Single cell transcriptomic analysis of external genitalia reveals complex and sexually dimorphic cell populations in the early genital tubercle.

External genital organs are among the most recognizable sexually dimorphic characters. The penis and clitoris develop from the embryonic genital tubercle, an outgrowth at the anterior margin of the cloaca that undergoes an extensive period of development in male and female embryos prior to the onset of sexual differentiation. In mice, differentiation into the penis and clitoris begins around embryonic day (E)15.5. Current knowledge of cell types that comprise the genital tubercle is limited to a few studies that have fate mapped derivatives of endoderm, mesoderm, and ectoderm. Here we use single cell transcriptomics to characterize the cell populations in the genital tubercles of male and female mouse embryos at E14.5, approximately 24 â€‹h before the onset of sexual differentiation, and we present the first comprehensive atlas of single-cell gene expression during external genital development. Clustering analyses and annotation using marker genes shows 19 distinct cell populations in E14.5 genital tubercles. Mapping of cell clusters to anatomical locations using in situ gene expression patterns revealed granularity of cellular specializations and positional identities. Although E14.5 precedes sexually dimorphic morphogenesis of the genital tubercle, comparative analysis of males and females identified sexual dimorphisms at the single cell level, including male-specific cell clusters with transcriptional signatures of smooth muscle and bone progenitors, both of which are known to be sexually dimorphic in adult genitalia, as well as immune cells. These results provide a new resource for classification of external genital cell types based on gene expression profiles and reveal sex-specific cellular specializations in the early genital tubercle.

Proliferative and nonproliferative lesions of the rat and mouse mammary, Zymbal's, preputial, and clitoral glands.

The mammary gland of laboratory rodents is an important organ for the evaluation of effects of xenobiotics, especially those that perturb hormonal homeostasis or are potentially carcinogenic. Mammary gland cancer is a leading cause of human mortality and morbidity worldwide and is a subject of major research efforts utilizing rodent models. Zymbal's, preputial, and clitoral glands are standard tissues that are evaluated in animal models that enable human risk assessment of xenobiotics. A widely accepted and utilized international harmonization of nomenclature for mammary, Zymbal's, preputial, and clitoral gland lesions in laboratory animals will improve diagnostic alignment among regulatory and scientific research organizations and enrich international exchanges of information among toxicologists and pathologists.

Clarification of the Innervation of the Bladder, External Urethral Sphincter and Clitoris: A Neuronal Tracing Study in Female Mongrel Hound Dogs.

Many studies examining the innervation of genitourinary structures focus on either afferent or efferent inputs, or on only one structure of the system. We aimed to clarify innervation of the bladder, external urethral sphincter (EUS) and clitoris. Retrograde dyes were injected into each end organ in female dogs. Spinal cord, mid-bladder, and spinal, caudal mesenteric, sympathetic trunk and pelvic plexus ganglia were examined for retrograde dye-labeled neurons. Neurons retrogradely labeled from the bladder were found primarily in L7-S2 spinal ganglia, spinal cord lateral zona intermedia at S1-S3 levels, caudal mesenteric ganglia, T11-L2 and L6-S2 sympathetic trunk ganglia, and pelvic plexus ganglia. The mid-bladder wall contained many intramural ganglia neurons labeled anterogradely from the pelvic nerve, and intramural ganglia retrogradely labeled from dye labeling sites surrounding ureteral orifices. Neurons retrogradely labeled from the clitoris were found only in L7 and S1 spinal ganglia, L7-S3 spinal cord lateral zona intermedia, and S1 sympathetic trunk ganglia, and caudal mesenteric ganglia. Neurons retrogradely labeled from the EUS were found in primarily at S1 and S2 spinal ganglia, spinal cord lamina IX at S1-S3, caudal mesenteric ganglia, and S1-S2 sympathetic trunk ganglia. Thus, direct inputs from the spinal cord to each end organ were identified, as well as multisynaptic circuits involving several ganglia, including intramural ganglia in the bladder wall. Knowledge of this complex circuitry of afferent and efferent inputs to genitourinary structures is necessary to understand and treat genitourinary dysfunction. Anat Rec, 2018. © 2018 Wiley Periodicals, Inc.

Preliminary report on the development and characterization of rabbit clitoral smooth muscle cell culture.

PURPOSE: Scientific model systems for physiological evaluation and investigation of pathophysiologies in clitoral function have been limited. The aim was to develop a New Zealand White rabbit clitoral corpus cavernosum smooth muscle cell culture. METHODS: Clitoral corpus cavernosum erectile tissue was harvested and placed in culture. Clitoral smooth muscle cells which migrated out from explants were grown to confluence and subcultured. Characterizations were performed by morphological and biochemical analyses. RESULTS: The cells exhibited typical morphologic characteristics of smooth muscle cells. Indirect immunofluorescence studies confirmed the presence of a-smooth muscle cell actin. Androgen and estrogen receptors were detected by specific antibodies and binding studies. The cells expressed subtypes of TGF-beta receptors. Treatment with 80 pM TGF-beta 1 24 h resulted in induction and/or increased availability of TGF-beta receptors. CONCLUSIONS: An in-vitro cell culture system using rabbit clitoral smooth muscle cells was developed. These smooth muscle cells retain their biochemical and functional integrity. This in-vitro cell culture system may facilitate studies aimed at understanding the molecular basis of female sexual function.

[Biological characteristics of female rabbit clitoral smooth muscle cell culture in vitro].

OBJECTIVE: To investigate the method of culturing rabbit clitoral smooth muscle cells(SMC) in vitro and their biological characteristics. METHODS: Rabbit clitoral SMCs were cultured in vitro by enzymatically dispersed method, and morphological observation, cell counting and immunohistochemical staining were employed to study their biological characteristics. RESULTS: Rabbit clitoral SMCs were spindle-shaped and parallelled along their longitudinal axis, and the attachment and proliferation in vitro were rapid. CONCLUSION: Rabbit clitoral SMCs cultured in vitro can grow and maintain their steady characteristics when provided with appropriate culture condition. Clitoral SMCs cultured in vitro can serve as a cell material for researches of clitoris.

Development and structure of the glandopreputial sulcus of the human clitoris with a special reference to glandopreputial glands.

The structure and development of the sulcus between the glans and prepuce of the human clitoris have hardly been investigated. Interest in its structure was raised when in the female, in contrast to the male, glands were found to develop from the solid lamella-like precursor of the glandopreputial sulcus. It prompted a further histological analysis of the sulcus in female fetuses and newborn and an extension of that study to clitorises of adult women. The investigation showed that in the clitoris, in contrast to the penis, the transformation of the glandopreputial lamella into the open sulcus was mostly incomplete and apparently remained so throughout life. As a most striking and probably exclusively female feature, two to eight eccrine glands developed from the base of the lamella in fetuses older than 14.5 weeks gestation. These glands formed secretory coils near and occasionally inside the adjacent distal corpora cavernosa. Some glands showed atresia, cystic dilatation, and squamous metaplasia. A remarkably similar picture was observed in the adult clitorises, in which the secretory coils were often found between the large blood vessels and nerves to the glans and were connected to the sulcus by long excretory ducts. All glands revealed unmistakably eccrine features. It is suggested that their secretion moistens the female glandopreputial sulcus, which is not lubricated by urethral secretion as in the male. The findings may explain the rare clitoral phimosis, cysts, and some pilonidal sinuses.

Sulcus nervi dorsalis penis/clitoridis: anatomic structure and clinical significance.

OBJECTIVES: The aim of this study was to correlate the anatomic and clinical significance of the prepubic course of dorsal nerve of penis and its groove on the pubic bone from the perspective of the surgeon. METHODS: The course of the dorsal nerve of the penis/clitoris was studied in six male and six female formalin-fixed cadavers. Several parameters of the pubis were quantified and analysed in 286 isolated pelvises. RESULTS: The course of the dorsal nerve of the penis is described in detail. This pubic sulcus was present in a majority of the pelvises examined. The dorsal nerve of the penis filled this groove, which we term the "sulcus nervi dorsalis penis/clitoridis." In contrast, the dorsal nerve and artery of the clitoris coursed in this groove in women. CONCLUSIONS: The course of dorsal nerve of the penis is described in detail including a previously unreported pubic structure-"sulcus nervi dorsalis penis/clitoridis." In the majority of individuals, the dorsal nerve of the penis/clitoris courses through the sulcus in the pubis. Based on the anatomy of the pelvic region, compression of the dorsal nerve is more apt to occur at the inferior border of the pubis or in the pubic sulcus than in the pudendal canal.

Oestrus cycle perturbations and hypotrophy of clitoral glands in malaria-infected female BALB/c mice.

An experimental host-parasite association involving BALB/c female mice infected with Plasmodium vinckei petteri was used with the aim of investigating the morphological and physiological alterations induced by the parasite in the genital tract of the host. The vaginal oestrous cycle was monitored as a daily clue to the sexual physiology of the female mice, and a complete histological analysis of the genital tract was performed 36 days following parasite inoculation. The oestrous cycle showed strong transitory alterations during the first 30 days following infection. The occurrence of oestrus days increased during the first 10 days post-infection and then decreased to a subnormal value during the following 20 days. Infected mice also showed a remarkable hypotrophy of their clitoral glands 30 days after the beginning of the malarial infection. A probable cause of such perturbations is a significant hormonal imbalance triggered by the erythrocytic proliferation of the Plasmodium. The relationship between the immune response of the host and these physiological and morphological alterations, as well as the outcomes of these alterations on the sexuality of the rodent host are discussed.

Postnatal development of vagina, clitoris and urethral glands of the golden hamster.

We have made a histological study of the postnatal development of the clitoris, preputial glands, urethral glands and vagina of the golden hamster. The 'phallic groove' of the clitoris is closed at 10 days of life, then the urethra has a cuboidal stratified, a stratified squamous and a stratified keratinized epithelium. The preputial glands are composed of branched saccular glands. These glands develop, with few changes during their maturation period. Formation of the urethral glands begins at 5 days and the alveoli are fully developed at puberty. The hamster vagina has two origins; the upper part is Müllerian, the caudal part is sinusal. The wall of the Müllerian vagina has a cylindrical epithelium at birth, which becomes 'double epithelium' at puberty and thereafter changes cyclically in connection with the estrous cycle. The sinusal vagina is solid at birth, its lumen being formed in the first 10 days of life and its wall having a cuboidal stratified epithelium. At 15 days it becomes a stratified keratinized epithelium, which will later line the vaginal pouch. At the 5th day, an ectodermic invagination (stratified keratinized epithelium) is observed in the zone of the future introitus. At the time of vaginal opening this zone forms the distal segment.