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.     

Androgenic and Oestrogenic Influences on Tyrosine Hydroxylase‐Immunoreactive Cells of the Prairie Vole Medial Amygdala and Bed Nucleus of the Stria Terminalis

The posterodorsal medial amygdala (MeApd) and principal nucleus of the bed nucleus of the stria terminalis (pBST) are densely interconnected sites integrating steroid hormone and olfactory information necessary for sociosexual behaviours in many rodents. Our laboratory recently reported sexually dimorphic populations of cells containing tyrosine hydroxylase (TH) located in the MeApd and pBST of prairie voles (Microtus ochrogaster), with males having many more TH‐immunoreactive (‐ir) cells in these sites than do females. Gonadal hormones circulating during adulthood were showm to regulate this sex difference because it was eliminated by castrating adult males or implanting females with testosterone‐filled capsules. In the present study, we demonstrate that many (25–65%) TH‐ir cells in the MeApd and pBST of adult virgin male and female prairie voles also contain immunoreactivity for either the androgen receptor or oestrogen receptor α. Subcutaneous implants of oestradiol benzoate mimicked the effects of testosterone and maintained high numbers of TH‐ir cells in these sites in castrated males. However, implants of dihydrotestosterone (DHT) did not, and these males had low numbers of TH‐ir cells similar to castrated males given empty capsules. A similar effect was found in females, where testosterone or oestradiol benzoate greatly increased the number of TH‐ir cells in these sites compared to intact or ovariectomised controls, but DHT did not. DHT implants did, however, maintain high seminal vesicle weights in males. Thus, many of the TH‐ir cells in the prairie vole MeApd and pBST are potentially sensitive to androgens and oestrogens, although maintaining immunocytochemically detectable levels of TH in these cells may depend more on an oestrogen‐mediated mechanism in both sexes. These data have implications for understanding how gonadal hormone release across the reproductive cycle modulates these species‐specific groups of catecholaminergic cells and socially monogamous behaviours in prairie voles.     

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.     

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.     

Sexual differentiation of the copulatory neuromuscular system in green anoles (Anolis carolinensis): normal ontogeny and manipulation of steroid hormones

The copulatory neuromuscular system of green anoles is sexually dimorphic and differentiates during embryonic development, although details of the process were unknown. In Experiment 1, we determined the time course of normal ontogeny. Both male and female embryos possessed bilateral copulatory organs (hemipenes) and associated muscles until incubation day 13; the structures completely regressed in female embryos by incubation day 19 (total incubation 34 days). In Experiment 2, we treated eggs with testosterone, dihydrotestosterone, estradiol, or vehicle on both incubation days 10 and 13 to determine whether these steroid hormones mediate sexual differentiation. These time points fall between gonadal differentiation, which was determined in Experiment 1 to complete before day 10, and regression of the peripheral copulatory system in females. Tissue was collected on the day of hatching. Gonads were classified as testes or ovaries; presence versus absence of hemipenes and muscles, and the number and size of copulatory motoneurons were determined. Copulatory system morphology of vehicle-treated animals matched their gonadal sex. Hemipenes and muscles were absent in estradiol-treated animals, and androgens rescued the hemipenes and muscles in most females. Both testosterone and dihydrotestosterone treatment also caused hypertrophy of the hemipenes, which were everted in animals treated with these steroids. Copulatory motoneurons, assessed on the day of hatching in both experiments, were not dimorphic in size or number. Steroid treatment significantly increased motoneuron size and number overall, but no significant differences were detected in pairwise comparisons. These data demonstrate that differentiation of peripheral copulatory neuromuscular structures occurs during embryonic development and is influenced by gonadal steroids (regression by estradiol and enhancement by androgens), but associated motoneurons do not differentiate until later in life.     

Hormonal control of neuron number in sexually dimorphic spinal nuclei of the rat: II. Development of the spinal nucleus of the bulbocavernosus in androgen-insensitive (Tfm) rats

The spinal nucleus of the bulbocavernosus (SNB) is a sexually dimorphic motor nucleus whose development is under the control of steroid hormones. The SNB contains many more motoneurons in adult male rats than in females, and this sex difference is produced by a sexually dimorphic motoneuron death which is regulated by androgens. To study further the role of androgens in the development of sex differences in SNB motoneuron number, we examined SNB development in males with the testicular feminization mutation (Tfm) which renders them insensitive to androgens. Counts of SNB motoneurons perinatally revealed that SNB development in normal male and female King-Holtzman rats was similar to that reported previously for Sprague-Dawley rats; SNB motoneuron number increased from initially low levels at embryonic day 18 through the day before birth, when motoneuron numbers in both sexes were substantially higher than adult levels. After this prenatal increase, motoneuron number declined in both sexes, until by postnatal day 10 motoneuron numbers were in their adult ranges and the sex difference was fully expressed. Females lost more motoneurons than did males during this period, and this loss was due to motoneuron death as revealed by counts of degenerating cells. SNB development in King-Holtzman Tfm males was similar to that of normal males through embryonic day 20, suggesting that androgens may not be necessary for the initial increase in motoneuron numbers in the SNB. Thereafter, SNB motoneuron numbers in Tfm males declined in a female-typical fashion; Tfm males and normal females did not differ at any postnatal age.(ABSTRACT TRUNCATED AT 250 WORDS)     

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.     

Connexin36 in gap junctions forming electrical synapses between motoneurons in sexually dimorphic motor nuclei in spinal cord of rat and mouse

Pools of motoneurons in the lumbar spinal cord innervate the sexually dimorphic perineal musculature, and are themselves sexually dimorphic, showing differences in number and size between male and female rodents. In two of these pools, the dorsomedial nucleus (DMN) and the dorsolateral nucleus (DLN), dimorphic motoneurons are intermixed with non‐dimorphic neurons innervating anal and external urethral sphincter muscles. As motoneurons in these nuclei are reportedly linked by gap junctions, we examined immunofluorescence labeling for the gap junction‐forming protein connexin36 (Cx36) in male and female mice and rats. Fluorescent Cx36‐labeled puncta occurred in distinctly greater amounts in the DMN and DLN of male rodents than in other spinal cord regions. These puncta were localized to motoneuron somata, proximal dendrites, and neuronal appositions, and were distributed either as isolated or large patches of puncta. In both rats and mice, Cx36‐labeled puncta were associated with nearly all (> 94%) DMN and DLN motoneurons. The density of Cx36‐labeled puncta increased dramatically from postnatal days 9 to 15, unlike the developmental decreases in these puncta observed in other central nervous system regions. In females, Cx36 labeling of puncta in the DLN was similar to that in males, but was sparse in the DMN. In enhanced green fluorescent protein (EGFP)–Cx36 transgenic mice, motoneurons in the DMN and DLN were intensely labeled for the EGFP reporter in males, but less so in females. The results indicate the presence of Cx36‐containing gap junctions in the sexually dimorphic DMN and DLN of both male and female rodents, suggesting coupling of not only sexually dimorphic but also non‐dimorphic motoneurons in these nuclei.     

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.     

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.     

Neonatal androgen maintains sexually dimorphic muscles in the absence of innervation

We examined the site of androgen action in maintaining the sexually dimorphic spinal nucleus of the bulbocavernosus (SNB) and its target perineal muscles, the bulbocavernosus (BC) and levator ani (LA), in rats. To determine whether androgen action on SNB motoneurons is crucial for BC/LA survival, we removed SNB cells in newborn female rats by lumbosacral spinalectomy, administered testosterone propionate (TP) on days 1 and 3 of life, and examined for the presence of BC/LA muscles in adulthood. BC/LA muscles were present in all TP-treated spinalectomized females, and staining of these muscles with alpha-bungarotoxin or for acetylcholinesterase showed no evidence of cholinergic innervation. Thus, complete neonatal denervation of the BC/LA does not prevent TP from sparing these muscles, suggesting that androgen acts directly upon BC/LA muscles to maintain them during development. This androgenic maintenance of the BC/LA may be crucial for the survival of SNB motoneurons.     

Sexual dimorphism and the influence of neonatal androgen in the dorsolateral motor nucleus of the rat lumbar spinal cord

There is a sexually dimorphic motor nucleus, the spinal nucleus of the bulbocavernosus (SNB) in the fifth and sixth lumbar segments of the rat spinal cord. We now report a second sex difference in the dorsolateral nucleus (DLN) in the ventral horn of the rat lumbar cord, which includes motoneurons innervating the ischiocavernosus muscle, a sexually dimorphic perineal muscle. Adult females possess fewer motoneurons in the DLN, probably because of an absence of neurons innervating the ischiocavernosus muscle, which females lack. The effect of a single dose of testosterone propionate on day 2 of life was confined to a specific rostrocaudal region of the adult DLN in which it partially masculinized the female DLN. Masculinized females have more DLN neurons than control females. The direction of change induced in DLN neuron number by the neonatal hormone treatment is compatible with the hypothesis that androgens are involved with the sexually dimorphic development of the DLN. In another motor nucleus, the retrodorsolateral nucleus, a small sex difference in neuron number was found in one study, but was not replicated in a second experiment.     

Neuroprotective effects of testosterone on the morphology and function of somatic motoneurons following the death of neighboring motoneurons

Motoneuron loss is a significant medical problem, capable of causing severe movement disorders or even death. We have previously shown that partial depletion of motoneurons from sexually dimorphic, highly androgen‐sensitive spinal motor populations induces dendritic atrophy in remaining motoneurons, and this atrophy is attenuated by treatment with testosterone. To test whether testosterone has similar effects in more typical motoneurons, we examined potential neuroprotective effects in motoneurons innervating muscles of the quadriceps. Motoneurons innervating the vastus medialis muscle were selectively killed by intramuscular injection of cholera toxin‐conjugated saporin. Simultaneously, some saporin‐injected rats were given implants containing testosterone or left untreated. Four weeks later, motoneurons innervating the ipsilateral vastus lateralis muscle were labeled with cholera toxin‐conjugated horseradish peroxidase, and dendritic arbors were reconstructed in three dimensions. Compared with intact normal males, partial motoneuron depletion resulted in decreased dendritic length in remaining quadriceps motoneurons, and this atrophy was attenuated by testosterone treatment. To examine the functional consequences of the induced dendritic atrophy, and its attenuation with testosterone treatment, the activation of remaining quadriceps motoneurons was assessed using peripheral nerve recording. Partial motoneuron depletion resulted in decreased amplitudes of motor nerve activity, and these changes were attenuated by treatment with testosterone, providing a functional correlate to the neuroprotective effects of testosterone treatment on quadriceps motoneuron morphology. Together these findings suggest that testosterone has neuroprotective effects on morphology and function in both highly androgen‐sensitive as well as more typical motoneuron populations, further supporting a role for testosterone as a neurotherapeutic agent in the injured nervous system. J. Comp. Neurol. 512:359–372, 2009. © 2008 Wiley‐Liss, Inc.     

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.     

Sexual dimorphism in brain weight of meadow voles: role of gonadal hormones

In most adult mammals, brain weights of males exceed those of females. The role of androgens in the genesis of this sex difference was assessed in meadow voles by acute neonatal or chronic postweaning manipulation of testosterone titers. Female voles given a single injection of testosterone propionate (TP) on the second day of postnatal life had brain weights in adulthood that were indistinguishable from those of male voles and significantly heavier than those of control females. Whole brain DNA content, a measure of cell number, was not increased by neonatal TP treatment. Females treated with TP from day 19 to 70 had lower brain weights than control females and males gonadectomized at 19 days of age had greater brain weights than did intact male voles at day 70. The sex dimorphism in brain weight reflects organizational effects of testosterone during perinatal development. Beginning at weaning, and continuing through postpubertal development, testosterone decreases brain weight in both sexes. We suggest that testosterone affects brain weight by altering cell size or non-cellular components rather than cell number.     

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.     

Sexual dimorphism in neuronal number of the posterodorsal medial amygdala is independent of circulating androgens and regional volume in adult rats

The posterodorsal medial amygdala (MePD) in rodents integrates olfactory and pheromonal information, which, coupled with the appropriate hormonal signals, may facilitate or repress reproductive behavior in adulthood. MePD volume and neuronal soma size are greater in male rats than in females, and these sexual dimorphisms are maintained by adult circulating hormone levels. Castration of adult males causes these measures to shrink to the size seen in females 4 weeks later, whereas testosterone treatment of adult females for 4 weeks enlarges these measures to the size of males. We used stereological methods to count the number of cells in the MePD and found that, in addition to the sex difference in regional volume and soma size, males also have more MePD neurons than do females, yet these numbers are unaffected by the presence or absence of androgen in adults of either sex. Males also have more glial cells than do females, but, in contrast to the effects on neuronal number, the number of glial cells is affected by androgen in the right MePD of both sexes and, therefore, may contribute to regional volume changes in adulthood in that hemisphere. Thus, regional volume, neuronal size, and glial numbers vary in the MePD of adult rats in response to circulating androgens, but neuronal number does not. These results suggest that the sex difference in neuronal number in the rat MePD may be "organized" by androgens prior to adulthood, whereas regional volume, neuronal size, and glial numbers can be altered by androgens in adulthood.     

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.     

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.     

Morphology of rat spinal motoneurons with normal and hormonally altered specificity

Potential determinants of motoneuronal morphology were examined by using a sexually dimorphic, steroid-sensitive neuromuscular system in the rat spinal cord. In males, the spinal nucleus of the bulbocavernosus (SNB) innervates the perineal muscles bulbocavernosus (BC) and levator ani (LA), and the dorsolateral nucleus (DLN) innervates the ischiocavernosus muscle (IC). Adult females normally lack these motoneurons and the peripheral targets. Prenatal exposure of females to the androgen dihydrotestosterone propionate (DHTP) partially masculinizes this neuromuscular system and alters moto-neuron-to-muscle specificity, resulting in retained SNB target muscles anomalously innervated by motoneurons in the DLN. Because the morphology of SNB and DLN motoneurons normally differs significantly, the influence of spinal cord location and peripheral target on motoneuron morphology can be directly compared. Injection of cholera toxin conjugated to horseradish peroxidase (CTHRP) into the LA of DHTP-treated females labeled motoneurons predominantly in the SNB. These (SNB-LA) motoneurons in DHTP females were identical in all morphological measures to those of normal males. CTHRP injection into the BC of DHTP females labeled motoneurons in both the SNB and the DLN. SNB-BC motoneurons in DHTP females resembled those of normal males in process number and orientation, but were significantly smaller in dendritic length per motoneuron and in soma size. The DLN motoneurons anomalously projecting to the BC in DHTP females differed significantly from SNB-BC motoneurons in soma size and number and orientation of primary processes. However, these motoneurons were identical in all respects to DLN-IC motoneurons in DHTP females; DLN-IC motoneurons were similar to those of normal males in the orientation of their dendritic arbor, but were significantly smaller in dendritic length, soma size, and number of primary processes. These comparisons make it clear that DHTP selectively affects motoneuronal specificity and morphology in specific motoneuron classes. Further, motoneuronal morphology in the SNB/DLN system appears to be influenced more by spinal cord location than by peripheral target.