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 α‐receptor (CNTFRα). To test the role of CNTFRα in the response of the developing SNB system to testosterone, CNTFRα +/+ 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 CNTFRα −/− 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 CNTFRα gene deletion on muscle size in males. However, the effect of TP treatment on BC and LA muscle sizes was blunted in CNTFRα −/− 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, CNTFRα knockout mice. Prenatal treatment with testosterone did not increase SNB motoneuron number in CNTFRα −/− 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 CNTFRα −/− mice. Moreover, the CNTFRα gene deletion influences perineal muscle development and the response of the perineal muscles to testosterone. Prenatal TP treatment of CNTFRα −/− males overcomes the effects of the gene deletion on the BC and LA muscles without a concomitant effect on SNB motoneuron number. © 1999 John Wiley & Sons, Inc. J Neurobiol 41: 317–325, 1999     

The spinal nucleus of the bulbocavernosus: firsts in androgen-dependent neural sex differences

Cell number in the spinal nucleus of the bulbocavernosus (SNB) of rats was the first neural sex difference shown to differentiate under the control of androgens, acting via classical intracellular androgen receptors. SNB motoneurons reside in the lumbar spinal cord and innervate striated muscles involved in copulation, including the bulbocavernosus (BC) and levator ani (LA). SNB cells are much larger and more numerous in males than in females, and the BC/LA target muscles are reduced or absent in females. The relative simplicity of this neuromuscular system has allowed for considerable progress in pinpointing sites of hormone action, and identifying the cellular bases for androgenic effects. It is now clear that androgens act at virtually every level of the SNB system, in development and throughout adult life. In this review we focus on effects of androgens on developmental cell death of SNB motoneurons and BC/LA muscles; the establishment and maintenance of SNB motoneuron soma size and dendritic length; BC/LA muscle morphology and physiology; and behaviors controlled by the SNB system. We also describe new data on neurotherapeutic effects of androgens on SNB motoneurons after injury in adulthood.     

Castration reduces motoneuron soma size but not dendritic length in the spinal nucleus of the bulbocavernosus of wild-type and BCL-2 overexpressing mice

Motoneurons in the spinal nucleus of the bulbocavernosus (SNB) and their target muscles, bulbocavernosus and levator ani (BC/LA), constitute an androgen-sensitive neuromuscular system. Testosterone regulates SNB soma size, SNB dendritic length, and BC/LA muscle mass in adult male rats. Recent evidence indicates that the cell death-regulatory protein, Bcl-2, may also play a role in adult neural plasticity. The present study examined whether gonadal hormones and/or the Bcl-2 protein influence the morphology of the SNB neuromuscular system in adult B6D2F1 mice. In Experiment 1, adult wild-type and Bcl-2 overexpressing males were castrated and implanted with silastic capsules containing testosterone or left blank. Six weeks after castration, cholera toxin-horseradish peroxidase was injected into the BC muscle to label SNB dendrites. Animals were killed 48 h later, and BC/LA muscle mass, SNB soma size, and SNB dendritic arbors were examined. In Experiment 2, wild-type and Bcl-2 overexpressing males were castrated or sham castrated, implanted with testosterone-filled or blank capsules, and examined 12 weeks later. In both experiments, BC/LA muscle mass and SNB soma size were significantly reduced in castrates receiving blank capsules. Surprisingly, however, there was no effect of hormone manipulation on any of several measures of dendritic length. Thus, the dendritic morphology of SNB motoneurons appears to be relatively insensitive to circulating androgen levels in B6D2F1 mice. Bcl-2 overexpression did not influence BC/LA muscle mass, SNB soma size, or SNB dendritic length, indicating that the morphology of this neuromuscular system and the response to castration are not altered by forced expression of the Bcl-2 protein.     

Evidence for androgen receptors in sexually dimorphic perineal muscles of neonatal male rats. Absence of androgen accumulation by the perineal motoneurons

During development, survival of the sexually dimorphic spinal nucleus of the bulbocavernosus (SNB) and its target perineal muscles, the bulbocavernosus (BC) and the levator ani (LA) is androgen-dependent. To define androgen's site of action in masculinizing SNB system structures, we examined whether or not androgen receptors are present in SNB motoneurons and/or BC/LA muscles of neonatal male rats. Using a receptor binding assay, we have identified androgen-binding factors in the neonatal BC/LA (Bmax = 13.5 fmol/mg protein; Kd = 4.69 nM) for the first time. In contrast, androgen autoradiography provided no evidence that neonatal spinal motoneurons accumulate androgens. These results support the hypothesis that BC/LA muscles are a primary site of androgen action for masculinizing SNB system structures, and that androgen need not interact with SNB motoneurons directly to sexually differentiate them.     

Castration reduces motoneuron soma size but not dendritic length in the spinal nucleus of the bulbocavernosus of wild‐type and BCL‐2 overexpressing mice

Motoneurons in the spinal nucleus of the bulbocavernosus (SNB) and their target muscles, bulbocavernosus and levator ani (BC/LA), constitute an androgen‐sensitive neuromuscular system. Testosterone regulates SNB soma size, SNB dendritic length, and BC/LA muscle mass in adult male rats. Recent evidence indicates that the cell death‐regulatory protein, Bcl‐2, may also play a role in adult neural plasticity. The present study examined whether gonadal hormones and/or the Bcl‐2 protein influence the morphology of the SNB neuromuscular system in adult B6D2F1 mice. In Experiment 1, adult wild‐type and Bcl‐2 overexpressing males were castrated and implanted with silastic capsules containing testosterone or left blank. Six weeks after castration, cholera toxin‐horseradish peroxidase was injected into the BC muscle to label SNB dendrites. Animals were killed 48 h later, and BC/LA muscle mass, SNB soma size, and SNB dendritic arbors were examined. In Experiment 2, wild‐type and Bcl‐2 overexpressing males were castrated or sham castrated, implanted with testosterone‐filled or blank capsules, and examined 12 weeks later. In both experiments, BC/LA muscle mass and SNB soma size were significantly reduced in castrates receiving blank capsules. Surprisingly, however, there was no effect of hormone manipulation on any of several measures of dendritic length. Thus, the dendritic morphology of SNB motoneurons appears to be relatively insensitive to circulating androgen levels in B6D2F1 mice. Bcl‐2 overexpression did not influence BC/LA muscle mass, SNB soma size, or SNB dendritic length, indicating that the morphology of this neuromuscular system and the response to castration are not altered by forced expression of the Bcl‐2 protein. © 2002 Wiley Periodicals, Inc. J Neurobiol 53: 403–412, 2002     

Seasonal variation in mammalian striated muscle mass and motoneuron morphology

Feral white-footed mice are seasonal breeders that undergo predictable cycles of reproductive function. Photoperiod-induced fluctuations in gonadal function of white-footed mice were associated with morphological changes in perineal muscles and their motoneurons. Exposure to short daylengths resulted in testicular regression, decreased perineal muscle mass, and shrinkage of somata and nuclei of motoneurons of the spinal nucleus of the bulbocavernosus (SNB). These effects were reversed by reinstatement of long daylengths. Similar reductions in muscle mass and SNB soma size were seen following gonadectomy of white-footed mice. In addition, dendritic trees of SNB motoneurons were reduced in gonadectomized mice compared with dendritic arbors of intact mice or castrates provided with testosterone capsules. Androgen-mediated annual changes in muscle mass and motoneuron morphology appear to be a natural part of this species' physiology.     

Sexual dimorphism and androgen regulation of satellite cell population in differentiating rat levator ani muscle

The bulbocavernosus (BC) and levator ani (LA) muscles of rats show remarkable androgen-dependent sexual dimorphism. These muscles are additionally of interest because they are thought to indirectly mediate sexual differentiation of innervating spinal motoneurons. This sexual differentiation of the BC/LA is thought to be due to an increase in muscle units in the male rat during the first week after birth. We examined the cellular basis of this differentiation by studying satellite cells in the LA of postnatal day 2.5 rats, when sexual dimorphism is already prominent. Two experiments were performed in which LA satellite cells were measured: (1) wild-type (WT) males were compared with females and to Tfm androgen receptor mutant males, which are androgen insensitive despite producing masculine amounts of testosterone, and (2) females treated prenatally and/or postnatally with testosterone proprionate were compared with females receiving vehicle injections. Our results indicate that WT males have a larger LA and a greater number of satellite cells in the LA muscle than females or Tfm males. However, satellite cell density was similar for all three groups. Prenatal testosterone treatment masculinized LA size and resulted in a corresponding increase in satellite cell populations, while postnatal TP treatment resulted in a tendency for increased satellite cell density without a significant increase in LA size. Taken together, these studies indicate that satellite cells in the neonatal LA muscle are sexually dimorphic, and that this dimorphism likely results from perinatal actions of androgens on androgen receptors.     

Maintenance of the spinal nucleus of the bulbocavernosus neuromuscular system is not influenced by physiological levels of glucocorticoids

The spinal nucleus of the bulbocavernosus (SNB) neuromuscular system mediates sexual reflexes, and is highly sexually dimorphic in rats. While maintenance of this system in adulthood is mainly dependent on androgens, there is also evidence to suggest that glucocorticoids may have a catabolic effect. We conducted a series of studies to fully examine the influence of basal glucocorticoids on the size of the SNB motoneurons and the associated bulbocavernosus (BC) and levator ani (LA) muscles. Specifically, we examined whether the muscles and motoneurons of the SNB neuromuscular system are affected by: (1) blockade of endogenous glucocorticoids via delivery of the antagonist RU-486 at doses ranging from low to high, (2) removal of endogenous glucocorticoids via adrenalectomy, or (3) restoration of physiological corticosterone levels via implants following adrenalectomy. In each study, we found that muscle and motoneuron size were unaffected by glucocorticoid manipulation. In contrast to previous results with supraphysiological levels of glucocorticoids, our results indicate that basal, nonstress levels of glucocorticoids do not influence the size of the BC/LA muscles or their associated SNB motoneurons.     

Androgen action at the target musculature regulates brain‐derived neurotrophic factor protein in the spinal nucleus of the bulbocavernosus

We have previously demonstrated that brain‐derived neurotrophic factor (BDNF) interacts with testosterone to regulate dendritic morphology of motoneurons in the highly androgen‐sensitive spinal nucleus of the bulbocavernosus (SNB). Additionally, in adult male rats testosterone regulates BDNF in SNB motoneurons and its target muscle, the bulbocavernosus (BC). Because BDNF is retrogradely transported from skeletal muscles to spinal motoneurons, we hypothesized that testosterone could regulate BDNF in SNB motoneurons by acting locally at the BC muscle. To test this hypothesis, we restricted androgen manipulation to the SNB target musculature. After castration, BDNF immunolabeling in SNB motoneurons was maintained at levels similar to those of gonadally intact males by delivering testosterone treatment directly to the BC muscle. When the same implant was placed interscapularly in castrated males it was ineffective in supporting BDNF immunolabeling in SNB motoneurons. Furthermore, BDNF immunolabeling in gonadally intact adult males given the androgen receptor blocker hydroxyflutamide delivered directly to the BC muscle was decreased compared with that of gonadally intact animals that had the same hydroxyflutamide implant placed interscapularly, or when compared with castrated animals that had testosterone implants at the muscle. These results demonstrate that the BC musculature is a critical site of action for the androgenic regulation of BDNF in SNB motoneurons and that it is both necessary and sufficient for this action. Furthermore, the local action of androgens at the BC muscle in regulating BDNF provides a possible mechanism underlying the interactive effects of testosterone and BDNF on motoneuron morphology. © 2013 Wiley Periodicals, Inc. Develop Neurobiol 73: 587–598, 2013     

Sexually dimorphic neuron number in lumbosacral dorsal root ganglia of the rat: Development and steroid regulation

Rats possess a sexually dimorphic neuromuscular system that controls penile reflexes critical for copulation. This system includes two motor nuclei in the lumbar cord and their target musculature in the perineum. The spinal nucleus of the bulbocavernosus (SNB) and the dorsolateral nucleus (DLN) motoneuron populations and their target perineal muscles are much larger in males than in females. The sex difference in motoneuron number develops via androgen-regulated differential cell death during the perinatal period; androgen also regulates retention of the target muscles. The developmental pattern and steroid sensitivity of peripheral afferents to the SNB/DLN motor nuclei were previously unknown. In order to characterize the peripheral sensory component of the dimorphic SNB/DLN system, the neurons of the relevant dorsal root ganglia (DRGs) were quantified in terms of number, size, and androgen sensitivity at various perinatal ages. DRG neuron number is greatest prenatally, then decreases in both sexes after birth; the timing and pattern of neuron number development are similar to those seen in the SNB and DLN. Postnatally, males have more DRG neurons than females, as a result of greater neuron death in the DRGs of females. Females treated with testosterone propionate during the perinatal period exhibit masculine development of DRG neuron number. Thus, the normal development of DRG neuron number parallels that of the SNB/DLN motor nuclei and target muscles in pattern and timing, is sexually dimorphic, and is regulated by androgen. © 1993 John Wiley & Sons, Inc.     

The role of cell death in sexually dimorphic muscle development: male-specific muscles are retained in female bax/bak knockout mice

The bulbocavernosus (BC) and levator ani (LA) muscles are present in males but absent or severely reduced in females, and the fate of these muscles controls the survival of motoneurons in the sexually dimorphic spinal nucleus of the bulbocavernosus. However, the mechanism underlying the sex difference in BC and LA development has been controversial. We examined the role of cell death in sexual differentiation of the bulbocavernosus BC/LA muscles in mice. Muscle development was mapped from embryonic day 16 (E16) to postnatal day 5 (P5). A sex difference (male>female) first arose on E17 (BC) or E18 (LA), and increased in magnitude postnatally. TUNEL labeling revealed dying cells in the BC and LA muscles of both sexes perinatally. However, females had a significantly higher density of TUNEL-positive cells than did males. A role for the proapoptotic factors, Bax and Bak, in BC/LA development was tested by examining mice lacking one or both of these proteins. In females lacking either Bax or Bak, the BC was absent and the LA rudimentary. Deletion of both bax and bak genes, however, rescued the BC, increased LA size approximately 20-fold relative to controls, and virtually eliminated TUNEL-positive cells in both muscles. We conclude that cell death plays an essential role in sexual differentiation of the BC/LA muscles. The presence of either Bax or Bak is sufficient for cell death in the BC/LA, whereas the absence of both prevents sexually dimorphic muscle cell death.     

Perineal muscles and motoneurons are sexually monomorphic in the naked mole-rat (Heterocephalus glaber)

Naked mole-rats are eusocial mammals that live in colonies with a single breeding female and one to three breeding males. All other members of the colony, known as subordinates, are nonreproductive and exhibit few sex differences in behavior or genital anatomy. This raises questions about the degree of sexual differentiation in subordinate naked mole-rats. The striated perineal muscles associated with the phallus [the bulbocavernosus (BC), ischiocavernosus (IC), and levator ani (LA) muscles], and their innervating motoneurons, are sexually dimorphic in all rodents examined to date. We therefore asked whether perineal muscles and motoneurons were also sexually dimorphic in subordinate naked mole-rats. Muscles similar to the LA and IC of other rodents were found in naked mole-rats of both sexes. No clear BC muscle was identified, although a large striated muscle associated with the urethra in male and female naked mole-rats may be homologous to the BC of other rodents. There were no sex differences in the volumes of the LA, IC, or the urethral muscles. Motoneurons innervating the perineal muscles were identified by retrograde labeling with cholera-toxin-conjugated horseradish peroxidase. All perineal motoneurons were found in a single cluster in the ventrolateral lateral horn, in a position similar to that of Onuf's nucleus of carnivores and primates. There was no sex difference in the size or number of motoneurons in Onuf's nucleus of naked mole-rats. Thus, unlike findings in any other mammal, neither the perineal muscles nor the perineal motoneurons appear to be sexually differentiated in subordinate naked mole-rats.     

Perineal muscles and motoneurons are sexually monomorphic in the naked mole‐rat (Heterocephalus glaber)

Naked mole‐rats are eusocial mammals that live in colonies with a single breeding female and one to three breeding males. All other members of the colony, known as subordinates, are nonreproductive and exhibit few sex differences in behavior or genital anatomy. This raises questions about the degree of sexual differentiation in subordinate naked mole‐rats. The striated perineal muscles associated with the phallus [the bulbocavernosus (BC), ischiocavernosus (IC), and levator ani (LA) muscles], and their innervating motoneurons, are sexually dimorphic in all rodents examined to date. We therefore asked whether perineal muscles and motoneurons were also sexually dimorphic in subordinate naked mole‐rats. Muscles similar to the LA and IC of other rodents were found in naked mole‐rats of both sexes. No clear BC muscle was identified, although a large striated muscle associated with the urethra in male and female naked mole‐rats may be homologous to the BC of other rodents. There were no sex differences in the volumes of the LA, IC, or the urethral muscles. Motoneurons innervating the perineal muscles were identified by retrograde labeling with cholera‐toxin‐conjugated horseradish peroxidase. All perineal motoneurons were found in a single cluster in the ventrolateral lateral horn, in a position similar to that of Onuf's nucleus of carnivores and primates. There was no sex difference in the size or number of motoneurons in Onuf's nucleus of naked mole‐rats. Thus, unlike findings in any other mammal, neither the perineal muscles nor the perineal motoneurons appear to be sexually differentiated in subordinate naked mole‐rats. © 2002 Wiley Periodicals, Inc. J Neurobiol 51: 33–42, 2002     

Short day lengths delay development of the SNB neuromuscular system in the Siberian hamster,Phodopus sungorus

The Siberian hamster, Phodopus sungorus, breeds seasonally. In the laboratory, the seasonal breeding can be controlled by photoperiod, which affects the durations of nightly melatonin secretions. Winterlike short day lengths induce gonadal regression in adult animals, and pups born and maintained in short days undergo gonadal development much later than animals born into long days. The spinal nucleus of the bulbocavernosus (SNB) and its target muscles, the bulbocavernosus (BC) and levator ani (LA), comprise a sexually dimorphic, androgen-sensitive neuromuscular system involved in male reproduction. The SNB neuromuscular system was studied in male Siberian hamsters maintained from conception in short-day (8:16 h light/dark cycle) versus long-day (16:8 h light/dark cycle) conditions. At 40–47 days of age, development of three components of the SNB neuromuscular system were all significantly delayed in hamsters raised in the short photoperiod: BC/LA muscle weight, the size of SNB motoneuronal somata, and the area of the neuromuscular junctions at the BC/LA muscles of short-day hamsters were each significantly reduced relative to those of long-day counterparts. Thus, development of the SNB reproductive system is delayed under short day lengths in this species. © 1998 John Wiley & Sons, Inc. J Neurobiol 35: 355–360, 1998     

Hormonal control of neuron number in sexually dimorphic spinal nuclei of the rat: IV. Masculinization of the spinal nucleus of the bulbocavernosus with testosterone metabolites

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 females. This sexual dimorphism is due to the presence of androgen during development; females treated with testosterone (T) perinatally have a masculine SNB system. To assess whether masculinization of the SNB could involve the conversion of testosterone into its active metabolites, dihydrotestosterone (DHT) and estrogen, we examined the development of the SNB in females treated perinatally with estrogen alone or in combination with dihydrotestosterone. Counts of motoneurons in the developing SNB in all groups showed the typical prenatal increase followed by a differential postnatal decline; the incidence of degenerating cells reflected this decline. Motoneuron numbers and the frequency of degenerating cells in females treated with estrogen (E) alone did not differ from those of normal females, with both groups losing large numbers of motoneurons and having a high incidence of degenerating cells. In contrast, females treated with both estrogen and dihydrotestosterone did not show the female-typical decline in motoneuron number and had a low, masculine incidence of degenerating cells. By postnatal day 10, females treated with estrogen and dihydrotestosterone had a fully masculine SNB motoneuron number, suggesting that dihydrotestosterone alone or in conjunction with estrogen may be involved in the development of the sexually dimorphic SNB system.     

Regulation of motoneuron death in the spinal nucleus of the bulbocavernosus.

A sexual dimorphism in the number of motoneurons in the spinal nucleus of the bulbocavernosus (SNB) of rats is engendered by a sex difference in ontogenetic cell death. Testicular secretions, specifically androgenic steroids, reduce SNB motoneuron death in males. The fate of the target muscles generally mirrors that of the motoneurons, and androgens appear to exert their effects upon the target muscles, sparing the motoneurons as a secondary consequence. Treatment with ciliary neurotrophic factor can also spare SNB motoneurons in newborn females, raising the possibility that this factor normally mediates androgen's effect upon motoneuron survival. The ontogeny of calcitonin gene-related peptide immunoreactivity is delayed in SNB cells compared with other motoneurons and is further delayed in the SNB cells of females. In both sexes, calcitonin gene-related peptide is detected after the period of SNB motoneuron death is complete. A sex difference in motoneuron number is also seen in the human homologue of the SNB and, because ontogenetic death of motoneurons in humans overlaps the period of androgen secretion, may arise in a manner similar to that in the rat SNB.     

Cellular analyses of hormone influence on motoneuronal development and function

The striated bulbocavernosus (BC) muscles of the rodent perineum are innervated by motoneurons in the spinal nucleus of the bulbocavernosus (SNB). In adulthood, the BC muscles are present in males only. However, newborn female rats have BC muscles, and SNB cells have made both anatomical and functional contact with them. Nevertheless, both motoneurons and muscles will degenerate unless androgens are administered perinatally. Such androgen treatment appears to be acting primarily on the BC muscles themselves, since the muscles are spared by androgen even after the loss of supraspinal neural afferents or even the entire lumbosacral spinal cord. Furthermore, androgen can spare SNB motoneurons that are themselves androgen insensitive. Perinatal steroid treatments can also alter the final spinal location of SNB cells as determined by retrograde tracing studies. Androgen continues to modify the morphology of the SNB system in adulthood, altering the size of both motoneurons and targets, which may be important for the reproductive function of BC muscles. Finally, the sexually dimorphic character of motoneuronal groups innervating perineal muscles seems to be common in mammals, since the homologue of the SNB, Onuf's nucleus, has more cells in males than in females in both dogs and humans.     

Adult plasticity in hormone-sensitive motoneuron morphology: methodological/behavioral confounds

Changes in androgen levels can alter the structure of motoneurons in the spinal nucleus of the bulbocavernosus (SNB), a motor nucleus that innervates perineal muscles involved in copulatory behavior. While sexual activity can alter androgen levels in normal males, it has no effect on SNB motoneuron soma size or dendritic morphology (Beversdorf, Kurz, and Sengelaub, 1990). However, Breedlove (1997) reported reductions in the size of SNB somata, nuclei, and target muscles of copulating versus noncopulating castrated rats maintained on subphysiological testosterone. To reconcile the results obtained using intact versus implant paradigms, we tested the hypothesis that the implant/behavior paradigm could produce differences in hormone levels, potentially confounding sexual behavior effects on the morphology of this androgen-sensitive neuromuscular system. Young adult male rats were castrated and immediately given 5-mm Silastic implants containing crystalline testosterone. One week later, blood samples were drawn and the males were housed with receptive females (copulators) or nonreceptive females (noncopulators) or housed alone (singles). After 27 days, blood samples were drawn again, and SNB target muscles and spinal cords removed. No differences in target muscle weight or SNB somata and nuclei size were observed between copulators, noncopulators, or singles; as expected, all measures were significantly reduced relative to intact males. Radioimmunoassay showed that testosterone declined differentially over the course of the behavioral manipulation across groups, being greatest in copulators and least pronounced in single males. These data indicate that differences in sexual or housing experience can alter testosterone titers under these implant conditions, potentially confounding hormone-sensitive measures of morphology.     

Regulation of motoneuron death in the spinal nucleus of the bulbocavernsus

A sexual dimorphism in the number of motoneurons in the spinal nucleus of the bulbocavernosus (SNB) of rats is engendered by a sex difference in ontogenetic cell death. Testicular secretions, specifically androgenic steroids, reduce SNB motoneuron death in males. The fate of the target muscles generally mirrors that of the motoneurons, and androgens appear to exert their effects upon the target muscles, sparing the motoneurons as a secondary consequence. Treatment with ciliary neurotrophic factor can also spare SNB motoneurons in newborn females, raising the possibility that this factor normally mediates androgen's effect upon motoneuron survival. The ontogeny of calcitonin gene-related peptide immunoreactivity is delayed in SNB cells compared with other motoneurons and is further delayed in the SNB cells of females. In both sexes, calcitonin gene-related peptide is detected after the period of SNB motoneuron death is complete. A sex difference in motoneuron number is also seen in the human homologue of the SNB and, because ontogenetic death of motoneurons in humans overlaps the period of androgen secretion, may arise in a manner similar to that in the rat SNB. © 1992 John Wiley & Sons, Inc.     

Aromatase inhibition reduces dendritic growth in a sexually dimorphic rat spinal nucleus

The rat lumbar spinal cord contains the steroid-sensitive spinal nucleus of the bulbocavernosus (SNB), whose motoneurons innervate perineal muscles involved in copulatory reflexes. In normal males, SNB motoneuron dendrites grow exuberantly through postnatal (P) day 28. This growth is steroid dependent: Dendrites fail to grow in males castrated at P7, but grow normally in castrates treated with testosterone or its metabolites, dihydrotestosterone combined with estrogen. Treatment with either metabolite alone supports dendritic growth, but not to the level of testosterone-treated or intact males. In this study, we tested the hypothesis that aromatization of androgens to estrogens was involved in the masculine development of SNB dendrites. Motoneuron morphology was assessed in normal males and males treated daily (P7-28) with fadrozole, a potent aromatase inhibitor (0.25 mg/kg, subcutaneously) or saline vehicle (n = 4-6/group). SNB motoneurons were retrogradely labeled with cholera toxin-horseradish peroxidase at P28 (when dendritic length is normally maximal) and reconstructed in three dimensions. Comparable labeling was seen across groups; it was equivalent in both the rostrocaudal and radial extents. However, dendritic lengths in fadrozole-treated males were significantly below those of intact or saline-treated males. Neither SNB somata size nor target muscle weight differed across groups. These results suggest that aromatization of androgens to estrogens is necessary for development of masculine SNB dendritic morphology.