Effect of Intraseptal Vasotocin and Vasoactive Intestinal Polypeptide Infusions on Courtship Song and Aggression in the Male Zebra Finch (Taeniopygia guttata)

The present experiments were conducted to test the hypothesis that septal arginine vasotocin (AVT) and vasoactive intestinal polypeptide (VIP) modulate directed song (a courtship behaviour) and aggression in male zebra finches ( Taeniopygia guttata ). Subjects were surgically fitted with a guide cannula directed at the septum. Following recovery they were tested for aggression and directed song following infusions of AVT, its antagonist (anti-vasopressin, AVP), and saline volume control. Infusion of the AVT antagonist significantly reduced all three aggressive behaviours measured (pecks, beak fences and chases); and AVT infusion significantly facilitated beak fencing. Vasoactive intestinal polypeptide treatment significantly reduced pecking. No treatment produced a change in directed song. Comparison with findings in mammals suggests that modulation of aggression by septal AVT (or AVP) is evolutionarily conserved in vertebrates, but modulation of aggression by VIP has not previously been reported for any vertebrate.     

Effect of Short‐and Long‐Term Gonadectomy on Neuroactive Steroid Levels in the Central and Peripheral Nervous System of Male and Female Rats

Significant levels of neuroactive steroids are still detected in the nervous system of rodents after the removal of peripheral steroidogenic glands. However, the influence of the plasma levels of gonadal steroids on the levels of neuroactive steroids in the nervous system has not so far been clarified in detail. Accordingly, by liquid chromatography tandem mass spectrometry, we have analysed the levels of neuroactive steroids in the sciatic nerve, in three central nervous system (CNS) regions (i.e. cerebellum, cerebral cortex and spinal cord) and in the plasma of male and female animals. The levels present in gonadally intact animals were compared with those present in short‐ and long‐term gonadectomised animals. We observed that: (i) changes in neuroactive steroid levels in the nervous system after gonadectomy do not necessarily reflect the changes in plasma levels; (ii) long‐term gonadectomy induces changes in the levels of neuroactive steroids in the peripheral nervous system (PNS) and the CNS that, in some cases, are different to those induced by short‐term gonadectomy; (iii) the effect of gonadectomy on neuroactive steroid levels is different between the PNS and the CNS and within different CNS regions; and (iv) the effects of gonadectomy on neuroactive steroid levels in the nervous system show sex differences. Altogether, these observations indicate that the nervous system adapts its local levels of neuroactive steroids in response to changes in gonadal hormones with sex and regional specificity and depending on the duration of the peripheral modifications.     

Association between iron‐deficiency anemia and depression: A web‐based Japanese investigation

Aim

This web‐based survey aimed to examine the relation between iron‐deficiency anemia and depression in 11 876 Japanese participants.

Methods

Participants consisted of 1000 individuals with self‐reported history of depression (mean age, 41.4 ± 12.3 years; 499 women) and 10 876 population‐based controls (mean age, 45.1 ± 13.6 years; 5185 women). The 6‐item Kessler Scale (K6) score was used as a psychological distress scale. The design of the study was cross‐sectional.

Results

The rate of self‐reported lifetime history of iron‐deficiency anemia was higher in the depression group in both men (depression, 7.2%; control, 4.0%; P < 0.001; odds ratio [OR], 1.86; 95% confidence interval [CI], 1.30–2.68) and women (depression, 33.4%; control, 25.8%; P < 0.001; OR, 1.45; 95%CI, 1.19–1.76). The K6 score in participants with self‐reported history of iron‐deficiency anemia was higher in both the depression (P = 0.004) and control (P < 0.001) groups. In addition, in all participants, the rate of individuals who showed a K6 cut‐off score of 13 or more was higher in those with a self‐reported history of iron‐deficiency anemia (P < 0.001; OR, 1.47; 95%CI, 1.31–1.65). Logistic regression analyses revealed that self‐reported history of depression and the K6 score were positively associated with self‐reported history of iron‐deficiency anemia (all P < 0.01).

Conclusion

Self‐reported history of iron‐deficiency anemia was associated with self‐reported history of depression. Furthermore, self‐reported history of iron‐deficiency anemia was associated with higher psychological distress.

     

Sex‐Specific Effects of Chronic Administration of Relaxin‐3 on Food Intake,Body Weight and the Hypothalamic‐Pituitary‐Gonadal Axis in Rats

The present study examined the effects of chronic central administration of relaxin‐3 (RLN3) on food intake, body weight and fat mass in intact and sterilised male and female rats, as well as on hypothalamic‐pituitary‐gonadal (HPG) axis activity in intact male and female rats that received i.c.v. infusions of RLN3 (400 pmol/day) or vehicle during a 14‐day period. The intact RLN3‐injected rats displayed a higher body weight than the vehicle‐treated groups, and this increase was statistically significantly stronger in female rats compared to male rats. In addition, feed efficiency and gonadal white adipose tissue weight were higher in female RLN3‐injected rats. Chronic i.c.v. administration of RLN3 activated the HPG axis in intact male rats, whereas inhibition of the HPG axis was observed in intact female rats. RLN3 significantly increased the plasma levels of luteinising hormone and follicular‐stimulating hormone in male rats but not in female rats. Conversely, hypothalamic expression of gonadotrophin‐releasing hormone mRNA was decreased by RLN3 in female rats but not in male rats. In addition, the plasma levels of oestradiol were significantly decreased by RLN3 administration in female rats. Consequently, intact RLN3‐injected female rats failed to display phasic inhibition of eating during oestrus. Sex‐specific effects of RLN3 on food intake and body weight were also observed in ovariectomised female and orchidectomised male rats, suggesting that the sex‐specific effects of RLN3 on energy metabolism are independent on the differential effects of RLN3 on HPG axis activity in male and female rats.     

Sex‐ and region‐specific pubertal maturation of the corticotropin‐releasing factor receptor system in the rat

One of the most reliable findings in psychiatry is in the incidence of anxiety and depression. Beginning at puberty, women develop mood disorders twice as often as men. Because corticotropin‐releasing factor (CRF) receptors are implicated, we compared CRF receptor binding in pre‐ and postpubertal rats. In each brain area, CRF receptor binding was sexually dimorphic, but no two areas were alike in the way the sexes differed. In the nucleus accumbens and olfactory tubercle, CRF₁ binding was initially the same in juveniles, but became greater in adult females. In piriform cortex, CRF₁ binding increased in females and decreased in males, again becoming sexually dimorphic. CRF₁ binding in the anterior cingulate was greater in females than in males at both ages. In CA3, CRF₁ binding was greater in males before puberty but decreased during puberty, abolishing the sex difference. CRF₂ binding in the posterior bed nucleus of the stria terminalis was greater in males irrespective of age. In contrast, in each of three subdivisions of the lateral septum, females had greater CRF₂ binding than males as juveniles, or, as juveniles and as adults. CRF₂ binding in the ventromedial hypothalamus was the same in juveniles, but binding levels increased in males, leading to an adult sex difference. Thus, eight CRF receptor‐expressing areas displayed eight distinct sex differences. These results show that sex differences pervade the CRF receptor system in juvenile and adult rats, and the mechanisms that control them are likely to be sex‐, region‐, and subtype‐specific. J. Comp. Neurol. 522:1284–1298, 2014. © 2013 Wiley Periodicals, Inc.     

Enteric neural crest‐derived cells and neural stem cells: biology and therapeutic potential

The enteric nervous system arises from two regions of the neural crest; the vagal neural crest which gives rise to the vast majority of enteric neurones throughout the gastrointestinal tract, and the sacral neural crest which contributes a smaller number of cells that are mainly distributed within the hindgut. The migration of vagal neural crest cells into, and along the gut is promoted by GDNF, which is expressed by the gut mesenchyme and is the ligand for the Ret/GFRα1 signalling complex present on migrating vagal‐derived crest cells. Sacral neural crest cells enter the gut after it has been colonized by vagal neural crest cells, but the molecular control of sacral neural crest cell development has yet to be elucidated. Under the influence of both intrinsic and extrinsic cues, neural crest cells differentiate into glia and different types of enteric neurones at different developmental stages. Recently, the potential for neural stem cells to form an enteric nervous system has been examined, with the ultimate aim of using neural stem cells as a therapeutic strategy for some gut disorders where enteric neurones are reduced or absent.     

Sex‐Related Differences in the Effects of Sports‐Related Concussion: A Review

Sports‐related concussion is a serious health challenge, and females are at higher risk of sustaining a sports‐related concussion compared to males. Although there are many studies that investigate outcomes following concussion, females remain an understudied population, despite representing a large proportion of the organized sports community. In this review, we provide a summary of studies that investigate sex‐related differences in outcome following sports‐related concussion. Moreover, we provide an introduction to the methods used to study sex‐related differences after sports‐related concussion, including common clinical and cognitive measures, neuroimaging techniques, as well as biomarkers. A literature search inclusive of articles published to March 2020 was performed using PubMed. The studies were reviewed and discussed with regard to the methods used. Findings from these studies remain mixed with regard to the effect of sex on clinical symptoms, concussion‐related alterations in brain structure and function, and recovery trajectories. Nonetheless, there is initial evidence to suggest that sex‐related differences following concussion are important to consider in efforts to develop objective biomarkers for the diagnosis and prognosis of concussion. Additional studies on this topic are, however, clearly needed to improve our understanding of sex‐related differences following concussion, as well as to understand their neurobiological underpinnings. Such studies will help pave the way toward more personalized clinical management and treatment of sports‐related concussion.     

Ontogeny of the vasopressinergic neurons of the suprachiasmatic nucleus and their extrahypothalamic projections in the rat brain—presence of a sex difference in the lateral septum

Immunocytochemical studies have revealed the presence of extensive vasopressinergic projections from the suprachiasmatic nucleus to the limbic system and other brain areas. Vibratome sections and the unlabeled antibody enzyme method were used to investigate the ontogeny of the vasopressinergic neurons of the suprachiasmatic nucleus and their exohypothalamic fibers in the rat brain. The first immunopositive neurons of this nucleus were revealed on the 2nd postnatal day. An adult appearance of the suprachiasmatic nucleus was detected on day 14. Although fibers appeared in the periventricular nucleus already on the 7th postnatal day, such fibers were visible in the lateral septum and lateral habenular nucleus only on day 10. From the 12th postnatal day onwards a marked sex difference developed with respect to the density of the vasopressin fibers in the lateral septum and, to a lesser extent, in the lateral habenular nucleus. In male rats the fiber density was higher in both areas. This sex difference persisted in adulthood.     

Brain development during adolescence: A mixed‐longitudinal investigation of cortical thickness,surface area,and volume

What we know about cortical development during adolescence largely stems from analyses of cross‐sectional or cohort‐sequential samples, with few studies investigating brain development using a longitudinal design. Further, cortical volume is a product of two evolutionarily and genetically distinct features of the cortex ‐ thickness and surface area, and few studies have investigated development of these three characteristics within the same sample. The current study examined maturation of cortical thickness, surface area and volume during adolescence, as well as sex differences in development, using a mixed longitudinal design. 192 MRI scans were obtained from 90 healthy (i.e., free from lifetime psychopathology) adolescents (11‐20 years) at three time points (with different MRI scanners used at time 1 compared to 2 and 3). Developmental trajectories were estimated using linear mixed models. Non‐linear increases were present across most of the cortex for surface area. In comparison, thickness and volume were both characterised by a combination of non‐linear decreasing and increasing trajectories. While sex differences in volume and surface area were observed across time, no differences in thickness were identified. Furthermore, few regions exhibited sex differences in the cortical development. Our findings clearly illustrate that volume is a product of surface area and thickness, with each exhibiting differential patterns of development during adolescence, particularly in regions known to contribute to the development of social‐cognition and behavioral regulation. These findings suggest that thickness and surface area may be driven by different underlying mechanisms, with each measure potentially providing independent information about brain development. Hum Brain Mapp 37:2027–2038, 2016. © 2016 Wiley Periodicals, Inc.     

Neonatal androgen‐dependent sex differences in lumbar spinal cord dopamine concentrations and the number of A11 diencephalospinal dopamine neurons

A₁₁ diencephalospinal dopamine (DA) neurons provide the major source of DA innervation to the spinal cord. DA in the dorsal and ventral horns modulates sensory, motor, nociceptive, and sexual functions. Previous studies from our laboratory revealed a sex difference in the density of DA innervation in the lumbar spinal cord. The purpose of this study was to determine whether sex differences in spinal cord DA are androgen dependent, influenced by adult or perinatal androgens, and whether a sex difference in the number of lumbar‐projecting A₁₁ neurons exists. Adult male mice have significantly higher DA concentrations in the lumbar spinal cord than either females or males carrying the testicular feminization mutation (tfm) in the androgen receptor (AR) gene, suggesting an AR‐dependent origin. Spinal cord DA concentrations are not changed following orchidectomy in adult male mice or testosterone administration to ovariectomized adult female mice. Administration of exogenous testosterone to postnatal day 2 female mice results in DA concentrations in the adult lumbar spinal cord comparable to those of males. Male mice display significantly more lumbar‐projecting A₁₁ DA neurons than females, particularly in the caudal portion of the A₁₁ cell body region, as determined by retrograde tract tracing and immunohistochemistry directed toward tyrosine hydroxylase. These results reveal an AR‐dependent sex difference in both the number of lumbar‐projecting A₁₁ DA neurons and the lumbar spinal cord DA concentrations, organized by the presence of androgens early in life. The AR‐dependent sex difference suggests thyat this system serves a sexually dimorphic function in the lumbar spinal cord. J. Comp. Neurol. 518:2423–2436, 2010. © 2010 Wiley‐Liss, Inc.     

Oestrogen‐Induced Activation of Preoptic Kisspeptin Neurones May be Involved in the Luteinising Hormone Surge in Male and Female Japanese Monkeys

The oestrogen‐induced luteinising hormone (LH) surge is evident in male primates, including humans, whereas male rodents never show the LH surge, even when treated with a preovulatory level of oestrogen. This suggests that the central mechanism governing reproductive hormones in primates is different from that in rodents. The present study aimed to investigate whether male Japanese monkeys conserve a brain mechanism mediating the oestrogen‐induced LH surge via activation of kisspeptin neurones. Adult male and female Japanese monkeys were gonadectomised and then were treated with oestradiol‐17β for 2 weeks followed by a bolus injection of oestradiol benzoate. Both male and female monkeys showed an oestrogen‐induced LH surge. In gonadectomised monkeys sacrificed just before the anticipated time of the LH surge, oestrogen treatment significantly increased the number of KISS1‐expressing cells in the preoptic area (POA) and enhanced the expression of c‐fos in POA KISS1‐positive cells of males and females. The oestrogen treatment failed to induce c‐fos expression in the arcuate nucleus (ARC) kisspeptin neurones in both sexes just prior to LH surge onset. Thus, kisspeptin neurones in the POA but not in the ARC might be involved in the positive‐feedback action of oestrogen that induces LH surge in male Japanese monkeys, as well as female monkeys. The present results indicate that oestrogen‐induced activation of POA kisspeptin neurones may contribute to the LH surge generation in both sexes. The conservation of the LH surge generating system found in adult male primates, unlike rodents, could be a result of the capability of oestrogen to induce POA kisspeptin expression and activation.     

Attempted suicide in Europe: rates,trend.S and sociodemographic characteristics of suicide attempters during the period 1989–1992. Results of the WHO/EURO Multicentre Study on Parasuicide

Schmidtke A, Bille-Brahe U, DeLeo D, Kerkhof A, Bjerke T, Crepet P, Haring C, Hawton K, Lönnqvist J, Michel K, Pommereau X, Querejeta I, Phillipe I, Salander-Renberg E, Temesvary B, Wasserman D, Fricke S, Weinacker B, Sampaio-Faria JG. Attempted suicide in Europe: rates, trends and sociodemographic characteristics of suicide attempters during the period 1989–1992. Results of the WHO/EURO Multicentre Study on Parasuicide. Acta Psychiatr Scand 1996: 93: 327–338. © Munksgaard 1996. The World Health Organization/EURO Multicentre Project on Parasuicide is part of the action to implement target 12 of the WHO programme, ‘Health for All by the Year 2000’, for the European region. Sixteen centres in 13 European countries are participating in the monitoring aspect of the project, in which trends in the epidemiology of suicide attempts are assessed. The highest average male age-standardized rate of suicide attempts was found for Helsinki, Finland (314/100000), and the lowest rate (45/100000) was for Guipuzcoa, Spain, representing a sevenfold difference. The highest average female age-standardized rate was found for Cergy-Pontoise, France (462/100000), and the lowest (69/100000) again for Guipuzcoa, Spain. With only one exception (Helsinki), the person-based suicide attempt rates were higher among women than among men. In the majority of centres, the highest person-based rates were found in the younger age groups. The rates among people aged 55 years or over were generally the lowest. For the majority of the centres, the rates for individuals aged 15 years or over decreased between 1989 and 1992. The methods used were primarily ‘soft’ (poisoning) or cutting. More than 50% of the suicide attempters made more than one attempt, and nearly 20% of the second attempts were made within 12 months after the first attempt. Compared with the general population, suicide attempters more often belong to the social categories associated with social destabilization and poverty.     

SSEA‐4 and YKL‐40 positive progenitor subtypes in the subventricular zone of developing human neocortex

The glycosphingolipid SSEA‐4 and the glycoprotein YKL‐40 have both been associated with human embryonic and neural stem cell differentiation. We investigated the distribution of SSEA‐4 and YKL‐40 positive cells in proliferative zones of human fetal forebrain using immunohistochemistry and double‐labeling immunofluorescence. A few small rounded SSEA‐4 and YKL‐40 labeled cells were present in the radial glial BLBP positive proliferative zones adjacent to the lateral ganglionic eminence from 12th week post conception. With increasing age, a similarly stained cell population appeared more widespread in the subventricular zone. At midgestation, the entire subventricular zone showed patches of SSEA‐4, YKL‐40, and BLBP positive cells. Co‐labeling with markers for radial glial cells (RGCs) and neuronal, glial, and microglial markers tested the lineage identity of this subpopulation of radial glial descendants. Adjacent to the ventricular zone, a minor fraction showed overlap with GFAP but not with nestin, Olig2, NG2, or S100. No co‐localization was found with neuronal markers NeuN, calbindin, DCX or with markers for microglial cells (Iba‐1, CD68). Moreover, the SSEA‐4 and YKL‐40 positive cell population in subventricular zone was largely devoid of Tbr2, a marker for intermediate neuronal progenitor cells descending from RGCs. YKL‐40 has recently been found in astrocytes in the neuron‐free fimbria, and both SSEA‐4 and YKL‐40 are present in malignant astroglial brain tumors. We suggest that the population of cells characterized by immunohistochemical combination of antibodies against SSEA‐4 and YKL‐40 and devoid of neuronal and microglial markers represent a yet unexplored astrogenic lineage illustrating the complexity of astroglial development. GLIA 2016;64:90–104     

Description of distributed features of the nestin‐containing cells in brains of adult mice: A potential source of neural precursor cells

The distribution of neural precursor cells (NPCs) in adult mice brain has so far not been described. Therefore, we investigated the distribution of NPCs by analyzing the nestin‐containing cells (NCCs) in distinct brain regions of adult nestin second‐intron enhancer‐controlled LacZ reporter transgenic mice through LacZ staining. Results showed that NCCs existed in various regions of adult mouse brain. In cerebellum, the greatest number of NCCs existed in cortex of the simple lobule, followed by cortex of the cerebellar lobule. In olfactory bulb, NCCs were most numerous in the granular cell layer, followed by the mitral cell layer and the internal plexiform, glomerular, and external plexiform layers. In brain nuclei (nu), NCCs were most numerous in the marginal nu, followed by the brainstem and diencephalon nu. NCCs in sensory nu of brainstem were more numerous than in motor nu, and NCCs in the dorsal of sensory nu were more numerous than in the ventral part. In brain ventricle systems, NCCs were largely distributed in the center of and external to the lateral ventricle, the inferior part of the third ventricle, the dorsal and inferior parts of the fourth ventricle, and the gray matter around the cerebral aqueduct. NCCs in the left vs. right brain were not significantly different. These data collectively indicate that NCCs were extensively distributed in the cerebellum and olfactory bulb, the partial nu of the marginal system, the partial brain nu adjacent to brain ventricle systems, the subependymal zone, and the cerebral cortex around the marginal lobe and were a potential source of NPCs. © 2009 Wiley‐Liss, Inc.     

Sex,drugs, and adult neurogenesis: Sex‐dependent effects of escalating adolescent cannabinoid exposure on adult hippocampal neurogenesis,stress reactivity,and amphetamine sensitization

Cannabinoid exposure during adolescence has adverse effects on neuroplasticity, emotional behavior, cognition, and reward sensitivity in adult rats. We investigated whether escalating doses of the cannabinoid receptor 1 (CB₁R) agonist, HU‐210, in adolescence would affect adult hippocampal neurogenesis and behavioral processes putatively modulated by hippocampal neurogenesis, in adult male and female Sprague‐Dawley rats. Escalating doses of HU‐210 (25, 50, and 100 µg/kg), or vehicle were administered from postnatal day (PND) 35 to 46. Animals were left undisturbed until PND 70, when they were treated with 5‐bromo‐2‐deoxyuridine (BrdU; 200 mg/kg) and perfused 21 days later to examine density of BrdU‐ir and BrdU/NeuN cells in the dentate gyrus. In another cohort, hypothalamic‐pituitary‐adrenal (HPA) axis reactivity to an acute restraint stress (30 min; PND 75) and behavioral sensitization to d‐amphetamine sulfate (1‐2 mg/kg; PND 105‐134) were assessed in adulthood. Adolescent HU‐210 administration suppressed the density of BrdU‐ir cells in the dentate gyrus in adult male, but not adult female rats. Adolescent HU‐210 administration also induced significantly higher peak corticosterone levels and reminiscent of the changes in neurogenesis, this effect was more pronounced in adult males than females. However, adolescent cannabinoid treatment resulted in significantly higher stereotypy scores in adult female, but not male, rats. Thus, adolescent CB₁R activation suppressed hippocampal neurogenesis and increased stress responsivity in adult males, but not females, and enhanced amphetamine sensitization in adult female, but not male, rats. Taken together, increased CB₁R activation during adolescence results in sex‐dependent, long‐term, changes to hippocampal structure and function, an effect that may shed light on differing vulnerabilities to developing disorders following adolescent cannabinoid exposure, based on sex. © 2013 Wiley Periodicals, Inc.     

Neonatal Kisspeptin is Steroid‐Independently Required for Defeminisation and Peripubertal Kisspeptin‐Induced Testosterone is Required for Masculinisation of the Brain: A Behavioural Study Using Kiss1 Knockout Rats

Rodents show apparent sex differences in their sexual behaviours. The present study used Kiss1 knockout (KO) rats to evaluate the role of kisspeptin in the defeminisation/masculinisation of the brain mechanism that controls sexual behaviours. Castrated adult Kiss1 KO males treated with testosterone showed no male sexual behaviours but demonstrated the oestrogen‐induced lordosis behaviours found in wild‐type females. The sizes of some of the sexual dimorphic nuclei of Kiss1 KO male rats are similar to those of females. Plasma testosterone levels at embryonic day 18 and postnatal day 0 (PND0) in Kiss1 KO males were high, similar to wild‐type males, indicating that perinatal testosterone is secreted in a kisspeptin‐independent manner. Long‐term exposure to testosterone from peripubertal to adult periods restored mounts and intromissions in KO males, suggesting that kisspeptin‐dependent peripubertal testosterone secretion is required to masculinise the brain mechanism. This long‐term testosterone treatment failed to abolish lordosis behaviours in KO males, whereas kisspeptin replacement at PND0 reduced lordosis quotients in Kiss1 KO males but not in KO females. These results suggest that kisspeptin itself is required to defeminise behaviour in the perinatal period, in cooperation with testosterone. Oestradiol benzoate treatment at PND0 suppressed lordosis quotients in Kiss1 KO rats, indicating that the mechanisms downstream of oestradiol work properly in the absence of kisspeptin. There was no significant difference in aromatase gene expression in the whole hypothalamus between Kiss1 KO and wild‐type male rats at PND0. Taken together, the present study demonstrates that both perinatal kisspeptin and kisspeptin‐independent testosterone are required for defeminisation of the brain, whereas kisspeptin‐dependent testosterone during peripuberty to adulthood is needed for masculinisation of the brain in male rats.     

Testosterone‐induced neuroendocrine changes in the medial preoptic area precede song activation and plasticity in song control nuclei of female canaries

Testosterone plays a key role in the control of seasonal changes in singing behavior and its underlying neural circuitry. After administration of exogenous testosterone, song quality and song control nuclei volumes change over the course of weeks, but song rate increases within days. The medial preoptic nucleus (POM) controls sexual motivation and testosterone action in POM increases sexually motivated singing. In this study, we investigated the time course of testosterone action in the song control nuclei and POM, at the gross anatomical and cellular level. Photosensitive female canaries were injected with BrdU to label newborn neurons. One day later they were transferred to a long‐day photoperiod and implanted with testosterone‐filled or empty implants. Brains and blood were collected 1, 2, 9 or 21 days later. Testosterone increased POM volume within 1 day, whereas the volume of song control nuclei increased significantly only on day 21 even if a trend was already observed for HVC on day 9. The density of newborn neurons in HVC, labeled by Bromodeoxyuridine (BrdU) and doublecortin, was increased by testosterone on days 9 and 21 although a trend was already detectable on day 2. In POM, testosterone increased the number and size of aromatase‐immunoreactive neurons already after 1 day. This rapid action of testosterone in POM supports its proposed role in controlling singing motivation. Although testosterone increased the number of newborn neurons in HVC rapidly (9, possibly 2 days), it is unlikely that these new neurons affect singing behavior before they mature and integrate into functional circuits.