antagonist decreased the serum LH levels to within the wild-type range and concurrently prevented development of the characteristic ovarian phenotype of cystic and hemorrhagic follicles. Furthermore, a lack of functional ERalpha within the ovary had no effect on the regulation of several genes required for follicular maturation and ovulation. However, the reduced numbers of ovulations following the administration of exogenous gonadotropins in the alphaERKO suggests an intraovarian role for ERalpha in follicular development and ovulation.

Induction of mammary gland development in estrogen receptor-alpha knockout mice.

Mammary glands from the estrogen receptor-a knockout (alphaERKO) mouse do not undergo ductal morphogenesis or alveolar development. Disrupted ERalpha signaling may result in reduced estrogen-responsive gene products in the mammary gland or reduced mammotropic hormones that contribute to the alphaERKO mammary phenotype. We report that circulating PRL is reduced in the female alphaERKO mouse. Implantation of an age-matched, heterozygous ERalpha pituitary isograft under the renal capsule of 25-day-old or 12-week-old alphaERKO mice increased circulating PRL and progesterone levels, and induced mammary gland development. Grafted alphaERKO mice also possessed hypertrophied corpora lutea demonstrating that PRL is luteotropic in the alphaERKO ovary. By contrast, ovariectomy at the time of pituitary grafting prevented mammary gland development in alphaERKO mice despite elevated PRL levels. Hormone replacement using pellet implants demonstrated that pharmacological doses of estradiol induced limited mammary ductal elongation, and estradiol in combination with progesterone stimulated lobuloalveolar development. PRL alone or in combination with progesterone or estradiol did not induce alphaERKO mammary growth. Estradiol and progesterone are required for the structural development of the alphaERKO mammary gland, and PRL contributes to this development by inducing ovarian progesterone levels. Therefore, the manifestation of the alphaERKO mammary phenotype appears due to the lack of direct estrogen action at the mammary gland and an indirect contributory role of estrogen signaling at the hypothalamic/pituitary axis.

LY191704 inhibits type I steroid 5 alpha-reductase in human scalp.

Conversion of testosterone to dihydrotestosterone (DHT) has been demonstrated to be catalyzed by two isoforms of steroid 5 alpha-reductase, designated types I and II. Although several classes of steroid-based inhibitors of the type II isoform have been identified, these agents have not demonstrated highly selective pharmacological activity against human type I 5 alpha-reductase. LY191704 is representative of a series of nonsteroidal agents that have potent [apparent inhibitory constant (Ki) = 11.3 nM] inhibitory activity in human scalp skin homogenates (pH 7.5), a source of type I 5 alpha-reductase. [3H]-DHT production in the presence and absence of LY191704 is consistent with a noncompetitive mode of inhibition. In human prostatic homogenates (pH 5.5), a source of type II 5 alpha-reductase, LY191704 is virtually inactive as an inhibitor [concentration of inhibitor producing 50% inhibition of enzymatic activity (IC50) > 1,000 nM] of [3H]-DHT formation. LY191704 does not inhibit the type I or type II isoforms of rat 5 alpha-reductase, nor does the compound compete for binding to the murine androgen receptor expressed in SF9 cells using a baculo virus expression system. The benzoquinolinones, as exemplified by LY191704, possess exquisite pharmacological selectivity and provide a tool to understand the role of human type I 5 alpha-reductase in normal and pathophysiological states. These agents may also find clinical utility in treating androgen-dependent dermatological conditions.

Effects of progestins on sexual behaviour in castrated lizards (Cnemidophorus inornatus).

In many male vertebrates, androgens stimulate, while progesterone inhibits, sexual behaviour. Testicular androgens also control sexual behaviour in males of the lizard Cnemidophorus inornatus. However, administration of progesterone will reinstate sexual behaviour in castrated animals. Increased doses of progesterone did not result in increased numbers of responders; at all doses tested approximately 36% (n = 45) of animals treated with progesterone responded as reliable courters. A high percentage (60-68%) of progesterone-sensitive individuals maintained their sensitivity over a period of months. A synthetic progestin 17,21-dimethyl-19-nor-4,9-pregnadiene-3,20-dione with low rates of metabolism and conversion, also stimulated sexual behaviour in castrated C. inornatus. Finally, the synthetic antiprogestin 11 beta-4-dimethylaminophenyl-17 beta-hydroxy-17 alpha-1-propinyl- 4,9-oestradiene-3-one inhibited the stimulatory effects of progesterone on sexual behaviour but failed to inhibit the stimulatory effects of androgens. These findings indicate an unusual departure from the characteristic pattern of steroid sensitivities of neural centres controlling sexual behaviour.

Estrogen-dependent gene regulation by an oxidative metabolite of diethylstilbestrol, diethylstilbestrol-4',4"-quinone.

Diethylstilbestrol (DES) is a well-characterized carcinogen in humans and animals although its mechanisms of carcinogenicity are not yet known. While the estrogenic activity of DES is important, there is evidence that oxidative metabolism also plays an important role for its toxicity. DES is oxidatively metabolized in vivo and in vitro to a number of compounds including diethylstilbestrol-4',4"-quinone (DQ), an unstable and reactive intermediate, and Z,Z-dienestrol (ZZ-DIEN). Estrogen receptor (ER) binding assays with mouse uterine cytosol indicate that DES, DQ and ZZ-DIEN have relative binding affinities of 286, 3.6 and 0.3, respectively, relative to estradiol as 100. In addition, DQ binds irreversibly and specifically to ER suggesting that DQ may be biologically active despite its rapid metabolism and lower binding affinity compared to DES. To test this, COS-1 cells were transfected with an estrogen responsive reporter construct containing of VitA2 estrogen response element (ERE) with or without an ER expression vector. In the presence of ER, treatments with DES, DQ and ZZ-DIEN resulted in 11, 10, and 2-fold induction of chloramphenicol acetyltransferase (CAT) activity, respectively. This induction was mediated by estrogen receptor since it was suppressed by pretreatment with a 10-fold excess of the pure antiestrogen ICI 182,780. These data indicate that DQ is a biologically active intermediate that is capable of transactivation of estrogen responsive genes through the ER. Furthermore, the data suggest that the ability of DQ to irreversibly bind ER may result in persistent stimulation of ER. This persistent stimulation may be related to the carcinogenicity of DES.

Interactions between progesterone and androgens in the stimulation of sex behaviors in male little striped whiptail lizards, Cnemidophorus inornatus.

Progesterone is believed to have a suppressive or inhibitory role in regulating androgen-dependent sex behaviors in male mammals and birds. Previous studies in this laboratory have revealed that in the little striped whiptail lizard (Cnemidophorus inornatus), progesterone (P) can stimulate sex behavior in a proportion of the males. The present study sought to determine (i) interactions between androgens and P in activating sex behaviors, and (ii) the overlap in behavioral sensitivities to androgens and P in male C. inornatus. With an increasing length of castration the behavioral sensitivity of males to exogenous P was reduced. However, priming of castrated males with subthreshold doses of exogenous dihydrotestosterone (DHT) greatly facilitated subsequent behavioral responses to exogenous P. Progesterone treatments of castrated males were more effective at reinstating sex behaviors in males that exhibited high-intensity sex behaviors prior to castration compared to males that exhibited low-intensity sex behaviors. Finally, exogenous DHT is more effective at reinstating sex behaviors in P-sensitive males than in P-insensitive males. These data are discussed in light of possible mechanisms underlying the unusual behavioral effects of P in the Cnemidophorus model system.

Psychobiology of sexual behavior in a whiptail lizard, Cnemidophorus inornatus.

This study investigated the effects of social environment on gonadal recrudescence and sexual behavior in male and female Little Striped Whiptail lizards (Cnemidophorus inornatus). The presence of sexually active males facilitates ovarian recrudescence in conspecific females. Similarly, the presence of reproductively active females facilitates testicular recrudescence in conspecific males. Males housed with females, however, had lower average circulating concentrations of testosterone and dihydrotestosterone, and higher average concentrations of corticosterone compared to intact males housed in isolation. In other studies, the presence of reproductively active females partially restored courtship behavior in castrated males compared to castrated males housed in isolation. Despite the stimulatory effects of females on castrates, exogenous androgens are required for complete restoration of all components of sexual behavior in male C. inornatus. Females are receptive to male courtship and copulatory behavior only during the vitellogenic stages; females in previtellogenic or postovulatory ovarian stages aggressively reject male courtship advances. These findings demonstrate reciprocal effects of sexual behaviors of males and females upon each other's reproductive behavior and physiology.

Individual variation in intensity of sexual behaviors in captive male Cnemidophorus inornatus.

The present studies investigated the source of individual variation in intensity of sexual behaviors in captive male whiptail lizards, Cnemidophorus inornatus. No correlation was found between an individual's circulating concentration of dihydrotestosterone, testosterone, or corticosterone at the time of capture or in the laboratory and their level of sexual behaviors observed in the laboratory. A large percentage of males that initially exhibited low intensity courtship remained low intensity courters, although some became more reliable courters following 6 months of acclimation to the laboratory. Similarly, following castration and androgen replacement, most low intensity courters continued to exhibit weak and infrequent sexual behaviors. The data suggest that individual variation in sexual behaviors exhibited by captive male C. inornatus is not due to (i) low circulating concentrations of androgens, (ii) elevated circulating concentrations of corticosterone, or (iii) different profiles of testicular steroidogenesis. Rather, the source of differences may lie in (i) an inability to respond to androgens, (ii) an inability to exhibit sexual behavior, or (iii) non-hormonal stress related to captivity.

Effects of progesterone and dihydrotestosterone on stimulation of androgen-dependent sex behavior, accessory sex structures, and in vitro binding characteristics of cytosolic androgen receptors in male whiptail lizards (Cnemidophorus inornatus).

Progestins often act as potent antiandrogens in male birds and mammals. Experiments with lizards find that progestins can both inhibit (when given in high dosages) or stimulate (when given in low dosages) male-typical sex behavior in gonadectomized individuals. This study shows that in the little striped whiptail lizard exogenous progesterone (P) facilitates androgen-dependent sex behaviors in males yet fails to stimulate seasonal activation of androgen-dependent accessory sex structures. Analysis of androgen receptors (AR) in brain and kidney cytosol of the little striped whiptail lizard reveals similarities with the AR of the mouse. The data indicate that despite the ability of P to mimic the actions of androgens in activating sex behaviors in males of this species, the characteristics of the AR are conserved with respect to other vertebrate species.

Hormonal control of courtship and copulatory behavior in male Cnemidophorus inornatus, a direct sexual ancestor of a unisexual, parthenogenetic lizard.

The hormonal control of courtship and copulatory behavior in male Cnemidophorus inornatus, one of the gonochoristic ancestral species of the all-female, parthenogenetic C. uniparens, was determined. Significantly fewer castrated males courted females than intact and sham-castrated control males. Silastic capsules containing dihydrotestosterone or testosterone reinstated courtship and copulatory behavior in a significant number of castrates. While significantly greater numbers of castrates treated with androgens resumed courtship, some of the castrates receiving progesterone implants also resumed intense courtship and copulatory behavior. Exogenous progesterone also maintained courtship behaviors in a significant number of intact males at a time when control males ceased to court. The implications of these findings for our understanding of the evolution of hormone-brain-behavior relationships and sex steroid hormone mechanisms of action are discussed.

Behavioral facilitation of reproduction in sexual and unisexual whiptail lizards.

All-female, parthenogenetic species afford a unique test of hypotheses regarding the nature and evolution of sexuality. Mating behavior accomplishes the transfer of gametes and stimulates the coordination of reproductive activity of the male and female. Cnemidophorus uniparens, a parthenogenetic species, is believed to have resulted from the hybridization of two extant gonochoristic species, Cnemidophorus inornatus and Cnemidophorus gularis. C. uniparens regularly and reliably perform behaviors identical in form to those performed during mating by male C. inornatus. We have determined experimentally that individuals of the parthenogenetic species demonstrating male-like pseudosexual behavior also share a similarity in function with males of the sexually reproducing species. The number of female C. inornatus ovulating increases, and the latency to ovulation decreases, if a sexually active conspecific male is present. A similar facilitatory effect on ovarian recrudescence occurs in the all-female C. uniparens in the presence of a male-like individual. These results show that behavioral facilitation of ovarian recrudescence is important in sexual and unisexual species. This may represent a potent selection pressure favoring the maintenance of male-typical behaviors, thus accounting for the display of behavioral traits usually associated with males in unisexual species of hybrid origin.