Galanin immunoreactivity increased in chicken supraoptic neurons after activation of the vasotocin system at oviposition

The avian arginine vasotocin (AVT) synthesized in the hypothalamic magnocellular neurons and released from the posterior pituitary is known to be involved in the regulation of uterine contractions for oviposition in chickens. However, regulation of AVT synthesis and release within the magnocellular hypothalamus has not been elucidated. Galanin, the oviposition inducing factor in the oviduct of the hen, has been demonstrated to have sexually dimorphic stimulatory action in oxytocin- and vasopressin neurons in the mammalian hypothalamus. In this study, galanin and AVT immunoreactivity was investigated around the time of oviposition in the supraoptic nucleus (SON) to determine if galanin modulates AVT synthesis and/or release. Within SON neurons increased AVT immunoreactivity before oviposition and the decreased AVT immunoreactivity after oviposition implied function-related peptide release. The significantly increased number of galanin neurons co-localizing with AVT immediately after oviposition suggests that galanin is involved in the negative feedback to limit AVT release in the SON. Thus, these data support the idea that AVT in the SON is involved in central regulation of oviposition and that AVT release could be modulated by the neuropeptide galanin.     

Preliminary observations on the fine structure of the medial preoptic nucleus in Coturnix japonica

The organization of the sexually dimorphic medial preoptic nucleus of the Japanese quail was studied at ultrastructural level. The region was characterized by clusters of parvocellular neurons showing a rich supply of axo-somatic synapses and a peculiar distribution of synthetic (rough endoplasmic reticulum) and secretory (Golgi complexes) structures. Further analyses are required to relate these features with sex, hormonal status and sexual behaviour of quail.     

Intracerebral Sex Differences in the Vasotocin System in Birds: Possible Implication in Behavioral and Autonomic Functions

The brain vasotocinergic system demonstrates clear sexual dimorphism in birds investigated so far. This paper examines the evidence obtained in studies on gallinaceous (domestic fowl, Japanese quail) and passerine (canary, junco, zebra finch) birds. Vasotocin (VT)-immunoreactive parvocellular neurons are present in the nucleus of stria terminalis of males, but they are less abundant or absent in the corresponding structure of females. A similar difference has been observed in the dorsal paraventricular area of domestic fowl. Sex-related differences in VT-gene expression have been confirmed byin situhybridization. Moreover, overall brain content of VT mRNA in cockerels is about twice that of hens, suggesting that VT synthesis may also be sexually dimorphic in other brain areas where morphological sex differences have not yet been revealed. The vasotocinergic system in birds is implicated in body fluid homeostasis, and during ontogeny it starts to respond to osmotic challenges in a sexually dimorphic way. Photoperiod, aging, or castration—all associated with changes in circulating testosterone levels—affect sexually dimorphic VT pathways and cell clusters. Sexually dimorphic vasotocinergic circuits are distributed in regions containing steroid-concentrating cells and are closely intermingled with aromatase-containing neurons that may mediate activational effects of gonadal steroids on this peptidergic system. However, it remains undetermined whether the observed neuroanatomical sex differences are related to sexually dimorphic autonomic and behavioral effects induced by VT. Most likely, VT in birds has a modulatory rather than a specific regulatory function in control of male sexual behavior and vocalization.     

Galanin immunoreactivity increased in chicken supraoptic neurons after activation of the vasotocin system at oviposition.

The avian arginine vasotocin (AVT) synthesized in the hypothalamic magnocellular neurons and released from the posterior pituitary is known to be involved in the regulation of uterine contractions for oviposition in chickens. However, regulation of AVT synthesis and release within the magnocellular hypothalamus has not been elucidated. Galanin, the oviposition inducing factor in the oviduct of the hen, has been demonstrated to have sexually dimorphic stimulatory action in oxytocin- and vasopressin neurons in the mammalian hypothalamus. In this study, galanin and AVT immunoreactivity was investigated around the time of oviposition in the supraoptic nucleus (SON) to determine if galanin modulates AVT synthesis and/or release. Within SON neurons increased AVT immunoreactivity before oviposition and the decreased AVT immunoreactivity after oviposition implied function-related peptide release. The significantly increased number of galanin neurons co-localizing with AVT immediately after oviposition suggests that galanin is involved in the negative feedback to limit AVT release in the SON. Thus, these data support the idea that AVT in the SON is involved in central regulation of oviposition and that AVT release could be modulated by the neuropeptide galanin.     

Muscle‐dependent and hormone‐dependent differentiation of the vocal control premotor nucleus robustus archistriatalis and the motornucleus hypoglossus pars tracheosyringealis of the zebra finch

Sex differences in the vertebrate brain (brain sex) are thought to develop owing to the tissue specific action of gonadal hormones similar to the development of secundary sex characteristics of the body. Small sex differences in body anatomy could, however, retrogradely control the sexual differentiation of the central nervous system. This possibility has so far been verified only for motorneuron pools, since the connectivity of sex‐specific higher brain areas to the sexual dimorphic periphery is frequently not well known. Here, we tested whether somatic sex differences feed back on higher brain areas by bilateral denervation of the syringeal musculature of zebra finches before, during, and after onset of estrogen‐sensitive sexual differentiation of forebrain vocal nuclei such as RA (nucleus robustus archistriatalis). In the zebra finch, the sound‐producing musculature (the syrinx), the syrinx motornucleus hypolossus pars tracheosyringealis (nXIIts), and the RA are much larger in males compared to females. Tract tracing studies revealed that the volume and neuron size distribution of the nXIIts was sexually dimorphic in intact but not in animals denervated as juveniles. In contrast, the volume of RA and size of RA neurons of denervated animals were highly sexually dimorphic. Furthermore, estrogen masculinized the RA of denervated females. Thus, sexual differentiation of the RA but not of the nXIIts appears independent of somatic sex differences. The syrinx muscles are, however, important for the soma size of those RA neurons that project to the nXIIts. © 2000 John Wiley & Sons, Inc. J Neurobiol 42: 220–231, 2000     

c-fos Expression in the forebrain and brainstem of White Leghorn hens following osmotic and cardiovascular challenges

In chickens, hyperosmolality and hemorrhage increase hypothalamic vasotocin (AVT) gene expression and stimulate the secretion of AVT from the posterior pituitary gland. In this study, c-fos expression was used to identify areas in the forebrain and brainstem of the domestic chicken that are activated following acute osmotic stress and hemorrhage-induced hypotension. Conscious hens were osmotically stimulated by administering a single intraperitoneal injection of 3 M NaCl (5 ml/kg). Urethane-anesthetized hens were bled to a mean systemic arterial pressure of 80-90 mm Hg and maintained at this blood pressure for 1 h with additional bleedings as required. In both studies, the expression of c-fos was determined in control and experimental birds by using Northern blot analysis and in situ hybridization analysis. Osmotic stress and hemorrhage-induced hypotension increased c-fos expression in the same brain regions. Prominent structures in the forebrain that expressed c-fos mRNA following acute osmotic stress and hemorrhage-induced hypotension included the supraoptic nucleus and paraventricular nucleus and nuclei within the hypothalamus that are anterior and ventral to the third ventricle. In the chicken, this region includes the organum subseptale, the o. vasculosum laminae terminalis, and the nucleus septalis medialis. In the brainstem, following either injection of 3 M NaCl or hemorrhage-induced hypotension, increased c-fos expression was observed in the nucleus of the solitary tract, parabrachial nucleus, area postrema, and locus ceruleus. Thus, the chicken central nervous system appears to use shared neuronal circuitry to stimulate hypothalamic AVT release in response to disturbances in body fluid composition and decreases in either systemic blood pressure or volume.     

Regulation of opioid peptides on the release of arginine vasotocin in the hen

Arginine vasotocin (AVT), an avian neurohypophysial hormone, is released during osmotic stimulation and oviposition. In the present study, the role of opioid peptides on AVT release was studied by examining the effects of an opioid agonist and antagonist on osmotic- and oviposition-induced secretion of AVT. The administration of hypertonic saline (1.5 M NaCl) induced an increase in the plasma levels of AVT. The simultaneous administration of morphine, an opioid receptor agonist, inhibited the osmotically induced increase in plasma levels of AVT in a dose-dependent manner. On the other hand, the co-administration of morphine with naloxone, an opioid receptor antagonist, attenuated the inhibitory effect of morphine. Moreover, injection of naloxone alone enhanced the osmotically induced increase in plasma levels of AVT. However, the administration of morphine did not inhibit the oviposition-induced increase in plasma levels of AVT. These results suggest that osmotic-induced release of AVT may be under opioid regulation, while oviposition-induced release of AVT may be controlled by a different mechanism. J. Exp. Zool. 286:481-486, 2000.     

The effects of sex steroids and vasotocin on behavioral responses to visual and olfactory sexual stimuli in ovariectomized female roughskin newts

Previous studies have found that vasotocin (AVT) administration to male roughskin newts (Taricha granulosa) enhances courtship clasping as well as appetitive responses to specific sexual stimuli and that treating female newts with androgens plus AVT induces the expression of male-typical courtship clasping (the selective clasping of females). However, the unique and/or interactive effects of sex steroids and AVT on appetitive responses to specific sexual stimuli have not yet been determined. To first identify male-typical, sexually dimorphic appetitive responses to female sexual stimuli, we tested intact newts during the breeding season and found that males, but not females, are attracted to female visual and pheromonal sexual stimuli. We then used ovariectomized (ovx) females implanted with empty silastic capsules (Blk) or with capsules containing testosterone (T), dihydrotestosterone (DHT), or estradiol (E2) and then injected with either saline or AVT to determine the effects of steroids and AVT, alone or in combination with each other, on male-typical behavioral responses to those stimuli. E2 treatment depressed responses toward female visual stimuli independently of AVT. On the other hand, only T-implanted, AVT-injected females displayed male-typical behavioral responses toward female olfactory stimuli, preferring to spend more time in proximity to female-scented than unscented newt models and selectively clasping the female-scented models. Together, these results support the conclusion that sex steroids and AVT influence behavioral responses to sexual stimuli via sensory-specific mechanisms. Furthermore, they suggest that T and AVT interact within the brain to influence sensorimotor processing in the pathways that integrate olfactory sexual stimuli into male-typical courtship behaviors.     

Development of vasotocin pathways in the bullfrog brain

The brain of adult bullfrogs (Rana catesbeiana) contains six populations of cells which are immunoreactive for the neurohypophysial peptide arginine vasotocin (AVT). It is unknown when some of these cell populations first appear during development and when the sexual differences in AVT distribution first become apparent. We therefore used immunocytochemistry to examine development of AVT pathways in developing bullfrog tadpoles and in newly metamorphosed froglets of both sexes. AVT-immunoreactive (AVT-ir) cells were already present in the three diencephalic areas (magnocellular preoptic nucleus, suprachiasmatic nucleus and hypothalamus) at stage III (Taylor and Kollros stages), the earliest stage examined. Cell size in the magnocellular nucleus was not bimodally distributed in either tadpoles or froglets. AVT-ir cells in the telencephalic septal nucleus and amygdala did not appear until stage VI. There was no sexual difference in the density of AVT-ir cells or fibers in the amygdala of tadpoles or froglets. Finally, cells in the hindbrain pretrigeminal nucleus appeared much later-after stage XX. Thus, different populations of neurons begin to express AVT at unique times during development. The sexual dimorphism in AVT content observed in the amygdala of adult bullfrogs must appear during juvenile development or at adulthood.     

Muscle-dependent and hormone-dependent differentiation of the vocal control premotor nucleus robustus archistriatalis and the motornucleus hypoglossus pars tracheosyringealis of the zebra finch

Sex differences in the vertebrate brain (brain sex) are thought to develop owing to the tissue specific action of gonadal hormones similar to the development of secondary sex characteristics of the body. Small sex differences in body anatomy could, however, retrogradely control the sexual differentiation of the central nervous system. This possibility has so far been verified only for motorneuron pools, since the connectivity of sex-specific higher brain areas to the sexual dimorphic periphery is frequently not well known. Here, we tested whether somatic sex differences feed back on higher brain areas by bilateral denervation of the syringeal musculature of zebra finches before, during, and after onset of estrogen-sensitive sexual differentiation of forebrain vocal nuclei such as RA (nucleus robustus archistriatalis). In the zebra finch, the sound-producing musculature (the syrinx), the syrinx motornucleus hypoglossus pars tracheosyringealis (nXIIts), and the RA are much larger in males compared to females. Tract tracing studies revealed that the volume and neuron size distribution of the nXIIts was sexually dimorphic in intact but not in animals denervated as juveniles. In contrast, the volume of RA and size of RA neurons of denervated animals were highly sexually dimorphic. Furthermore, estrogen masculinized the RA of denervated females. Thus, sexual differentiation of the RA but not of the nXIIts appears independent of somatic sex differences. The syrinx muscles are, however, important for the soma size of those RA neurons that project to the nXIIts.     

Effect of acute and chronic cardiac denervation on osmotic release of vasotocin in chickens

Hypo- and hyperosmotic NaCl were infused intravenously to examine osmotic release of arginine vasotocin (AVT) in anesthetized, acutely cardiac-denervated chickens and in conscious, chronically denervated birds. Mean arterial blood pressure was consistently higher in denervated compared to sham-operated chickens but heart rates were similar in experimental and control groups. Plasma AVT concentrations (pAVT) were significantly higher than controls in acutely, but not chronically, denervated chickens. The slope of the regression line relating pAVT to plasma sodium concentration was higher in denervated birds indicating that removal of cardiac receptor activity increases the osmotic sensitivity of the AVT system. The results suggest that cardiac end-net receptor activity may participate in the regulation of blood pressure and can modulate the release of antidiuretic hormone in the chicken.     

Initial sex differences in neuron growth and survival within an avian song nucleus develop in the absence of afferent input

Only male zebra finches (Poephila guttata) sing, and nuclei implicated in song behavior exhibit marked sex differences in neuron number. In the robust nucleus of the anterior neostriatum (RA), these sex differences develop because more neurons die in young females than in males. However, it is not known whether the sexually dimorphic survival of RA neurons is a primary event in sexual differentiation or a secondary response to sex differences in the number of cells interacting trophically with RA neurons. In particular, since sexual differentiation of the RA parallels the development of dimorphisms in the numbers of neurons providing afferent input from the lateral magnocellular nucleus of the anterior neostriatum (lMAN) and the high vocal center (HVC), it has been hypothesized that sex differences in the size of these afferent populations trigger differential RA neuron survival and growth. To test this hypothesis, we lesioned either the lMAN or both the lMAN and HVC unilaterally in 12-day-old male and female zebra finches. Subsequently, RA cell death and RA neuron number and size were measured. Unilateral lMAN lesions increased cell death and decreased neuron number and size within the ipsilateral RA of both sexes. However, even in the lMAN-lesioned hemisphere, these effects were less pronounced in males than in females, so that by day 25 the volume, number, and size of neurons were sexually dimorphic in both the contralateral and ipsilateral RA. Similarly, the absence of both lMAN and HVC afferents did not prevent the emergence of sex differences in the number and size of RA neurons by 25 day posthatching. We conclude that these sex differences within the RA are not a secondary response to dimorphisms in the numbers of lMAN or HVC neurons providing afferent input. © 1995 John Wiley & Sons, Inc.     

Sex and seasonal co-variation of arginine vasotocin (AVT) and gonadotropin-releasing hormone (GnRH) neurons in the brain of the halfspotted goby

Gonadotropin-releasing hormone (GnRH) and arginine vasotocin (AVT) are critical regulators of reproductive behaviors that exhibit tremendous plasticity, but co-variation in discrete GnRH and AVT neuron populations among sex and season are only partially described in fishes. We used immunocytochemistry to examine sexual and temporal variations in neuron number and size in three GnRH and AVT cell groups in relation to reproductive activities in the halfspotted goby (Asterropteryx semipunctata). GnRH-immunoreactive (-ir) somata occur in the terminal nerve, preoptic area, and midbrain tegmentum, and AVT-ir somata within parvocellular, magnocellular, and gigantocellular regions of the preoptic area. Sex differences were found among all GnRH and AVT cell groups, but were time-period dependent. Seasonal variations also occurred in all GnRH and AVT cell groups, with coincident elevations most prominent in females during the peak- and non-spawning periods. Sex and temporal variability in neuropeptide-containing neurons are correlated with the goby's seasonally-transient reproductive physiology, social interactions, territoriality and parental care. Morphological examination of GnRH and AVT neuron subgroups within a single time period provides detailed information on their activities among sexes, whereas seasonal comparisons provide a fine temporal sequence to interpret the proximate control of reproduction and the evolution of social behavior.     

Fibroblast growth factor-2 stimulates cell proliferation and decreases sexually dimorphic cell death in an avian song control nucleus

The neural system controlling song in birds has proven a useful model for investigating how neuronal growth and survival are regulated by sexual differentiation. The present study focused on one song control area, the robust nucleus of the archistriatum (RA), and explored how sex differences in the proliferation of putative glia cells in this region influence sexually dimorphic cell survival. In zebra finches (Poephila guttata), RA neuron death is much greater in young females than in males, resulting in marked sex differences in RA neuron number. An earlier study indicated that just prior to this sexually dimorphic neuron death the proliferation of putative glia cells within the RA is significantly lower in females than in males and remains so throughout the peak of neuron death. This suggests that sex differences in glia (or glia-derived molecules) might regulate neuron survival during sexual differentiation of the RA. To determine whether increased cell proliferation within the RA favors increased cell survival, we infused the potent glia mitogen fibroblast growth factor-2 (FGF-2) into the RA unilaterally in young females. We find that FGF-2 infusions increase RA cell proliferation and concurrently decrease the incidence of degenerating RA cells, results consistent with the hypothesis that glia exert neurotrophic effects on RA neurons during sexual differentiation. © 1998 John Wiley & Sons, Inc. J Neurobiol 37: 573–581, 1998     

The interstitial nuclei of the human anterior hypothalamus: an investigation of variation with sex, sexual orientation, and HIV status

The interstitial nuclei of the human anterior hypothalamus (INAH1-4) have been considered candidates for homology with the sexually dimorphic nucleus of the preoptic area of the rat. Volumetric sexual dimorphism has been described for three of these nuclei (INAH1-3), and INAH3 has been reported to be smaller in homosexual than heterosexual men. The current study measured the INAH in Nissl-stained coronal sections in autopsy material from 34 presumed heterosexual men (24 HIV- and 10 HIV+), 34 presumed heterosexual women (25 HIV- and 9 HIV+), and 14 HIV+ homosexual men. HIV status significantly influenced the volume of INAH1 (8% larger in HIV+ heterosexual men and women relative to HIV- individuals), but no other INAH. INAH3 contained significantly more neurons and occupied a greater volume in presumed heterosexual males than females. No sex difference in volume was detected for any other INAH. No sexual variation in neuronal size or density was observed in any INAH. Although there was a trend for INAH3 to occupy a smaller volume in homosexual men than in heterosexual men, there was no difference in the number of neurons within the nucleus based on sexual orientation.     

Stereological evaluation and Golgi study of the sexual dimorphisms in the volume,cell numbers,and cell size in the medial preoptic nucleus of the rat

The medial preoptic nucleus (MPN) and the sexually dimorphic nucleus of the preoptic area (SDN-POA) stand out as prominent sexually dimorphic cell groups of the rat brain. However, quantitative data on sex-related differences in these nuclei in the adult rat are confined to their volume. We have used stereological methods and Golgi-impregnated material to examine whether, in young adult rats, the sexual dimorphism in the volume of the MPN, including its divisions, and of the SDN-POA, reflect similar differences in the number and size of their neurons. We found that the total number of neurons in all MPN divisions is higher and the mean somatic volume larger in males than in females. In addition, the total dendritic length of MPN neurons is greater, but the dendritic spine density is smaller, in males than in females. Likewise, in the SDN-POA the total number and size of its neurons is greater in males than in females. The sex differences in all quantitative parameters evaluated accounted for the larger volume of the MPN and SDN-POA in males relative to females. In addition, the MPN neuropil also displays sex-related differences in its volume, and these differences closely match those detected for the volume of each MPN division. It deserves to be emphasised that the numerical density of neurons was the only parameter found to be significantly higher in females than in males in all MPN divisions and in the SDN-POA. Our results show that the MPN and the SDN-POA display sex differences in the volume, total number of neurons, and size of neuronal cell bodies and dendritic trees. Furthermore, they also indicate that the neuropil is critical for the establishment of sexual dimorphism in the size of the MPN.     

Hormonal Regulation of Opioid Peptide Neurons in the Anteroventral Periventricular Nucleus

Steroid hormones provide a means of coordinating the activity of widespread neural systems that mediate endocrine, autonomic, and somatomotor aspects of reproductive processes that are essential for the propagation of mammalian species. Because these processes are quite different in each sex, the neural pathways that control them are also sexually differentiated. The anteroventral periventricular nucleus (AVPV) of the preoptic region occupies a nodal point in sexually dimorphic forebrain circuits and appears to play a critical role in regulating gonadotropin secretion. The AVPV contains sexually dimorphic populations of opioid peptide containing neurons that display different patterns of development and are differentially regulated in adult animals by gonadal steroids. Moreover, estrogen (ER) and progesterone (PR) receptors are expressed in AVPV neurons in a transmitter-specific way, and the expression of these nuclear transacting factors is differentially regulated by sex steroids. Thus, neurons in the AVPV show distinct patterns of hormonal regulation of gene expression, and distinct hormone receptor profiles.     

Nonapeptides and social behavior in fishes

The nonapeptide hormones arginine vasotocin and isotocin play important roles in mediating social behaviors in fishes. Studies in a diverse range of species demonstrate variation in vasotocin neuronal phenotypes across within and between sexes and species as well as effects of hormone administration on aggressive and sexual behaviors. However, patterns vary considerably across species and a general explanatory model for the role of vasotocin in teleost sociosexual behaviors has proven elusive. We review these findings, examine potential explanations for the lack of agreement across studies, and propose a model based on the parvocellular AVT neurons primarily mediating social approach and subordinance functions while the magnocellular and gigantocellular AVT neurons mediate courtship and aggressive behaviors. Isotocin neuronal phenotypes and effects on behavior are relatively unstudied, but research to date suggests this will be a fruitful line of inquiry. This article is part of a Special Issue entitled Oxytocin, Vasopressin, and Social Behavior.     

Dehydration-induced drinking decreases Fos expression in hypothalamic paraventricular neurons expressing vasopressin but not corticotropin-releasing hormone

Water-restricted (WR) rats exhibit a rapid suppression of plasma corticosterone following drinking. The present study monitored Fos-like immunoreactivity (Fos) to assess the effect of WR-induced drinking on the activity of vasopressin (VP)-positive magnocellular and parvocellular neurons and corticotropin-releasing hormone (CRH)-positive parvocellular neurons in the paraventricular nucleus of the hypothalamus. Adult male rats received water for 30 min (WR) in the post meridiem (PM) each day for 6 days and were killed without receiving water or at 1 h after receiving water for 15 min. In WR rats, Fos increased in VP magnocellular and parvocellular neurons but not CRH neurons. After drinking, Fos was reduced in VP magnocellular and parvocellular neurons but did not change in CRH neurons. To assess the severity of osmotic stress, rats were sampled throughout the final day of WR. Plasma osmolality, hematocrit and plasma VP were increased throughout the day before PM rehydration, and plasma ACTH and corticosterone were elevated at 1230 and 1430, respectively, showing that WR activates hypothalamic-pituitary-adrenal activity during the early PM before the time of rehydration. To determine the effects of WR-induced drinking on CRH neurons activated by acute stress, WR rats underwent restraint. Restraint increased plasma ACTH and corticosterone and Fos in CRH neurons; although rehydration reduced plasma ACTH and Fos expression in VP neurons, Fos in CRH neurons was not affected. These results suggest that inhibition of VP magnocellular and parvocellular neurons, but not CRH parvocellular neurons, contributes to the suppression of corticosterone after WR-induced drinking.