Spinal Nucleus Of The Bulbocavernosus Motoneuron Number Research Articles

Sexual Differentiation of the Nervous System: Where the Action Is

Sexual Differentiation of the Nervous System: Where the Action Is

Sexual Differentiation of the Spinal Nucleus of the Bulbocavernosus Is Not Mediated Solely by Androgen Receptors in Muscle Fibers

The spinal nucleus of the bulbocavernosus (SNB) neuromuscular system is a highly conserved and well-studied model of sexual differentiation of the vertebrate nervous system. Sexual differentiation of the SNB is currently thought to be mediated by the direct action of perinatal testosterone on androgen receptors (ARs) in the bulbocavernosus/levator ani muscles, with concomitant motoneuron rescue. This model has been proposed based on surgical and pharmacological manipulations of developing rats as well as from evidence that male rats with the testicular feminization mutation (Tfm), which is a loss of function AR mutation, have a feminine SNB phenotype. We examined whether genetically replacing AR in muscle fibers is sufficient to rescue the SNB phenotype of Tfm rats. Transgenic rats in which wild-type (WT) human AR is driven by a human skeletal actin promoter (HSA-AR) were crossed with Tfm rats. Resulting male HSA-AR/Tfm rats express WT AR exclusively in muscle and nonfunctional Tfm AR in other tissues. We then examined motoneuron and muscle morphology of the SNB neuromuscular system of WT and Tfm rats with and without the HSA-AR transgene. We observed feminine levator ani muscle size and SNB motoneuron number and size in Tfm males with or without the HSA-AR transgene. These results indicate that AR expression in skeletal muscle fibers is not sufficient to rescue the male phenotype of the SNB neuromuscular system and further suggest that AR in other cell types plays a critical role in sexual differentiation of this system.

Maternal licking influences dendritic development of motoneurons in a sexually dimorphic neuromuscular system

Maternal licking of pups' perineal regions affects the development of the spinal nucleus of the bulbocavernosus (SNB), a sexually dimorphic motor nucleus in the lumbar spinal cord that controls penile reflexes involved with copulation. Maternal licking influences SNB motoneuron number, with reductions in licking resulting in fewer motoneurons. Reduced maternal licking also has functional consequences in adulthood, resulting in increased latency to ejaculation and postejaculatory intromission and longer interintromission intervals. In this experiment, we assessed the potential effect of maternal licking on the development of SNB dendritic morphology. To reduce maternal licking, dams were treated with intranasal application of zinc sulfate during the first two postnatal weeks, which interferes with their ability to detect the pup odors that drive the licking behavior. At either postnatal day (P) 28 (when SNB dendritic length is normally maximal) or P49 (when SNB dendritic morphology is normally mature), SNB motoneurons were retrogradely labeled with cholera toxin-conjugated HRP, and dendritic arbor was reconstructed in three dimensions. At P28, the dendritic arbor of reduced maternal licking pups was not different from controls; however at P49, reduced licking pups showed a 23% reduction in dendritic arbor in the SNB, an effect that was especially pronounced in the rostral end of the nucleus, where reductions reached 48%. These results suggest that reductions in perineal stimulation provided by maternal licking could affect adult male copulatory behavior via alterations in SNB motoneuron morphology, and thus support maternal licking as an important factor in normal neural and behavioral development.

Effects of Bax Gene Deletion on Muscle and Motoneuron Degeneration in a Sexually Dimorphic Neuromuscular System

Motoneurons in the spinal nucleus of the bulbocavernosus (SNB) and their target muscles in the perineum, bulbocavernosus (BC), and levator ani (LA) normally degenerate in female rodents. Death of the motoneurons and muscles can be prevented by androgen treatments around the time of birth. To identify the intracellular mechanisms underlying hormone-dependent survival of this neuromuscular system, we examined mice with a targeted disruption of the pro-death gene Bax. SNB motoneuron number was increased in female Bax-/- mice, whether measured using immunolabeling for a motoneuron-specific marker or retrograde labeling with the fluorescent tracer Fluoro-Gold. Based on retrograde tracing, the sex difference in SNB cell number is eliminated in Bax-/- mice. Thus, Bax is required for sexually dimorphic motoneuron death in the SNB, and motoneurons rescued by Bax deletion project their axons to the periphery. Mean soma size in the SNB of Bax-/- females is reduced, however, and there is a subpopulation of very small cells in the SNB of female knock-outs. In addition, the BC muscle was not identified in any female, regardless of Bax gene status. All females possessed a small LA muscle, and Bax deletion resulted in a tripling of LA fiber number in females. This increase was small, however, relative to the >50-fold sex difference in LA muscle fiber number. Thus, the sex difference in the perineal muscles is mostly unaffected by the absence of Bax protein, and SNB motoneuron number is dissociated from target muscle size in Bax-/- animals.

Steroid Receptor Coactivator-1 Is Not Required for Androgen-Mediated Sexual Differentiation of Spinal Motoneurons

Steroid receptor coactivator-1 (SRC-1) amplifies genomic steroid hormone signal transduction and has been implicated in steroid-mediated sexual differentiation of the mammalian nervous system. We investigated the possible effect of an SRC-1 null mutation on 2 morphological endpoints of androgenic signaling: the number and size of motoneurons within the spinal nucleus of the bulbocavernosus (SNB). In wild-type C57/BL6 mice, SRC-1 immunoreactive nuclei were observed within the SNB and one of its target muscles, the levator ani. However, SRC-1 null mice were indistinguishable from sex-matched wild-type littermates in both SNB number and cross-sectional area of SNB motoneurons. Similarly, we found no difference between SRC-1 null and wildtype littermates in the number or size of motoneurons in the retrodorsolateral nucleus, a motor pool that is not typically sexually differentiated in either number or size. These results demonstrate that SRC-1 is not essential for the development and maintenance of a sexually dimorphic neuromuscular system.

Early postnatal response of the spinal nucleus of the bulbocavernosus and target muscles to testosterone in male gerbils

This study examined the response of the spinal nucleus of the bulbocavernosus (SNB) and the bulbocavernosus (BC) muscle, to testosterone in male Mongolian gerbils (Meriones unguiculatus) during the early postnatal period. Male gerbil pups were given testosterone propionate (TP) or vehicle for 2 days, then perfused on postnatal day (PND) 3, 5, 10 or 15. The BC and levator ani (LA) muscles were removed, weighed, and sectioned. Cross-sections of BC muscle fibers were measured and muscle fiber morphology examined. Spinal cords were removed and coronally sectioned in order to count and measure the SNB motoneurons. Following TP treatment, male pups of all ages had significantly heavier BC–LA muscles and larger fibers in the BC muscle compared to age-matched controls. The increase in muscle weight following TP treatment was greatest at PND10, while fiber size increased to a similar degree at all ages suggesting that hyperplasia as well as hypertrophy was responsible for the increase in muscle mass at this time. SNB motoneurons increased significantly in number and size with age and TP treatment. We hypothesize that the increase in SNB motoneuron number during normal ontogeny that can be augmented by TP treatment and represents an unusual means of establishing sexual dimorphism in the nervous system of a mammal through cell recruitment to the motor pool of a postnatal animal.

Long-term castration effects motoneuron size but not number in the spinal nucleus of the bulbocavernosus in the adult male Mongolian gerbil

The spinal nucleus of the bulbocavernosus (SNB) is a sexually dimorphic group of motoneurons in Rexed’s Lamina X of the lumbosacral spinal cord of the Mongolian gerbil. The SNB innervates the perineal musculature, the bulbocavernosus (BC), levator ani (LA), and external anal sphincter (EAS). Recent studies demonstrated a peripubertal component to the masculinization of the gerbil SNB with an apparent increase in both motoneuron size and number after puberty as measured with a Nissl stain. However, these studies could not determine if the apparent change in motoneuron number were due to the long-term castration involved in the methodology or due to a loss of motoneuron size beyond the point of being recognizable as motoneurons. Therefore, the current study was undertaken to examine this possibility by repeating the experimental protocol from previous peripubertal studies, on adult male gerbils with the addition of retrograde tract-tracing. Adult male gerbils were castrated at postnatal day (PND) 150 and given subcutaneous implants of testosterone proprionate (TP), dihydrotestosterone (DHT), estradiol benzoate (EB), EB and DHT, no steroid, or left intact. At PND215, the animals were injected with fluorogold (FG), a retrograde tract-tracer. At PND220, the animals were aldehyde perfused transcardially. The spinal cords were sectioned and alternate sections processed for either thionin-stain or FG visualization. Results indicated that long-term castration had no effect on SNB motoneuron number but did decrease SNB motoneuron size. TP, and to a lesser extent DHT, treatment could prevent the reduction in motoneuron size, however, EB could not. This study indicates that the maintenance of the adult male gerbil SNB–BC system is androgen, not estrogen, dependent and that long-term castration does not reduce motoneuron size to the point where they can no longer be distinguished.

Blockade of Endogenous Neurotrophic Factors Prevents the Androgenic Rescue of Rat Spinal Motoneurons

Target-derived neurotrophic factors are assumed to regulate motoneuron cell death during development but remain unspecified. Motoneuron cell death in the spinal nucleus of the bulbocavernosus (SNB) of rats extends postnatally and is controlled by androgens. We exploited these features of the SNB system to identify endogenously produced trophic factors regulating motoneuron survival. Newborn female rat pups were treated with the androgen, testosterone propionate, or the oil vehicle alone. In addition, females received trophic factor antagonists delivered either into the perineum (the site of SNB target muscles) or systemically. Fusion molecules that bind and sequester the neurotrophins (trkA-IgG, trkB-IgG, and trkC-IgG) were used to block activation of neurotrophin receptors, and AADH-CNTF was used to antagonize signaling through the ciliary neurotrophic factor receptor-alpha (CNTFRalpha). An acute blockade of trkB, trkC, or CNTFRalpha prevented the androgenic sparing of SNB motoneurons when antagonists were delivered to the perineum. Trophic factor antagonists did not significantly reduce SNB motoneuron number when higher doses were injected systemically. These findings demonstrate a requirement for specific, endogenously produced trophic factors in the androgenic rescue of SNB motoneurons and further suggest that trophic factor interactions at the perineum play a crucial role in masculinization of this neural system.

Sexual dimorphism in the number and size of SNB motoneurons: delayed development during normal ontogeny

The spinal nucleus of the bulbocavernosus (SNB) is a sexually dimorphic pool of motoneurons that innervates the perineal musculature. In the Mongolian gerbil, the SNB lies dorsolateral to the central canal within the lumbosacral spinal cord. Previously, no information was available on the normal development of the sexual dimorphism of this structure in the Mongolian gerbil, although evidence exists for a peripubertal development of the SNB in the gerbil. At each age from postnatal day 1 (PND1) through PND15 and at PND25, male and female gerbils were aldehyde perfused. Spinal cords were gelatin-embedded, cryoprotected, frozen and sectioned coronally through the lumbosacral transition zone and stained for Nissl substance with thionin. Examination by light microscopy revealed that the number of visible male and female SNB motoneurons significantly increased from PND1 through PND25. The size of the motoneurons also significantly increased in both sexes, however, until PND15 male gerbil SNB showed two significantly different sized populations of motoneurons. These data suggest the development of the SNB in the Mongolian gerbil is delayed, compared to the rat, and may continue well beyond the perinatal time period.

Additive Effects of Ciliary Neurotrophic Factor and Testosterone on Motoneuron Survival; Differential Effects on Motoneuron Size and Muscle Morphology

Testosterone and ciliary neurotrophic factor (CNTF) each enhance motoneuron survival in the spinal nucleus of the bulbocavernosus (SNB) of newborn rats. Here we directly compared the effects of CNTF and testosterone, alone and in combination, on SNB motoneuron number, SNB cell size, and morphology of the levator ani (LA) target muscle. Female rat pups were treated daily from postnatal day 1 through 6 (P1–P6) with recombinant human CNTF (hCNTF), testosterone propionate (TP), both hCNTF and TP, or neither. Effects of treatment were assessed on P7. TP and hCNTF each increased the number of SNB motoneurons and did so to a similar degree. Females treated with both hCNTF and TP had significantly more SNB cells than those receiving either hCNTF or TP alone. TP administered from P1 to P6 also increased SNB motoneuron size on P7. In contrast, hCNTF alone did not significantly affect SNB cell size, and hCNTF in combination with TP antagonized the effect of TP on motoneuron size. TP also increased LA muscle fiber number and LA fiber size, whereas hCNTF did not significantly influence LA muscle morphology in this study. Immunohistochemistry established that virtually all SNB motoneurons of both males and females express the CNTF α receptor (CNTFRα) between embryonic day 20 and postnatal day 6. Thus, effects of TP and hCNTF on SNB motoneuron survival were additive, and increases in motoneuron survival were dissociated from changes in target muscle morphology in hCNTF-treated animals. SNB motoneurons express CNTFRα perinatally and are therefore potential direct sites of hCNTF action.

Effects of testosterone on the development of a sexually dimorphic neuromuscular system in ciliary neurotrophic factor receptor knockout mice.

Motoneurons in the spinal nucleus of the bulbocavernosus (SNB) innervate the perineal muscles, bulbocavernosus (BC), and levator ani (LA). Testosterone regulates the survival of SNB motoneurons and BC/LA muscles during perinatal life. Previous findings suggest that effects of testosterone on this system may be mediated by trophic factors-in particular, by a factor acting through the ciliary neurotrophic factor alpha-receptor (CNTFRalpha). To test the role of CNTFRalpha in the response of the developing SNB system to testosterone, CNTFRalpha +/+ and -/- mice were treated with testosterone propionate (TP) or oil during late embryonic development. BC/LA muscle size and SNB motoneuron number were evaluated on the day of birth. Large sex differences in BC and LA muscle size were present in newborn mice of both genotypes, but muscle volumes were reduced in CNTFRalpha -/- animals relative to same-sex, wild-type controls. Prenatal testosterone treatment completely eliminated the sex difference in BC/LA muscle size in wild-type animals, and eliminated the effect of the CNTFRalpha gene deletion on muscle size in males. However, the effect of TP treatment on BC and LA muscle sizes was blunted in CNTFRalpha -/- females. SNB motoneuron number was sexually dimorphic in oil-treated, wild-type mice. In contrast, there was no sex difference in SNB motoneuron number in oil-treated, CNTFRalpha knockout mice. Prenatal treatment with testosterone did not increase SNB motoneuron number in CNTFRalpha -/- mice, but also did not significantly increase SNB motoneuron number in newborn wild-type animals. These findings confirm the absence of a sex difference in SNB motoneuron number in CNTFRalpha -/- mice. Moreover, the CNTFRalpha gene deletion influences perineal muscle development and the response of the perineal muscles to testosterone. Prenatal TP treatment of CNTFRalpha -/- males overcomes the effects of the gene deletion on the BC and LA muscles without a concomitant effect on SNB motoneuron number.

Sexual Dimorphism in the Spinal Cord Is Absent in Mice Lacking the Ciliary Neurotrophic Factor Receptor

Ciliary neurotrophic factor (CNTF) has potent survival-promoting effects on motoneurons in vitro and in vivo. We examined knockout mice with null mutations of the gene for either CNTF itself or the alpha-subunit of the CNTF receptor (CNTFRalpha) to assess whether CNTF and/or its receptors are involved in the development of a sexually dimorphic neuromuscular system. Male rodents have many more motoneurons in the spinal nucleus of the bulbocavernosus (SNB) than do females. This sex difference is caused by hormone-regulated death of SNB motoneurons and their target muscles. Sexual dimorphism of SNB motoneuron number developed completely normally in CNTF knockout (CNTF -/-) mice. In contrast, a sex difference in the SNB was absent in CNTFRalpha -/- animals: male mice lacking a functional CNTF alpha-receptor had fewer than half as many SNB motoneurons than did wild-type males and no more than did their female counterparts. Size of the bulbocavernosus and levator ani muscles, the main targets of SNB motoneurons, was not affected in either CNTF or CNTFRalpha knockout males. These observations suggest that signaling through the CNTF receptor is involved in sexually dimorphic development of SNB motoneuron number and that target muscle survival per se is not sufficient to ensure motoneuron survival in this system. In addition, our observations are consistent with the suggestion that CNTF itself is not the only endogenous ligand for the CNTF receptor. A second, as yet unknown, ligand may be important for neural development, including sexually dimorphic motoneuron development.

Ontogeny of androgen receptor immunoreactivity in lumbar motoneurons and in the sexually dimorphic levator ani muscle of male rats

We documented the ontogeny of androgen receptor (AR) immunoreactivity for rat lumbar motoneurons of the sexually dimorphic motor pools, the spinal nucleus of the bulbocavernosus (SNB) and the dorsolateral nucleus (DLN), and for the sexually monomorphic retrodorsolateral nucleus (RDLN). We also assessed the ontogeny of AR immunoreactivity in the rat sexually dimorphic levator ani (LA), which is a target muscle for SNB motoneurons. Lumbar spinal cords and LA muscles from gonadally intact males at ages postnatal days (P)7, P10, and P14 and as adults were incubated with the rabbit antiserum PG-21. Half of the prepubertal males (P7–P14) received 200 μg of testosterone propionate (TP) 2 hours prior to death to enhance immunodetection of ARs. We found that SNB motoneurons developed AR immunoreactivity first and achieved adult levels by P10. In contrast, the number of RDLN motoneurons with AR-immunopositive nuclei during development remained well below the adult number. Development of AR immunoreactivity in the DLN shared characteristics with both the SNB and the RDLN. AR immunoreactivity developed in some DLN motoneurons by P10, although the percentage of labelled motoneurons remained below that in adulthood. Acute TP treatment significantly increased the number of SNB motoneurons with AR-positive nuclei at P7. The LA showed a robust pattern of AR immunostaining from P7 to adulthood. Immunostaining was present only in nuclei and constituted only a subpopulation of the nuclei present in muscle. The present results confirm and extend previous results based on steroid autoradiography and steroid binding assays regarding regional and developmental differences in the expression of ARs. J. Comp. Neurol. 379:88-98, 1997. © 1997 Wiley-Liss, Inc.

Ontogeny of androgen receptor immunoreactivity in lumbar motoneurons and in the sexually dimorphic levator ani muscle of male rats

We documented the ontogeny of androgen receptor (AR) immunoreactivity for rat lumbar motoneurons of the sexually dimorphic motor pools, the spinal nucleus of the bulbocavernosus (SNB) and the dorsolateral nucleus (DLN), and for the sexually monomorphic retrodorsolateral nucleus (RDLN). We also assessed the ontogeny of AR immunoreactivity in the rat sexually dimorphic levator ani (LA), which is a target muscle for SNB motoneurons. Lumbar spinal cords and LA muscles from gonadally intact males at ages postnatal days (P)7, P10, and P14 and as adults were incubated with the rabbit antiserum PG-21. Half of the prepubertal males (P7-P14) received 200 micrograms of testosterone propionate (TP) 2 hours prior to death to enhance immunodetection of ARs. We found that SNB motoneurons developed AR immunoreactivity at first and achieved adult levels by P10. In contrast, the number of RDLN motoneurons with AR-immunopositive nuclei during development remained well below the adult number. Development of AR immunoreactivity in the DLN shared characteristics with both the SNB and the RDLN. AR immunoreactivity developed in some DLN motoneurons by P10, although the percentage of labelled motoneurons remained below that in adulthood. Acute TP treatment significantly increased the number of SNB motoneurons with AR-positive nuclei at P7. The LA showed a robust pattern of AR immunostaining from P7 to adulthood. Immunostaining was present only in nuclei and constituted only a subpopulation of the nuclei present in muscle. The present results confirm and extend previous results based on steroid autoradiography and steroid binding assays regarding regional and developmental differences in the expression of ARs.

Short- and long-term effects of ciliary neurotrophic factor on androgen-sensitive motoneurons in the lumbar spinal cord.

Motoneuron death in the spinal nucleus of the bulbocavernosus (SNB) and the dorsolateral nucleus (DLN) of the lumbar spinal cord is androgen regulated. As a result, many more SNB and DLN motoneurons die in perinatal female rats than in males, whereas treatment of newborn females with androgen results in a permanent sparing of the motoneurons and their target muscles. We previously observed that a neurotrophic molecule, ciliary neurotrophic factor (CNTF), also arrests the death of SNB motoneurons and their target musculature, at least in the short term. The present study compares the short- and long-term consequences of perinatal CNTF treatment on motoneuron number in the SNB, the DLN, and the retrodorsolateral nucleus (RDLN), a motor pool in the lower lumbar cord that does not exhibit hormone-regulated cell death. Female pups were treated with CNTF or vehicle alone from embryonic day 22 through postnatal day 6 (P6). Motoneuron number in the each nucleus was then determined immediately after treatment on P7, or 10 weeks later (P77). CNTF treatment significantly elevated motoneuron number in the SNB and DLN on P7; the volume of SNB target muscles on P7 was also greater in the CNTF-treated group. These effects were transient, however, as motoneuron number and ratings of muscle size were not different in CNTF-and vehicle-treated females on P77. Perinatal CNTF treatment did not alter cell number in the RDLN at either age. The finding that effects of CNTF on SNB and DLN motoneuron number are short lived contrasts with the permanent effects of early androgen treatment, and has implications for molecular models of the actions of androgen and neurotrophic factors on the developing spinal cord.

Intrauterine position affects motoneuron number and muscle size in a sexually dimorphic neuromuscular system

The intrauterine position occupied by a rodent fetus influences the amount of testosterone to which it is exposed before birth. Animals that are gestated between two male fetuses (2M) are exposed to higher circulating levels of testosterone than are animals positioned between two female fetuses (2F) and there are reliable differences in the reproductive physiology and behavior of 2M and 2F animals when adult. To determine whether intrauterine position modifies development of the central nervous system, we examined the sexually dimorphic spinal nucleus of the bulbocavernosus (SNB) in male and female gerbils from known intrauterine positions. We found that adult 2M female gerbils had 16% more SNB motoneurons than did 2F females. 2M males did not differ from 2F males in SNB motoneuron number, but the bulbocavernosus muscle, which is innervated by SNB motoneurons, was approximately 50% larger in 2M than in 2F males. These data indicate that intrauterine position can influence the morphology of the sexually dimorphic SNB neuromuscular system.

Intrauterine position affects motoneuron number and muscle size in a sexually dimorphic neuromuscular system

The intrauterine position occupied by a rodent fetus influences the amount of testosterone to which it is exposed before birth. Animals that are gestated between two male fetuses (2M) are exposed to higher circulating levels of testosterone than are animals positioned between two female fetuses (2F) and there are reliable differences in the reproductive physiology and behavior of 2M and 2F animals when adult. To determine whether intrauterine position modifies development of the central nervous system, we examined the sexually dimorphic spinal nucleus of the bulbocavernosus (SNB) in male and female gerbils from known intrauterine positions. We found that adult 2M female gerbils had 16% more SNB motoneurons than did 2F females. 2M males did not differ from 2F males in SNB motoneuron number, but the bulbocavernosus muscle, which is innervated by SNB motoneurons, was approximately 50% larger in 2M than in 2F males. These data indicate that intrauterine position can influence the morphology of the sexually dimorphic SNB neuromuscular system.

Motoneurons dorsolateral to the central canal innervate perineal muscles in the mongolian gerbil

The Mongolian gerbil provides a model in which sexually dimorphic areas in the hypothalamus are correlated with sociosexual behaviors such as scent marking and male copulatory behavior. To extend this model, investigations were conducted to determine whether sexually dimorphic areas existed in the spinal cord that could be relevant to male sexual behavior. The focus of these investigations was the perineal muscles associated with the penis. Therefore, this research identified the spinal motoneurons that innervate the bulbocavernosus, levator ani, anal sphincter, and ischiocavernosus muscles of Mongolian gerbils. The motoneuron pool that innervates the bulbocavernosus, levator ani, and anal sphincter was designated the spinal nucleus of the bulbocavernosus (SNB), as for other species of rodents. The motoneuron pool innervating the ischiocavernosus was identified as the dorsolateral nucleus, again, to be consistent with the designation for other rodents. The motoneurons of the gerbil SNB were distributed dorsolateral to the central canal in the lumbosacral transition zone of the spinal column. These motoneurons are located in the region classically defined as area X of the spinal cord. The number of SNB motoneurons was sexually dimorphic, with male gerbils having about five times as many SNB motoneurons as do female gerbils. The size of SNB motoneurons was also sexually dimorphic. The SNB motoneurons of males were 1.5 times larger than the SNB motoneurons of females. The effects of adult castration on the male SNB were also studied. After castration, the size, but not the number, of SNB motoneurons in males was significantly decreased. This decrease was prevented by testosterone treatment. The percentage of calcitonin gene-related peptide (CGRP)-immunoreactive SNB motoneurons was also affected by adult castration. The percentage of CGRP-immunoreactive motoneurons was significantly decreased after adult castration. Again, this decrease was reversed by testosterone treatment. These findings suggest that the SNB of gerbils is sexually dimorphic and is sensitive to circulating levels of gonadal steroids. The unique placement of the SNB motoneurons suggests that an alternative laminar organizational scheme may be necessary for Mongolian gerbil.

Calcitonin gene-related peptide-like immunoreactivity in spinal motoneurons of the male mouse is affected by castration and genotype

Calcitonin gene-related peptide (CGRP) is found in motoneurons of the mammalian spinal cord, including motoneurons of the androgen-dependent spinal nucleus of the bulbocavernosus (SNB) of the mouse. Motoneurons of the SNB innervate the bulbocavernosus (BC), a striated muscle involved in penile reflexes. CGRP is thought to be a trophic factor produced by motoneurons to regulate the expression of the acetylcholine receptor at the neuromuscular junction. In rats, the number of SNB motoneurons containing CGRP is increased by gonadal steroids. This regulation appears to rely on an activity-dependent factor produced by the BC muscle. The purpose of the present study was to examine, using immunohistochemistry, the steroid dependence of CGRP in the SNB of male house mice. Genotypic differences in the steroid regulation of CGRP immunoreactivity were examined in three strains of mice that differ in their behavioral sensitivity to castration. The results demonstrate that castration reduces the number of CGRP-positive SNB motoneurons in mice. The magnitude of the change in CGRP in response to castration and the length of time required following castration to alter CGRP were dependent on genotype. Interestingly, the effect of castration in mice, to reduce the number of CGRP-immunoreactive SNB motoneurons, is opposite in direction from the increase in CGRP SNB motoneurons observed in rats observed following castration. These experiments suggest that androgens may alter neuromuscular junction function of mouse SNB by regulating the production of CGRP in a species-specific, genotypically dependent fashion.

Timing and duration of dihydrotestosterone treatment affect the development of motoneuron number and morphology in a sexually dimorphic rat spinal nucleus

The spinal nucleus of the bulbocavernosus (SNB) is a sexually dimorphic motor nucleus in the rat lumbar spinal cord. SNB motoneurons and their perineal target muscles are present in adult males, but reduced or absent in adult females. This dimorphism is due to the presence of androgens during development. Perinatal treatment of females with testosterone (T), or a combination of dihydrotestosterone (DHT) and estrogen (E+D females) from embryonic (E) day 16 through postnatal (P) day 5, results in a masculine number of SNB motoneurons and the retention of the target muscles. Perinatal treatment with estrogen alone does not masculinize the SNB; prenatal treatment with DHT alone from E17-E22 results in a feminine number of SNB motoneurons and a significantly altered motoneuron morphology and connectivity. To determine if masculinization of the SNB involves the interaction of estrogen and DHT or results from a longer exposure to DHT alone, the number, morphology, and connectivity of SNB motoneurons in females treated with DHT both pre- and post-natally (from E16-P5) were examined. At E22, DHTP (E16-P5) females have SNB motoneuron numbers identical to E+D and normal females, but far fewer than normal males, thus indicating that T is essential for prenatal masculinization. After E22, SNB motoneuron number declines precipitously in normal females but remains stable in DHTP (E16-P5) females and E+D females, which do not differ from normal males at P10. These results demonstrate that DHT can completely masculinize SNB motoneuron number without any synergistic actions with estrogen, and suggest that the development of SNB motoneuron number is strictly an androgen-mediated event. In adulthood, horseradish peroxidase histochemistry reveals that the connectivity, dendritic length, and soma size of SNB motoneurons in DHTP (E16-P5) females are identical to those of normal males but differ significantly from those of DHTP (E17-E22) females. These data suggest that the altered connectivity in DHTP (E17-E22) females is not simply a hormone-specific effect, but the result of a truncated hormone exposure. Thus, DHT can fully masculinize SNB morphology and connectivity if given during the appropriate period of development. It is suggested that while T may be required to masculinize the SNB prenatally, DHT may be involved in masculinizing postnatal aspects of SNB development.