Androgen receptors are required for full masculinization of nitric oxide synthase system in rat limbic-hypothalamic region

The neuronal nitric oxide synthase (nNOS) is involved in the control of male and female sexual behavior and its distribution in several regions of the limbic–hypothalamic system, as well as its coexistence with gonadal hormones' receptors, suggests that these hormones may play a significant role in controlling its expression. However, data illustrating the role of gonadal hormones in controlling the nNOS expression are, at present, contradictory, even if they strongly suggest an involvement of testosterone (T) in the regulation of nNOS. The action of T may be mediated through androgen (AR) or, after aromatization to estradiol (E₂), through estrogen receptors.To elucidate the role of AR on nNOS expression, we compared male and female rats with a non-functional mutation of AR (Tfm, testicular feminization mutation) to their control littermates. We investigated some hypothalamic and limbic nuclei involved in the control of sexual behavior [medial preoptic area (MPA), paraventricular (PVN), arcuate (ARC), ventromedial (VMH) and stria terminalis (BST) nuclei]. In BST (posterior subdivision), VMH (ventral subdivision), and MPA we detected a significant sexual dimorphism in control animals and a decrease of nNOS positive elements in Tfm males compared to their littermate. In addition, we observed a significant increase of nNOS positive elements in BST (posterior) of Tfm females. No significant changes were observed in the other nuclei. These data indicate that, contrary to current opinions, androgens, through the action of AR may have a relevant role in the organization and modulation of the nNOS hypothalamic system.     

Effects of testosterone on the development of neuromuscular systems and their target tissues involved in courtship and copulation in green anoles (Anolis carolinensis)

Male green anole lizards court females using a red throat fan (dewlap) and copulate by intromitting one of two penises (hemipenes). These structures begin sexually monomorphic, but by adulthood males have larger dewlaps, only males have hemipenes, and many of the neuromuscular components of both systems show male-biased dimorphisms. We hypothesized that testosterone (T), which increases in juvenile males but not females about a month after hatching, facilitates masculinization. To test this idea, on post-hatching day 30, gonadally intact females received either a blank or T implant, and males were either castrated or sham-castrated. At day 90, juveniles were euthanized and the length of the cartilage and cross-sectional areas of the muscle fibers and motoneurons required for dewlap extension were examined. We also measured the cross-sectional areas of the hemipenes and associated muscle fibers and motoneurons, and counted the motoneurons. T-treated females had longer cartilages and larger dewlap muscle fibers compared to those with blank implants. No effects on motoneurons were detected, and no females possessed hemipenes or associated musculature. In males, castration produced shorter dewlap cartilages and smaller hemipenes; other measures were not affected by treatment. These data indicate that components of the dewlap system differentiate relatively late in development, that T likely mediates the process, and that although components of the copulatory system are plastic in juvenile males, sexual differentiation of peripheral features is complete before day 30. The data also suggest that target structures (dewlap cartilage and hemipenes), compared to their neuromuscular effectors, are particularly sensitive to developmental T exposure.     

Sexually dimorphic expression of the sex chromosome-linked genes cntfa and pdlim3a in the medaka brain

In vertebrates, sex differences in the brain have been attributed to differences in gonadal hormone secretion; however, recent evidence in mammals and birds shows that sex chromosome-linked genes, independent of gonadal hormones, also mediate sex differences in the brain. In this study, we searched for genes that were differentially expressed between the sexes in the brain of a teleost fish, medaka (Oryzias latipes), and identified two sex chromosome genes with male-biased expression, cntfa (encoding ciliary neurotrophic factor a) and pdlim3a (encoding PDZ and LIM domain 3 a). These genes were found to be located 3–4 Mb from and on opposite sides of the Y chromosome-specific region containing the sex-determining gene (the medaka X and Y chromosomes are genetically identical, differing only in this region). The male-biased expression of both genes was evident prior to the onset of sexual maturity. Sex-reversed XY females, as well as wild-type XY males, had more pronounced expression of these genes than XX males and XX females, indicating that the Y allele confers higher expression than the X allele for both genes. In addition, their expression was affected to some extent by sex steroid hormones, thereby possibly serving as focal points of the crosstalk between the genetic and hormonal pathways underlying brain sex differences. Given that sex chromosomes of lower vertebrates, including teleost fish, have evolved independently in different genera or species, sex chromosome genes with sexually dimorphic expression in the brain may contribute to genus- or species-specific sex differences in a variety of traits.     

Intrasexual selection in Verreaux's sifaka (Propithecus verreauxi verreauxi)

Studies of sexual selection show that both female choice and male-male competition can influence the evolution and expression of male phenotypes. In this regard, it is important to determine the functional basis through which male traits influence variation in male reproductive success. In this study, we estimate the strength and type of sexual selection acting on adult males in a population of wild lemur, Verreaux's sifaka (Propithecus verreauxi verreauxi). The data used in this study were collected at Beza Mahafaly Special Reserve, southwest Madagascar. We conducted paternity analyses on 70 males in order to estimate the distribution of reproductive success in this population. Paternity data were combined with morphometric data in order to determine which morphological traits covary with male fitness. Five morphological traits were defined in this analysis: body size, canine size, torso shape, arm shape, and leg shape. We utilized phenotypic selection models in order to determine the strength and type of selection acting directly on each trait. Our results show that directional selection acts on leg shape (a trait that is functionally related to locomotor performance), stabilizing selection acts on body mass and torso shape, and negative correlational selection acts on body mass and leg shape. We draw from biomechanical and kinematic studies of sifaka locomotion to provide a functional context for how these traits influence male mating competition within an arboreal environment. Verreaux's sifaka and many other gregarious lemurs are sexually monomorphic in body mass and canine size, despite a high frequency and intensity of male-male aggressive competition. Our results provide some insight into this paradox: in our population, there is no directional selection acting on body mass or canine size in males. The total pattern of selection implicates that behaviors relating to locomotor performance are more important than behaviors relating to fighting ability during intrasexual contests.     

Sex-specific repression of dachshund is required for Drosophila sex comb development

The origin of new morphological structures requires the establishment of new genetic regulatory circuits to control their development, from initial specification to terminal differentiation. The upstream regulatory genes are usually the first to be identified, while the mechanisms that translate novel regulatory information into phenotypic diversity often remain obscure. In particular, elaborate sex-specific structures that have evolved in many animal lineages are inevitably controlled by sex-determining genes, but the genetic basis of sexually dimorphic cell differentiation is rarely understood. In this report, we examine the role of dachshund (dac), a gene with a deeply conserved function in sensory organ and appendage development, in the sex comb, a recently evolved male-specific structure found in some Drosophila species. We show that dac acts during metamorphosis to restrict sex comb development to the appropriate leg region. Localized repression of dac by the sex determination pathway is necessary for male-specific morphogenesis of sex comb bristles. This pupal function of dac is separate from its earlier role in leg patterning, and Dac at this stage is not dependent on the pupal expression of Distalless (Dll), the main regulator of dac during the larval period. Dll acts in the epithelial cells surrounding the sex comb during pupal development to promote sex comb rotation, a complex cellular process driven by coordinated cell rearrangement. Our results show that genes with well-conserved developmental functions can be re-used at later stages in development to regulate more recently evolved traits. This mode of gene co-option may be an important driver of evolutionary innovations.     

Abdominal-B is essential for proper sexually dimorphic development of the Drosophila gonad

Sexual dimorphism requires the integration of positional information in the embryo with the sex determination pathway. Homeotic genes are a major source of positional information responsible for patterning along the anterior-posterior axis in embryonic development, and are likely to play a critical role in sexual dimorphism. Here, we investigate the role of homeotic genes in the sexually dimorphic development of the gonad in Drosophila. We have found that Abdominal-B (ABD-B) is expressed in a sexually dimorphic manner in the embryonic gonad. Furthermore, Abd-B is necessary and sufficient for specification of a sexually dimorphic cell type, the male-specific somatic gonadal precursors (msSGPs). In Abd-B mutants, the msSGPs are not specified and male gonads now resemble female gonads with respect to these cells. Ectopic expression of Abd-B is sufficient to induce formation of extra msSGPs in additional segments of the embryo. Abd-B works together with abdominal-A to pattern the non-sexually dimorphic somatic gonad in both sexes, while Abd-B alone specifies the msSGPs. Our results indicate that Abd-B acts at multiple levels to regulate gonad development and that Abd-B class homeotic genes are conserved factors in establishing gonad sexual dimorphism in diverse species.     

Notch and Numb are required for normal migration of peripheral glia in Drosophila

A prominent feature of glial cells is their ability to migrate along axons to finally wrap and insulate them. In the embryonic Drosophila PNS, most glial cells are born in the CNS and have to migrate to reach their final destinations. To understand how migration of the peripheral glia is regulated, we have conducted a genetic screen looking for mutants that disrupt the normal glial pattern. Here we present an analysis of two of these mutants: Notch and numb. Complete loss of Notch function leads to an increase in the number of glial cells. Embryos hemizygous for the weak Notch(B-8X) allele display an irregular migration phenotype and mutant glial cells show an increased formation of filopodia-like structures. A similar phenotype occurs in embryos carrying the Notch(ts1) allele when shifted to the restrictive temperature during the glial cell migration phase, suggesting that Notch must be activated during glial migration. This is corroborated by the fact that cell-specific reduction of Notch activity in glial cells by directed numb expression also results in similar migration phenotypes. Since the glial migration phenotypes of Notch and numb mutants resemble each other, our data support a model where the precise temporal and quantitative regulation of Numb and Notch activity is not only required during fate decisions but also later during glial differentiation and migration.     

Sexual dimorphism of craniological characters in Eurasian badgers, Meles spp. (Carnivora, Mustelidae)

An analysis of 30 craniological characters of Eurasian badgers (Meles spp.) revealed different levels of sexual size dimorphism (SSD) and geographic variation in the three different species. SSD is displayed mostly in the general size of the skull (condylobasal length, zygomatic width, width of rostrum, and cranial height) and mandible (height of the vertical mandibular ramus, total length of the mandible, and length between the angular process and infradentale), and in some dental characters (length of the upper molar M¹). The most stable size dimorphism is manifested in the size of the canines, which is pronounced in all studied samples. SSD is not apparent in the length of the auditory bulla, the postorbital width, the minimum palatal width, the length of the lower molar M₂, and the talonid length of the lower carnassial tooth M₁.In both the European badgers, Meles meles, and the Asian badgers, Meles leucurus, two geographic forms that differ in the degree of SSD have been found. The most pronounced SSD of cranial characters is found in the Transcaucasian form of Meles meles and the Far-Eastern form of Meles leucurus. In the large European form of Meles meles, SSD is less pronounced in both absolute and relative criteria than in the smaller Transcaucasian form. An analogous situation is observed in the larger Siberian and smaller Far-Eastern forms of Meles leucurus. In the Japanese badgers, Meles anakuma, a downsizing of the skull is accompanied by a decrease in SSD (except in canine size) in comparison to the continental species. The significant differences in the manifestation of SSD in the Eurasian badgers support an earlier hypothesis (Abramov, 2001. Proc. Zool. Inst. Russ. Acad. Sci. 288, 221-233; Abramov, 2002. Russ. J. Theriol. 1 (1), 57-60; Abramov, 2003. Small Carnivore Conserv. 29, 5-7) that Meles meles, Meles leucurus, and Meles anakuma are separate species. The differences in the level of SSD among and within these three species of badgers may be attributed not to differences in the diet or social structure, but to the history of speciation events and formation of intraspecific distribution ranges.     

Ontogenetic changes and sexual dimorphism in the mandible of adult woolly mammoths (Mammuthus primigenius)

Morphometric studies are a fundamental tool in paleontology to answer taxonomic, functional and evolutionary questions. In particular, appropriate functional interpretation often requires consideration of ontogenetic changes in the structures studied. The woolly mammoth (Mammuthus primigenius) is one of the most representative large mammal species of the Eurasian Pleistocene. Available ontogenetic studies on woolly mammoth mandible have focused on the first ontogenetic stages of the mandible development up to 4-5 years and have suggested that the symphysial process is sexually dimorphic. In the present work, we studied ontogenetic changes and sexual dimorphism in 45 mandibles from subadult and adult stages (8-56 African Elephant Years). Our results show positive correlations among almost all the morphometric variables measured, as well as an increase of mandible size with age. This increase does not differ among the variables examined, although the highest values are related with the symphysis height and the opening of the horizontal branches, and the lower ones with the greatest length (dimension), which implies the increase in the relative mandible width and height throughout the individual life. Sexual dimorphism in the mandible is at best slight, and the symphysial process is not diagnostic for sexing purposes. In addition, differences in age were an important confounding factor to assess sexual dimorphism and should be considered in future uses of sexual dimorphism assessment techniques.     

Foxh1 and Foxa2 are not required for formation of the midgut and hindgut definitive endoderm

The definitive endoderm forms during gastrulation and is rapidly transformed into the gut tube which is divided along the anterior-posterior axis into the foregut, midgut, and hindgut. Lineage tracing and genetic analysis have examined the origin of the definitive endoderm during gastrulation and demonstrated that the majority of definitive endoderm arises at the anterior end of the primitive streak (APS). Foxh1 and Foxa2 have been shown to play a role in specification of the APS and definitive endoderm. However, prior studies have focused on the role of these factors in specification of foregut definitive endoderm, while their role in the specification of midgut and hindgut definitive endoderm is less understood. Furthermore, previous analyses of these mutants have utilized definitive endoderm markers that are restricted to the anterior endoderm, expressed in extraembryonic endoderm, or present in other germ layers. Here, we characterized the expression of several novel definitive and visceral endoderm markers in Foxh1 and Foxa2 null embryos. In accordance with previous studies, we observed a deficiency of foregut definitive endoderm resulting in incorporation of visceral endoderm into the foregut. Interestingly, this analysis revealed that formation of midgut and hindgut definitive endoderm is unaffected by loss of Foxh1 or Foxa2. This finding represents a significant insight into specification and regionalization of mouse definitive endoderm.     

Intercontinental effect on sexual shape dimorphism and allometric relationships in the beetle pest Diabrotica virgifera virgifera LeConte (Coleoptera: Chrysomelidae)

Hind wing shape variation was examined in 686 adult Diabrotica virgifera virgifera collected from maize plants in Europe and the USA Corn Belt, using geometric morphometric techniques. Sexual dimorphism at an Intercontinental scale was assessed using canonical variates analysis, a multivariate statistical method used to find the shape characters that best distinguish among groups of specimens. Our results showed that each of the populations of D. v. virgifera investigated in this study showed high levels of sex based hind wing shape dimorphism. In particular a stronger and more obvious pattern of hind wing shape variation was found in the USA than in Europe. These results support previous studies on D. v. virgifera wing shape that show that female D. v. virgifera have more elongated wings than males. These differences raise the question of whether sexual dimorphism may be modulated by natural selection.     

Experimental manipulation of yolk testosterone affects digit length ratios in the ring-necked pheasant (Phasianus colchicus)

In humans, most of the mammals and one bird species studied so far, the relative length of individual digits is sexually dimorphic. Most studies of humans have been concerned with the ratio between second (2D) and fourth digits (4D), whereas some studies of humans and other mammals have also investigated other digit ratios. Inter- and intra-sexual variation in 2D:4D may depend on differential exposure to androgens during embryonic life, and the genetic mechanisms linking 2D:4D to androgens may be mediated by Hox genes. Because Hox genes are conserved in vertebrates, similar patterns of variation in digit ratios might be expected across vertebrate classes. The observation of correlations between digit ratios and physiological, psychological and performance traits in humans has generated interest in exploring the possibility that digit ratios are a marker of embryonic exposure to androgens, which have diverse consequences on several phenotypic traits. However, the hypothesis that digit ratios depend on androgen effects during development has never been tested experimentally. In this study, we increased testosterone concentration in ring-necked pheasant eggs and measured length ratios between the second, third and fourth digits of both feet in fully grown offspring. Females from testosterone-injected eggs had larger 2D:3D in the left foot, whereas this was not the case in males. The other digit ratios were unaffected by hormone treatment in both sexes. However, digit ratios showed no sexual dimorphism among controls. Thus, present results are consistent with the hypothesis that variation in testosterone levels during development affects digit ratios.     

Causes of secondary sexual differences in plants — Evidence from extreme leaf dimorphism in Leucadendron (Proteaceae)

In extreme cases leaves in male plants of the dioecious genus Leucadendron (Proteaceae) are up to an order of magnitude smaller than female leaves. This secondary sexual dimorphism (SSD) in leaf size has previously been suggested to be due to intra-male sexual selection, leading to an increase in male allocation to reproduction in dimorphic species. After critically evaluating previous data provided to support this hypothesis, I suggest on both theoretical grounds and on re-analysis that this argument is unlikely and unsupported. Leaf size dimorphism could theoretically evolve directly due to disruptive ecological selection between genders, leading to niche dimorphism either within or between habitats. I test this ecological causation hypothesis by providing data on specific leaf area (sla) and water use efficiency (δ ¹³C) of leaves from males and females of several Leucadendron species. Results confirm the expectation of minimal gender differences. I argue that leaf dimorphism is a consequence of selection on flower size and architecture.     

Androgen in free-living arctic ground squirrels: seasonal changes and influence of staged male-male aggressive encounters

We tested the responsiveness of plasma androgen in free-living male arctic ground squirrels (Spermophilus parryii) to challenges from conspecific males by staging aggressive encounters between males and then comparing androgen concentrations in manipulated and unmanipulated animals. The effect of staged encounters on androgen depended on time of year. In spring, androgen levels significantly increased after staged encounters, except during the peak of mating activity when levels were maximal in all reproductive males. In late summer, staged encounters had no significant effect on androgen levels. In spring but not summer, androgen was positively correlated to the relative intensity of encounters. These results suggest that circulating levels of androgen in male arctic ground squirrels result from a combination of seasonal and social factors. Responsiveness of androgen to encounters during spring but not late summer support the challenge hypothesis for this polygynous mammal.     

Babesia bigemina sexual stages are induced in vitro and are specifically recognized by antibodies in the midgut of infected Boophilus microplus ticks

Babesia bigemina, a causative agent of bovine babesiosis, is transmitted from one bovine to another only by infected ticks. The life cycle of B. bigemina includes a sexual phase in the tick host; however, molecules from sexual stages of any Babesia species have not been characterized. This is the first report of the induction of sexual stages of any Babesia species in vitro, free of tick antigens. Intraerythrocytic parasites were cultured in vitro for 20h using an induction medium. Extraerythrocytic parasites were first seen 3h post induction; elongated stages with long projections appeared at 6h post induction and by 9h they paired and fused to form larger stages. Round zygotes appeared 20h post induction. Moreover, by using Percoll gradients, sexual stages were purified free of contaminating intraerythrocytic stages. Purified parasites were used to generate polyclonal antibodies, which specifically bound to antigens expressed in sexual stages induced in vitro, but not to antigens expressed in intraerythrocytic stages. Importantly, these antibodies specifically identified sexual stages from midguts of female Boophilus microplus ticks fed on infected cattle.