Hormonal control of neuron number in sexually dimorphic spinal nuclei of the rat: I. Testosterone-regulated death in the dorsolateral nucleus

Adult male rats have substantially more motoneurons than do females in two motor nuclei in the lumbar spinal cord: the spinal nucleus of the bulbocavernosus (SNB) and the dorsolateral nucleus (DLN). Previous studies of the development of the SNB revealed that the sex difference in SNB motoneuron number is established through a differential motoneuron death which is under the control of androgens. In this study the development of the sexually dimorphic DLN was examined to test the hypothesis that early androgen action also determines the sex difference in DLN motoneuron number by regulating normally occurring motoneuron death. Because SNB motoneurons may migrate from the DLN, quantitative examination of DLN development was necessary in order to understand more completely the cellular mechanisms contributing to the establishment of dimorphic motoneuron number. At 5 days before birth, the number of motoneurons in the DLN is significantly higher than in adulthood in both sexes, and no sex difference is present. There is a decrease in motoneuron numbers prenatally in both sexes, which is consistent with the emigration of presumptive SNB motoneurons. Motoneuron number declines differentially through the first week of postnatal life and by postnatal day 10 motoneuron numbers are in the adult range and the sex difference is fully expressed. Females lose significantly more DLN motoneurons than males through a differential death as revealed by the higher incidence of degenerating cell profiles. Females treated with testosterone propionate have a male-typical motoneuron loss and incidence of degenerating cells. These results indicate that steroid hormones establish the sex difference in DLN motoneuron number by regulating normally occurring cell death.     

Androgen regulation of dendritic growth and retraction in the development of a sexually dimorphic spinal nucleus

The spinal nucleus of the bulbocavernosus (SNB) is a sexually dimorphic group of motoneurons whose development and maintenance are under androgenic control. Exposure to androgens early in development permanently alters SNB motoneuron number and soma size; in adulthood, androgens regulate dendritic and synaptic architecture. The present set of experiments investigates the influence of androgens on the development of SNB dendritic morphology. In normal males, SNB dendritic growth is biphasic, reaching exuberant lengths by the fourth postnatal week and then retracting to adult lengths by 7 weeks of age. This dendritic growth is androgen dependent--males castrated on postnatal day (P) 7 and given daily injections of testosterone propionate (TP) had exuberant dendritic lengths similar to those of normal males; dendritic length in oil-treated males remained at P7 levels. The early exuberant dendritic length was retained in TP-treated males through P49. The retraction of SNB dendrites after P28 is also influenced by androgens. Males castrated at P28 and given testosterone implants retained exuberant dendritic length at P49; blank-implanted males had significantly shorter dendritic lengths by P70. These results suggest that androgens are necessary for the early exuberant growth of SNB dendrites. Furthermore, the subsequent retraction of SNB dendrites may be halted when testosterone titers reach a critical level during puberty, stabilizing their adult length.     

Development and loss of early projections in a sexually dimorphic rat spinal nucleus

The spinal nucleus of the bulbocavernosus (SNB) contains 3-4 times more motoneurons in adult male rats compared to females. This large dimorphism in motoneuron number is produced perinatally by an androgen-regulated cell death. To determine if the early projections of the SNB to its target musculature may be involved in the creation of this sexual dimorphism, and how these projections might interact with androgens, HRP tracing techniques were used to retrogradely label SNB motoneurons during prenatal and postnatal development in males, females, and masculinized females. HRP labeling revealed that the prenatal formation of early projections of the SNB in males and females is comparable. SNB motoneuron number increases through the day before birth in all groups, and during this increase, labeled cells can be seen outside the SNB, which we hypothesize are in the process of migrating into the SNB from the lateral motoneuron column. Postnatally, SNB motoneuron number declines, especially in females, and by postnatal day 10 the sexual dimorphism in cell number and projections has been established. These results indicate that although masculine androgen levels are critical in determining SNB motoneuron survival, they are not necessary for initial axon outgrowth of SNB motoneurons. However, androgens may be involved in the regulation of SNB motoneuron migration and the stabilization of the peripheral projections of the SNB. Both male and female SNB motoneuron axons are present at their target muscles during the time in which sex differences in motoneuron number develop, suggesting that the interaction of SNB motoneurons with their targets could be involved in the dimorphic regulation of cell survival.     

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.     

Motoneuron morphology in the dorsolateral nucleus of the rat spinal cord: Normal development and androgenic regulation

The rat lumbar spinal cord contains two sexually dimorphic motor nuclei, the spinal nucleus of the bulbocavernosus (SNB), and the dorsolateral nucleus (DLN). These motor nuclei innervate anatomically distinct perineal muscles that are involved in functionally distinct copulatory reflexes. The motoneurons in the SNB and DLN have different dendritic morphologies. The dendrites of motoneurons in the medially positioned SNB have a radial, overlapping arrangement, whereas the dendrites of the laterally positioned DLN have a bipolar and strictly unilateral organization. During development, SNB motoneuron dendrites grow exuberantly and then retract to their mature lengths. In this experiment we determined whether the adult difference in SNB and DLN motoneuron morphology was reflected in different patterns of dendritic growth during normal development. Furthermore, the development of both these nuclei is under androgenic control. In the absence of androgens, SNB dendrites fail to grow; testosterone replacement supports normal dendritic growth. Thus, we also examined the development of DLN dendrites for similar evidence of androgenic regulation. By using cholera toxin-horseradish peroxidase (BHRP) to label motoneurons retrogradely, we measured the morphology of DLN motoneurons in normal males, and in castrates treated with testosterone or oil/blank implants at postnatal day (P) 7, P28, P49, and P70. Our results demonstrate that in contrast to the biphasic pattern of dendritic development in the SNB, dendritic growth in the DLN was monotonic; the dendritic length of motoneurons increased more than 500% between P7 and P70. However, as in the SNB, development of DLN motoneuron morphology is androgen-dependent. In castrates treated with oil/blank implants, DLN somal and dendritic growth were greatly attenuated compared to those of normal or testosterone-treated males. Thus, while androgens are clearly necessary for the growth of motoneurons in both the SNB and DLN, their different developmental patterns suggest that other factors must be involved in regulating this growth. © 1993 Wiley-Liss, Inc.     

Motoneuron development after deafferentation. I. Dorsal rhizotomy does not alter growth in the spinal nucleus of the bulbocavernosus (SNB)

The spinal nucleus of the bulbocavernosus (SNB) and the dorsolateral nucleus (DLN) are sexually dimorphic motor nuclei in the rat lumbar spinal cord. During postnatal development, SNB and DLN motoneurons grow substantially in measures of soma size, dendritic length, and radial dendritic extent. SNB motoneurons exhibit a biphasic pattern of dendritic growth, where there is an initial period of exuberant growth followed by a period of retraction to mature lengths by 7 weeks. In this experiment, we examined whether primary afferent input to the SNB nucleus was necessary for the normal postnatal growth of SNB motoneurons. We partially deafferented the SNB via unilateral dorsal rhizotomy of lumbosacral dorsal roots in male rats at 1 week of age. Using cholera toxin horseradish peroxidase (BHRP) to visualize SNB motoneurons, we examined SNB motoneuron morphology at 4 and 7 weeks of age. SNB motoneurons in rhizotomized males developed normally; measures of dendritic length in rhizotomized males were typically exuberant at 4 weeks of age, and declined significantly to mature lengths by 7 weeks of age. In addition, dorsal rhizotomy did not alter the development of SNB motoneuron soma size or radial dendritic extent. These results are discussed in reference to sensorimotor connections in the SNB, the extent of the deafferentation, and dendrodendritic interactions.     

Sexually dimorphic motor nucleus in the rat lumbar spinal cord: Response to adult hormone manipulation, absence in androgen-insensitive rats

There is a sexually dimorphic motor nucleus in the fifth and sixth lumbar segments of the rat spinal cord, consisting of motoneurons innervating two striated perineal muscles, the levator ani and the bulbocavernosus. This nucleus, which is diminished or absent in female rats, has been named the spinal nucleus of the bulbocavernosus (SNB)³. We now report that the number of neurons in the SNB of either male or female rats is not altered by adult gonadectomy or treatment with testosterone propionate. However, the size of individual SNB neurons is increased in the presence of androgen in either sex. Genetically male rats with the testicular feminization mutation which results in reduced androgen receptors have a markedly feminine SNB. These results support the hypothesis that the sexually dimorphic nature of the SNB depends on neither the adult hormonal state nor the presence of a Y chromosome, but on the interaction of androgens with their receptors early in development.     

Sexual dimorphism and hormone responsiveness in the spinal cord of the socially monogamous prairie vole (Microtus ochrogaster)

Prairie voles (Microtus ochrogaster) are exceptional among rodents in that many aspects of their brain and behavior are not masculinized by exogenous aromatizable androgens. However, the sexually differentiated endpoints studied to date rely on estrogenic mechanisms in other mammals. We examined whether sexual differentiation of an androgen receptor‐dependent sex difference would be similarly distinct in prairie voles. Male mammals have more and larger motoneurons projecting to perineal muscles than do females. This sex difference normally arises from males' perinatal androgen exposure and can be eliminated by treating developing females with androgens. Gross dissection revealed bulbospongiosus muscles in adult male, but not female, prairie voles. Retrograde tracing from males' bulbocavernosus muscles and the external anal sphincter from both sexes revealed sexually dimorphic populations of labeled motoneurons in the ventral horn of the lumbar spinal cord. Similar to other rodents, males had twice as many motoneurons as females, although no sex difference in motoneuron size was detected. Unexpectedly, prenatal or early postnatal exposure to testosterone propionate had no effect on adult females' motoneuron number or size. In adulthood, gonadectomy alone or followed by chronic testosterone treatment also had no effect on females' motoneuron size or number, although castration reduced motoneuron size in males. Comparing gonadally intact weanlings confirmed that the sex difference in motoneuron number exists before adulthood. As with some other sexually dimorphic traits, and perhaps related to their unique social organization, sexual differentiation of the prairie vole spinal cord differs from that found in most other laboratory rodents. J. Comp. Neurol. 516:117–124, 2009. © 2009 Wiley‐Liss, Inc.     

N-methyl-D-aspartate receptor blockade inhibits estrogenic support of dendritic growth in a sexually dimorphic rat spinal nucleus

The lumbar spinal cord of rats contains the sexually dimorphic, steroid-sensitive spinal nucleus of the bulbocavernosus (SNB). Dendritic development of SNB motoneurons requires the action of both androgens and estrogens. Estrogenic effects are limited to the initial growth of SNB dendrites through 4 weeks of age. During this postnatal period, dendritic growth in other spinal motoneurons is regulated by N-methyl-D-aspartate (NMDA) receptor activation. In this study, we tested whether NMDA receptor activation was involved in SNB dendritic growth and whether the estrogenic support of SNB dendritic growth was dependent on the activation of NMDA receptors. Motoneuron morphology was assessed in normal males, intact males treated daily with the NMDA receptor antagonist MK-801, castrated males treated with estradiol benzoate (EB), and castrated males treated with both EB and MK-801. SNB motoneurons were retrogradely labeled with cholera toxin-horseradish peroxidase at 4 weeks of age (when dendritic length is normally maximal) and reconstructed in three dimensions. Somal area and dendritic length of SNB motoneurons in MK-801-treated, intact males were below those of normal males. Dendritic growth was partially supported in EB-treated castrates, but this growth was blocked by MK-801 treatment. These results suggest that, as in other motoneurons, dendritic development in the SNB involves NMDA receptors and, furthermore, that the estrogen-sensitive component of SNB dendritic development requires their activation.     

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. © 1995 Wiley-Liss, Inc.     

Hormonal control of neuron number in sexually dimorphic spinal nuclei of the rat: III. Differential effects of the androgen dihydrotestosterone

The spinal cord of the rat contains two sexually dimorphic nuclei: the spinal nucleus of the bulbocavernosus (SNB) and the dorsolateral nucleus (DLN). These nuclei and the perineal muscles they innervate are present in males but reduced or absent in females. The sex difference in motoneuron number in these nuclei is due to an androgen-regulated motoneuron death. Developing females treated with the androgen testosterone propionate (TP) have a fully masculine number of SNB and DLN motoneurons and retain the perineal muscles they would normally have lost. Paradoxically, females treated prenatally with the androgen dihydrotestosterone propionate (DHTP) also retain the perineal musculature but as adults lack the SNB motoneurons which would normally innervate them. The SNB target muscles retained by DHTP females are anomalously innervated by motoneurons in the DLN. Counts of motoneurons and degenerating cells in the developing SNB of DHTP-treated females showed that their feminine number is the result of a failure of DHTP to prevent the death of SNB motoneurons. Furthermore, the peak number of SNB motoneurons was below that of normal females, suggesting that DHTP treatment may also have inhibited motoneuronal migration. However, DHTP treatment fully masculinized both motoneuron number and degenerating cell counts in the DLN of these females, and it is this masculinized DLN that gives rise to the anomalous projection. Taken together, these results suggest that the effects of different androgens during development are specific and complex, involving the regulation of motoneuron death, migration, and specification of peripheral projections.     

Hormonal control of a developing neuromuscular system. I. Complete Demasculinization of the male rat spinal nucleus of the bulbocavernosus using the anti-androgen flutamide

Prenatal treatment of male rats with the anti-androgen, flutamide (FL), demasculinizes the sexually dimorphic spinal nucleus of the bulbocavernosus (SNB) by reducing the number of SNB neurons, the size of the somas and nuclei of SNB neurons, and the size of their target muscles in adulthood. However, FL does not affect mounting or the traditional, postural measure of intromission, indicating that the SNB system does not play a major role in the mediation of these particular behaviors. Postnatal testosterone propionate (TP) treatment of male rats castrated on the day of birth results in more male copulatory behaviors in adulthood and masculinizes all measures of the SNB system. The postnatal masculinization by TP is more pronounced in males treated prenatally with FL, for morphological but not behavioral measures. The combined treatment of prenatal FL and day 1 castration without TP therapy results in a male with a completely demasculinized SNB system. Specifically, such males have SNB neurons that are as scarce and as small as those of females and, like females, they lack the target muscles of the SNB. These results support the hypothesis that perinatal androgens normally direct the sexually dimorphic development of the SNB and its target muscles.     

Testosterone fails to save androgen-sensitive rat motoneurons following early target removal.

Axotomy during development can result in the death of up to 100% of the affected motoneurons. However, axotomy-induced death can be significantly reduced by administration of androgens in young rats. Motoneuron death in the spinal nucleus of the bulbocavernosus (SNB) has previously been shown to be regulated by androgens during development. the present experiment examined the effects of androgen treatment on the survival of SNB motoneurons after target removal and concomitant axotomy early in development. On the day of birth, two target muscles of SNB motoneurons of male and female rats were bilaterally extirpated. Target removal resulted in a dramatic loss of SNB motoneurons within 48 h of surgery, with an ultimate loss of virtually all motoneurons projecting to the extirpated muscles by postnatal day 10. Treatment with testosterone failed to save SNB motoneurons from target removal-induced death. Pups treated with testosterone after target removal did not differ in the pattern or timing of motoneuron loss from untreated pups at any age examined. Counts of degenerating cells in the SNB reflected the extensive motoneuron loss and also did not differ with testosterone treatment. These results indicate that testosterone cannot save the androgen-sensitive SNB motoneurons from death after target removal and concomitant axotomy early in development. The failure of testosterone treatment to rescue SNB motoneurons in the absence of the SNB target musculature further suggests that during normal development, both androgens and target muscles are necessary for SNB motoneuron survival.     

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.     

Motoneuron development after deafferentation: II. Dorsal rhizotomy does doe block estrogen-supported growth in the dorsolateral nucleus (DLN)

The lumbar spinal cord of the rat contains two sexually dimorphic motor nuclei, the spinal nucleus of the bulbocavernosus (SNB) and the dorsolateral nucleus (DLN). Postnatally, SNB and DLN motoneurons grow substantially and reach their adult morphology by 7 weeks of age. The masculinization of SNB and DLN motoneuron dendrites depends upon steroid hormones. After early castration, the growth of SNB and DLN dendrites is markedly attenuated, but testosterone replacement restores this growth. In the SNB, initial dendritic growth is also supported in castrates treated with estrogen. By using castration and hormone replacement techniques, we examined the development of DLN motoneuron morphology in estrogen-treated castrated rats to determine if estrogen also supports the growth of DLN motoneurons. In addition, given that dorsal root ganglia may be a site of estrogen action, we tested the hypothesis that estrogen acts at primary afferents to support DLN dendritic growth. Thus, we attempted to block the potential trophic effect of estrogen by performing unilateral dorsal rhizotomies in estrogen-treated castrates. DLN motoneuron morphology was analyzed at 4 and 7 weeks of age by using cholera toxin horseradish peroxidase (BHRP) histochemistry. As found for SNB motoneurons, estrogen treatment transiently supported development. DLN motoneurons in estrogen-treated castrates developed normally through 4 weeks of age, but by 7 weeks, DLN motoneuron morphology in estrogen-treated castrates was no longer different from that in oil-treated castrates. Moreover, deafferentation via unilateral dorsal rhizotomy did not inhibit estrogen's ability to masculinize the early development of DLN motoneurons. Thus, the trophic effect of estrogen did not appear to act via the dorsal root ganglia to support the early postnatal development of DLN motoneurons.     

Hormone-sensitive periods for the control of motoneuron number and soma size in the dorsolateral nucleus of the rat spinal cord

The dorsolateral nucleus (DLN) of the rat lumbosacral spinal cord is sexually dimorphic, with males having more and larger DLN motoneurons than do females. The development of this dimorphism depends on the presence of perinatal androgens. The present study sought to determine the periods in development during which the DLN is sensitive to the masculinizing effects of the androgen testosterone propionate (TP). The size and number of DLN motoneurons in neonatally ovariectomized female rats that were exposed to TP during either the late prenatal, early postnatal, or late postnatal period were compared to control males and females. Both late prenatal and early postnatal TP injections significantly increased DLN number by 48% and 50%, respectively, but the sensitive period for TP masculinization of soma size seems to be primarily postnatal, because prenatal TP injections had little or no effect on that measure. The sensitive period for TP masculinization of DLN neuron number is similar to that of the sexually dimorphic spinal nucleus of the bulbocavernosus (SNB). However, the sensitive period for TP masculinization of DLN soma size appears to begin later than for the SNB.     

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.     

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.     

Sexual dimorphism and androgen effects on spinal motoneurons innervating the rat flexor digitorum brevis.

The foot muscle, flexor digitorum brevis, is innervated by motoneurons in the retrodorsolateral nucleus of the lumbar spinal cord in rats. We found this muscle to be sexually dimorphic, but insensitive to the anabolic or catabolic effects of androgen manipulation in adulthood: the flexor digitorum brevis was larger in adult male rats than in females, with no decrease in mass after castration in males nor any increase in size after androgen treatment of ovariectomized females. The cross-sectional area of motoneurons innervating this muscle was also sexually dimorphic, i.e., the motoneurons were larger in males. In contrast to the absence of an androgen effect on target muscle size, however, cross-sectional area of motoneurons decreased in adult males as a result of castration, and increased in adult females after androgen treatment. The dissociation of androgen effects on muscle mass and motoneuron size suggests the possibility of steroid effects upon motoneurons independent of effects upon target musculature.     

Neurogenesis of motoneurons in the sexually dimorphic spinal nucleus of the bulbocavernosus in rats

The spinal nucleus of the bulbocavernosus (SNB) consists of motoneurons innervating striated perineal muscles in male rats. The adult number of SNB motoneurons can be increased or decreased by perinatal manipulations with androgen or anti-androgen. The present results with thymidine autoradiography demonstrate that SNB motoneurons undergo their final mitosis on the fourteenth day of gestation. Because testosterone production in male rats does not begin until after gestational day 14, androgens are unlikely to affect SNB neurogenesis in normal males. By extension, hormonal manipulations more than a week later, which affect the number of SNB cells, are probably mediated by alterations in the death or specification of cells, but not their proliferation.