Interaction of BDNF and testosterone in the regulation of adult perineal motoneurons

In androgen-sensitive motoneurons of the spinal nucleus of the bulbocavernosus (SNB), we investigated the interaction of BDNF (brain-derived neurotrophic factor) and testosterone to understand whether each factor gates the ability of the other to regulate androgen receptor expression and soma size, and whether each factor requires the presence of the other for its action. We axotomized SNB motoneurons and applied BDNF or PBS (phosphate-buffered saline) to the cut ends of the axons in rats that were castrated and treated with either testosterone or placebo. Control groups were either not castrated or not axotomized, or had intact SNB axons and were castrated and treated with testosterone or placebo. We found that testosterone determined the expression of nuclear androgen receptor, and this effect was enhanced by both BDNF and contact with the target muscles. The effect of BDNF on androgen receptor expression was seen only when testosterone was present. In the regulation of soma size, BDNF dominated. The application of BDNF completely compensated for the loss of testosterone in castrated males so that the testosterone effect on soma size was seen only in intact SNB motoneurons and in axotomized motoneurons treated with PBS. Moreover, testosterone increased androgen receptor and soma size in axotomized SNB motoneurons, indicating that testosterone can act on sites other than the target muscles of the SNB to regulate each of these. These results indicate that the regulation of androgen receptor by testosterone does not require BDNF, but the regulation of androgen receptor by BDNF does require testosterone. The regulation of soma size by BDNF does not require high expression of nuclear androgen receptor.     

Distribution of androgen receptor immunoreactivity in the spinal cord of wild-type,androgen-insensitive and gonadectomized male rats

The polyclonal antiserum PG21 was used to detect androgen receptor (AR) protein in three motoneuronal pools of the male rat lumbar spinal cord. In gonadally intact, wild-type males, the spinal nucleus of the bulbocavernosus (SNB), dorsolateral nucleus (DLN), and retrodorsolateral nucleus (RDLN) all displayed immunolabeling of cell nuclei. The percentage of motoneurons displaying such labeling was highest in the SNB and lowest in the RDLN. This pattern of AR immunocytochemical labeling agrees well with previous steroid autoradiographic studies of androgen accumulation in the rat spinal cord. In contrast, virtually no motoneurons in any of the three pools displayed nuclear AR immunostaining in long-term gonadectomized males or in gonadally intact males carrying the Tfm mutation, which renders the AR incompetent. In gonadectomized males, labeling was restored in the SNB and DLN, but not the RDLN, 30 min after an injection of replacement testosterone. Eight hours of testosterone exposure restored immunolabeling in all three motor nuclei. Apparent cytoplasmic staining was seen in SNB motoneurons of untreated castrates and Tfm rats, but not intact rats, suggesting that AR residing in the cytoplasm translocates to the nucleus on binding to androgen in these motoneurons. © 1995 John Wiley & Sons, Inc.     

Estrogenic support of motoneuron dendritic growth via the neuromuscular periphery in a sexually dimorphic motor system

The lumbar spinal cord of rats contains the sexually dimorphic, steroid-sensitive spinal nucleus of the bulbocavernosus (SNB). In males, the growth of SNB dendrites is steroid-dependent: dendrites fail to grow after castration, but grow in castrates treated with androgens or estrogens. Blocking estradiol synthesis or estrogen receptors in gonadally intact males attenuates SNB dendritic growth, suggesting that estrogens are required and must be able to act at their receptors to support normal masculine dendritic growth. However, SNB motoneurons do not accumulate estrogens, suggesting that estrogens act indirectly to support SNB dendritic growth. In this experiment, we examined whether local estrogen action in the neuromuscular periphery was involved in the postnatal development of SNB motoneurons. Motoneuron morphology was assessed in gonadally intact and castrated males. Gonadally intact males were left untreated or given either blank or tamoxifen implants sutured to the target musculature, or tamoxifen interscapular implants. Castrated males were left untreated or were given estradiol by muscle or interscapular implants or systemic injection during the period of SNB dendritic growth. At postnatal day 28, when SNB dendritic length is normally maximal, SNB motoneurons were retrogradely labeled with cholera toxin-HRP and reconstructed in three dimensions. While interscapular tamoxifen implants were ineffective, blocking estrogen receptors at the target musculature resulted in attenuation of SNB dendritic growth. In contrast, while interscapular implants of estradiol were ineffective, local treatment with estradiol at the target musculature in castrated males resulted in masculinization of dendritic growth. Thus, estrogens may act by an indirect action in the neuromuscular periphery to support SNB dendritic growth.     

Testosterone metabolites differentially maintain adult morphology in a sexually dimorphic neuromuscular system

The lumbar spinal cord of rats contains the sexually dimorphic, steroid‐sensitive spinal nucleus of the bulbocavernosus (SNB). Androgens are necessary for the development of the SNB neuromuscular system, and in adulthood, continue to influence the morphology and function of the motoneurons and their target musculature. However, estrogens are also involved in the development of the SNB system, and are capable of maintaining function in adulthood. In this experiment, we assessed the ability of testosterone metabolites, estrogens and nonaromatizable androgens, to maintain neuromuscular morphology in adulthood. Motoneuron and muscle morphology was assessed in adult normal males, sham‐castrated males, castrated males treated with testosterone, dihydrotestosterone, estradiol, or left untreated, and gonadally intact males treated with the 5α‐reductase inhibitor finasteride or the aromatase inhibitor fadrozole. After 6 weeks of treatment, SNB motoneurons were retrogradely labeled with cholera toxin‐HRP and reconstructed in three dimensions. Castration resulted in reductions in SNB target muscle size, soma size, and dendritic morphology. Testosterone treatment after castration maintained SNB soma size, dendritic morphology, and elevated target muscle size; dihydrotestosterone treatment also maintained SNB dendritic length, but was less effective than testosterone in maintaining both SNB soma size and target muscle weight. Treatment of intact males with finasteride or fadrozole did not alter the morphology of SNB motoneurons or their target muscles. In contrast, estradiol treatment was completely ineffective in preventing castration‐induced atrophy of the SNB neuromuscular system. Together, these results suggest that the maintenance of adult motoneuron or muscle morphology is strictly mediated by androgens. © 2009 Wiley Periodicals, Inc. Develop Neurobiol 70: 206–221, 2010.     

Castration‐induced upregulation of muscle ERα supports estrogen sensitivity of motoneuron dendrites in a sexually dimorphic neuromuscular system

The spinal cord of rats contains the sexually dimorphic motoneurons of the spinal nucleus of the bulbocavernosus (SNB). In males, SNB dendrites fail to grow after castration, but androgen or estrogen treatment supports dendritic growth in castrated males. Estrogenic support of SNB dendrite growth is mediated by estrogen receptors (ER) in the target muscle. ERα expression in cells lacking a basal lamina (referred to as “extra‐muscle fiber cells”) of the SNB target musculature coincides with the period of estrogen‐dependent SNB dendrite growth. In the SNB target muscle, extra‐muscle fiber ERα expression declines with age and is typically absent after postnatal (P) day 21 (P21). Given that estradiol downregulates ERα in skeletal muscle, we tested the hypothesis that depleting gonadal hormones would prevent the postnatal decline in ERα expression in the SNB target musculature. We castrated male rats at P7 and assessed ERα immunolabeling at P21; ERα expression was significantly greater in castrated males compared with normal animals. Because ERα expression in SNB target muscles mediates estrogen‐dependent SNB dendrogenesis, we further hypothesized that the castration‐induced increase in muscle ERα would heighten the estrogen sensitivity of SNB dendrites. Male rats were castrated at P7 and treated with estradiol from P21 to P28; estradiol treatment in castrates resulted in dendritic hypertrophy in SNB motoneurons compared with normal males. We conclude that early castration results in an increase in ERα expression in the SNB target muscle, and this upregulation of ERα supports estrogen sensitivity of SNB dendrites, allowing for hypermasculinization of SNB dendritic arbors. © 2013 Wiley Periodicals, Inc. Develop Neurobiol 73: 921–935, 2013     

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     

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.     

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.     

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.     

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.     

Critical period for estrogen‐dependent motoneuron dendrite growth is coincident with ERα expression in target musculature

The spinal cord of rats contains the sexually dimorphic, steroid‐sensitive motoneurons of the spinal nucleus of the bulbocavernosus (SNB). In males, SNB dendrite growth is dependent on gonadal steroids: dendrite growth is inhibited after castration, but supported in androgen‐ or estrogen‐treated castrated males. Furthermore, estrogenic support of SNB dendrite growth is mediated by estrogen action at the target musculature, inhibited by estrogen receptor (ER) blockade at the muscle and supported by local estradiol treatment. However, this estrogenic support is restricted to the early postnatal period, after which the morphology of SNB dendrites is insensitive to estrogens. To test if the developmentally restricted effects of estrogens on SNB dendrite growth coincide with the transient expression of ER in the target musculature, ERα expression was assessed during development and in adulthood. ERα expression in extra‐Muscle fiber cells was greatest from postnatal day 7 (P7) to P14 and declined after P21. Because this pattern of ERα expression coincided with the period of estrogen‐dependent dendrite growth, we tested if limiting hormone exposure to the period of maximal ERα expression in extra‐muscle fiber cells could fully support estrogen‐dependent SNB dendrite growth. We restricted estradiol treatment in castrated males from P7 to P21 and assessed SNB dendritic morphology at P28. Treating castrates with estradiol implants at the muscle from P7 to P21 supported dendrite growth to normal levels through P28. These data suggest that the transient ERα expression in target muscle could potentially define the critical period for estrogen‐dependent dendrite growth in SNB motoneurons. © 2011 Wiley Periodicals, Inc. Develop Neurobiol, 2013     

Exogenous testosterone prevents motoneuron atrophy induced by contralateral motoneuron depletion

Gonadal steroids exhibit neuroprotective and neurotherapeutic effects. The lumbar spinal cord of male rats contains a highly androgen-sensitive population of motoneurons, the spinal nucleus of the bulbocavernosus (SNB), whose morphology and function are dependent on testosterone in adulthood. Unilateral SNB motoneuron depletion induces dendritic atrophy in contralateral SNB motoneurons, but this atrophy is reversed in previously castrated males treated with testosterone. In the present experiment we test the hypothesis that the morphology of SNB motoneurons is protected from atrophy after contralateral motoneuron depletion by exogenous testosterone alone (i.e., with no delay between castration and testosterone replacement). We unilaterally depleted SNB motoneurons by intramuscular injection of cholera toxin conjugated saporin. Simultaneously, some saporin-injected rats were castrated and immediately given replacement testosterone. Four weeks later, contralateral SNB motoneurons were labeled with cholera toxin conjugated HRP, soma sizes were measured, and dendritic arbors were reconstructed. Contralateral SNB motoneuron depletion induced somal atrophy and dendritic retraction, but testosterone treatment prevented both of these effects. Thus, the presence of high-normal levels of testosterone prevents motoneuron atrophy induced by contralateral motoneuron depletion. These data support a therapeutic role for testosterone in preventing atrophy induced by motoneuron injury.     

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.     

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.     

Estrogen alters excitability but not morphology of a sexually dimorphic neuromuscular system in adult rats

In rats, motoneurons of the spinal nucleus of the bulbocavernosus (SNB) innervate the bulbocavernosus (BC) muscle, which surrounds the base of the penis. The SNB/BC is a sexually dimorphic, steroid-sensitive neuromuscular system, which is critically important in male reproductive behavior. Androgens are necessary for the development, morphology, and function of the SNB/BC system. However, estradiol (E) is also necessary for the development of the SNB/BC system, and E is capable of maintaining BC EMG activity in adulthood. In this study, we used electrophysiological and anatomical methods to examine estrogenic effects on BC EMG activity. We used a modified H-reflex testing method to investigate polysynaptic reflex characteristics in intact males, castrates, and castrates treated short term with estradiol benzoate (EB). Measures of EMG activity, response latency, and spike count were altered in castrates, but maintained in EB-treated castrates to the levels of intact males. Furthermore, estrogenic effects were found in EMG activity that could be isolated to the periphery of the SNB/BC system. BC NMJ size and muscle fiber area have been demonstrated to be hormone sensitive, and we examined these for possible correlates of E's effects on BC EMG activity. BC muscles of intact males, castrates, and short-term EB-treated castrates were fixed and stained with zinc iodide and osmium tetroxide. NMJ size and muscle fiber area did not differ between groups. Together, these data suggest that E treatment results in changes in the neuromuscular periphery that maintain BC EMG activity, but this effect cannot be accounted for by changes in NMJ size or muscle fiber area.     

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     

Hormonally mediated plasticity of motoneuron morphology in the adult rat spinal cord: A cholera toxin-HRP study

The dorsolateral nucleus (DLN) and the spinal nucleus of the bulbocavernosus (SNB) of the rat lumbar spinal cord are sexually dimorphic groups of motoneurons that innervate striated perineal muscles involved in male copulatory behavior. Androgens control the development of these motoneurons and their target muscles, and continue to influence the system in adulthood. Given that several features of SNB motoneuron morphology have been shown to be androgen sensitive in adult male rats, we examined the effects of androgen manipulations on the morphology of motoneurons in the DLN in adult rats. Adult male rats were castrated and implanted with testosterone-filled or blank implants, or were subjected to a sham-castration procedure. Six weeks after treatment, motoneurons in the DLN were retrogradely labeled with cholera toxin-horseradish peroxidase (HRP) after injection into the ischiocavernosus (IC) muscle and their morphology assessed. Measures of the radial extent and coverage of the dendritic arbor of DLN motoneurons projecting to the IC (DLN-IC motoneurons) were similar across the groups, indicating comparable degrees of HRP transport. However, DLN-IC motoneurons in castrates with blank implants possessed both shorter dendritic lengths and smaller somas than those of castrates treated with testosterone. Castrates with testosterone implants had DLN-IC motoneurons that were significantly larger than those of sham castrates in dendritic length and soma area. These results suggest that motoneurons in the DLN, like those in the SNB, possess a significant degree of structural plasticity in adulthood which is influenced by androgens.     

Hormonally mediated plasticity of motoneuron morphology in the adult rat spinal cord: a cholera toxin-HRP study.

The dorsolateral nucleus (DLN) and the spinal nucleus of the bulbocavernosus (SNB) of the rat lumbar spinal cord are sexually dimorphic groups of motoneurons that innervate striated perineal muscles involved in male copulatory behavior. Androgens control the development of these motoneurons and their target muscles, and continue to influence the system in adulthood. Given that several features of SNB motoneuron morphology have been shown to be androgen sensitive in adult male rats, we examined the effects of androgen manipulations on the morphology of motoneurons in the DLN in adult rats. Adult male rats were castrated and implanted with testosterone-filled or blank implants, or were subjected to a sham-castration procedure. Six weeks after treatment, motoneurons in the DLN were retrogradely labeled with cholera toxin-horseradish peroxidase (HRP) after injection into the ischiocavernosus (IC) muscle and their morphology assessed. Measures of the radial extent and coverage of the dendritic arbor of DLN motoneurons projecting to the IC (DLN-IC motoneurons) were similar across the groups, indicating comparable degrees of HRP transport. However, DLN-IC motoneurons in castrates with blank implants possessed both shorter dendritic lengths and smaller somas than those of castrates treated with testosterone. Castrates with testosterone implants had DLN-IC motoneurons that were significantly larger than those of sham castrates in dendritic length and soma area. These results suggest that motoneurons in the DLN, like those in the SNB, possess a significant degree of structural plasticity in adulthood which is influenced by androgens.     

Ciliary neurotrophic factor arrests muscle and motoneuron degeneration in androgen-insensitive rats

Steroid hormones and neurotrophic factors exert profound and widespread effects on the developing nervous system, including regulation of the size, connectivity, and survival of neurons. Androgenic control of the survival of motoneurons in the spinal nucleus of the bulbocavernosus (SNB) of rats has been well documented. We previously found that ciliary neurotrophic factor (CNTF) mimics many effects of androgen on the developing SNB. Whether effects of CNTF depend on the presence of a functional androgen receptor was evaluated in the present study. Androgen-insensitive male rats bearing the testicular feminization mutation, Tfm, and female littermates were treated with CNTF or with vehicle alone from embryonic day 22 through postnatal day 3. On postnatal day 4 SNB cell number was elevated in both groups receiving CNTF. Volumes of the bulbocavernosus (BC) and levator ani (LA) muscles, targets of SNB motoneurons, were also markedly increased by CNTF. Since the BC appears to degenerate completely in untreated females, these results indicate that CNTF can delay or prevent muscle fiber death. The relative sparing of muscles and motoneurons did not differ for Tfm males and females, demonstrating that effects of CNTF on the SNB neuromuscular system do not require functional androgen receptors. © 1995 John Wiley & Sons, Inc.