Involvement of postnatal apoptosis on sex difference in number of cells generated during late fetal period in the sexually dimorphic nucleus of the preoptic area in rats

doi:10.1016/j.neulet.2012.04.017

Neuroscience Letters

Volume 516, Issue 2, 16 May 2012, Pages 290-295

https://doi.org/10.1016/j.neulet.2012.04.017 Get rights and content

Abstract

Postnatal apoptosis is involved in formation of the sex difference in neuron number of the sexually dimorphic nucleus of the preoptic area (SDN-POA) in rats. In this study, we examined the origin of neurons that die with apoptosis on the postnatal period to exhibit the sex difference in neuron number of the SDN-POA. First, we measured the number of cells that were labeled with 5-bromo-2′-deoxyuridine (BrdU) on embryonic day (ED) 17, ED18, and ED19 in the SDN-POA of rats on postnatal day (PD) 4 and PD8. The SDN-POA had many more cells labeled with BrdU on ED17 and ED18 than those on ED19. Significantly fewer cells labeled with BrdU on ED18 in the female SDN-POA from PD4 to PD8 resulted in a significant sex difference in the number at PD8. Next, combination analyses of BrdU-labeling and immunohistochemistry for single-stranded DNA (ssDNA), an apoptotic marker, were succeeded to investigate whether SDN-POA neurons generated during ED17–18 were removed by apoptosis. Many more ssDNA-immunoreactive cells that had been labeled with BrdU during ED17–18 were found in the SDN-POA of PD8 females, but few in the SDN-POA of PD8 males and PD4 females and males. These results suggest that the sex difference in the number of SDN-POA neurons generated during the late fetal period was caused by postnatal apoptosis.

Highlights

► SDN-POA has many more neurons in male rats than female rats in adulthood. ► Postnatal apoptosis is involved in the sex difference in SDN-POA neuron number. ► We examined the origin of SDN-POA neurons that die with postnatal apoptosis. ► Female rats had more neurons born on ED17–18 and died with apoptosis on PD8. ► Male rats had few neurons born on ED17–18 and died with apoptosis on PD8.

Introduction

The sexually dimorphic nucleus of the preoptic area (SDN-POA) of male rats is five to seven times larger and has more neurons than that in female rats [10], [11]. The physiological functions of the SDN-POA have yet to be clearly determined. However, lesions of the SDN-POA delay the onset and decrease the incidence of male sexual behavior in sexually naive male rats [7] but not in sexually experienced male rats [3], suggesting that the SDN-POA is involved in modulating sexuality in male rats.

A sex difference in the number of cells dying by apoptosis was found in the SDN-POA of rats in the early postnatal period. The number of apoptotic cells in the SDN-POA was larger in female rats than male rats between postnatal day (PD) 7 and PD10 (day of birth = PD1) [6]. The number of apoptotic cells in the female SDN-POA peaks on PD8 [5]. Thus, the sex difference in number of apoptotic cells in the postnatal period is negatively correlated with the sex difference in number of neurons in adulthood, supporting the idea that the control of neuron number by postnatal apoptosis is involved in formation of the morphological sex differences in the SDN-POA. In the SDN-POA of postnatal rats, the level of antiapoptotic Bcl-2 expression is higher in males than females [16], [17], whereas females had stronger expression of proapoptotic Bax [17]. The sex differences in Bcl-2 and Bax are followed by the sex difference in apoptosis induced by caspases, because the number of cells containing active caspase-3 in the SDN-POA of PD8 female rats was greater than that in same-aged males [17]. Thus, molecular mechanisms by which the sex differences in apoptosis arise in the SDN-POA of postnatal rats were partially uncovered. However, the origin of neurons that exhibit the sex difference in apoptosis has yet been clearly determined.

Neurons that comprise the SDN-POA are generated between embryonic day 15 (ED15) and ED19 (ED1 = day of fertilization) in rats, and generation of SDN-POA neurons reaches a peak level during ED18–19 [4]. The number of neurons generated on ED18 in the male SDN-POA remains elevated during PD2-30, but the number in the female SDN-POA decreases dramatically between PD4 and PD7, reaching a minimal number by PD10 [8]. In addition, the number of SDN-POA neurons born on ED17 is larger in male rats than in female rats by PD30 [12]. These findings indicate that SDN-POA neurons generated on the late fetal period exhibit the sex difference in their number, although the mechanism responsible for the sex difference is unclear. We hypothesized that the sex difference is caused by migration of neurons to exit from the SDN-POA or loss of neurons by apoptosis within the SDN-POA. In the current study, we tested the hypothesis by 5-bromo-2′-deoxyuridine (BrdU)-labeling method and detection of apoptosis in BrdU-labeled cells.

Section snippets

Animals

Pregnant Wistar rats were purchased as breeding stock (Japan Laboratory Animals, Inc., Tokyo, Japan). Fertilization of these animals had been checked by the presence of a vaginal plug, and the day of fertilization was designed ED1. Pregnant mothers were housed in a 12-h light/12-h dark cycle at 23 °C with free access to water and food. They were checked daily to identify the day of parturition, and the day when pups were found was designated PD1. Male and female pups at PD4 and PD8 were used for

The volume of the SDN-POA in postnatal rats

The SDN-POA of males became larger with age, although the size of the SDN-POA of females did not change (Fig. 2A). The volume of the SDN-POA was significantly affected by age (F_1,42 = 4.42, p < 0.05), sex (F_1,42 = 6.70, p < 0.05), and by the interaction between age and sex (F_1,42 = 17.6, p < 0.0005). The volume of the SDN-POA in PD8 males was significantly larger than that in PD8 females and in PD4 males (Fig. 2B). There was no significant difference in SDN-POA volume between PD4 and PD8 females.

The number of BrdU-labeled cells in the SDN-POA of postnatal rats

Many cells

Discussion

Labeling cells with BrdU in the current study showed that the number of cells born on ED18 significantly decreased in the SDN-POA of females from PD4 to PD8. The decrease in the number of BrdU-labeled cells in the female SDN-POA was responsible for the significant difference in the cell number between sexes at PD8, although no sex difference in the number of BrdU-labeled cells was found on PD4. These results suggest that the same number of neurons is generated in both sexes at ED18 in the

Acknowledgments

This work was supported in part by the Project Research Grant from the Research Management Bureau of Saitama University to S.T. (A10-20 and A11-34) and by Grants-in-Aid for Scientific Research from the Ministry of Education, Science, Culture and Sports of Japan (23310043) to S.T.

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Involvement of postnatal apoptosis on sex difference in number of cells generated during late fetal period in the sexually dimorphic nucleus of the preoptic area in rats

Abstract

Highlights

Introduction

Section snippets

Animals

The volume of the SDN-POA in postnatal rats

The number of BrdU-labeled cells in the SDN-POA of postnatal rats

Discussion

Acknowledgments

Neuroscience Research

Brain Research Bulletin

Brain Research

Brain Research Bulletin

Brain Research

Neuroscience Letters

Neuroscience Research

Estimation of nuclear population from microtome sections

Anatomical Record

Development of the preoptic area: time and site of origin, migratory routes, and settling patterns of its neurons

Journal of Comparative Neurology