Research ReportPregnancy inhibits cell proliferation and neuroblast differentiation without neuronal damage in the hippocampal dentate gyrus in C57BL/6N mice
Introduction
The hippocampus is involved in some forms of learning and memory (Miyagawa et al., 2007). It is relatively susceptible to exogenous stress and shows remarkable degree of plasticity against stress (Blaise et al., 2008, Howland and Wang, 2008). Accordingly, progenitor cells in the dentate gyrus (DG), a part of hippocampus, retain the ability to proliferate and differentiate into neurons in all mammals, including non-human and human primates, forming functional synaptic inputs similar to those by mature (Song et al., 2002, Ambrogini et al., 2004, Llorens-Martín et al., 2006, Ngwenya et al., 2008, Snyder et al., 2009). There are some markers to visualize newly produced cells. Especially, Ki67 is a marker for all proliferating cells except early G1 phase (Gerdes et al., 1983, Cattoretti et al., 1992), and doublecortin (DCX) for progenitors differentiating into neurons (Brown et al., 2003).
There are neuronal changes affecting the dam during gestation and postpartum period (Numan, 2007, Brusco et al., 2008). The brain size has been shown to decrease across pregnancy, returning to preconception size after delivery in humans (Oatridge et al., 2002) and rodents (Galea et al., 2000). During pregnancy, there are dramatic fluctuations of many steroid hormones (Cameron and Gould, 1994, Ormerod et al., 2003, Casolini et al., 2007). Estradiol and corticosterone are regulators of adult neurogenesis. Estradiol initially (within 4 h) increases and subsequently suppresses (within 48 h) cell proliferation via adrenal steroids (Ormerod et al., 2003), while corticosteroids strongly inhibit cell proliferation in the hippocampus of adult female rodent (Cameron and Gould, 1994). Furthermore, corticosteroids are transferred to the pups by lactating (Casolini et al., 2007). In particular, corticosterone levels are significantly changed along gestation (Dalle et al., 1978), which is required. It is required for the prolactin receptor gene expression in the mammary gland of the late pregnant mouse (Mizoguchi et al., 1997) and corticosterone levels fall from day 17 of gestation until birth in the maternal plasma, thereafter remaining stable (Dalle et al., 1978).
There are studies focusing on changes of neurogenesis during gestation and postpartum period (Banasr et al., 2001, Shingo et al., 2003, Furuta and Bridges, 2005, Rolls et al., 2008), but only upon one time point or 7–10 days after gestation. In the present study, we investigated on (i) cell death using terminal deoxynucleotidyl dUTP nick-end labeling (TUNEL) and Fluoro-jade B (F-J B) staining and NeuN immunohistochemistry, (ii) cell proliferation and (iii) neuroblast differentiation using immunohistochemistry for Ki67 and DCX, and western blot for DCX in the DG at late stages of gestation in C57BL/6N mice (GD 14.5, 16.5 and 18.5).
Section snippets
Effects of pregnancy on mature neurons
In the control group, NeuN-immunoreactive (+) mature neurons were well detected in the hippocampus including the DG (Figs. 1A and B). In the GD14.5, GD16.5 and GD18.5 groups, the number and the distribution pattern of NeuN+ neurons were similar to the control group (Figs. 1C–H).
Effects of pregnancy on neuronal death
In the age-matched control group, TUNEL or F-J B positive (+) cells were rarely observed in the DG (Figs. 2A and B). In GD14.5, GD16.5 and GD18.5 groups, only few TUNEL+ or F-J B+ cells were detected in the DG (Figs. 2
Discussion
We studied the effects of pregnancy on (i) mature neurons using NeuN immunohistochemistry, (ii) cell death using TUNEL and F-J B staining, and (iii) cell proliferation and (iv) neuroblast differentiation using Ki67 and DCX immunohistochemistry, respectively, in the hippocampal DG at various time points during gestation in C57BL/6N mice.
In the present study, we could not find any significant change in numbers of mature neurons in any of the subregions during gestation based on NeuN
Experimental animals
Eight-week-old male and female C57BL/6N mice were purchased from Orient Bio Inc. (Seongnam, South Korea) and mated. The next morning, the plug was observed and considered as gestation day 0.5 (GD0.5). Pregnant mice were housed in a conventional state under adequate temperature (23 °C) and humidity (60%) control with a 12-h light/12-h dark cycle, and free access to food and water. The procedures for handling and caring for animals adhered to the guidelines that are in compliance with the current
Acknowledgments
The authors would like to thank Mr. Seok Han, Mr. Seung Uk Lee and Ms. Hyun Sook Kim for their technical help in this study. This research was supported by the National Research Foundation of Korea (NRF) funded by the Ministry of Education, Science and Technology (2009-0084089) and by a grant (20090K001290) from Brain Research Center of the 21st Century Frontier Research Program funded by the Ministry of Education, Science and Technology, the Republic of Korea.
References (36)
- et al.
Learning may reduce neurogenesis in adult rat dentate gyrus
Neurosci. Lett.
(2004) - et al.
Analysis of neurogenesis and programmed cell death reveals a self-renewing capacity in the adult rat brain
Neurosci. Lett.
(2000) - et al.
Neonatal isolation stress alters bidirectional long-term synaptic plasticity in amygdalo-hippocampal synapses in freely behaving adult rats
Brain Res.
(2008) - et al.
Plasma hormonal profiles and dendritic spine density and morphology in the hippocampal CA1 stratum radiatum, evidenced by light microscopy, of virgin and postpartum female rats
Neurosci. Lett.
(2008) - et al.
Adult neurogenesis is regulated by adrenal steroids in the dentate gyrus
Neuroscience
(1994) - et al.
Gestation-induced cell proliferation in the rat brain
Brain Res. Dev. Brain Res.
(2005) - et al.
Spatial working memory and hippocampal size across pregnancy in rats
Horm. Behav.
(2000) - et al.
Synaptic plasticity in learning and memory: stress effects in the hippocampus
Prog. Brain Res.
(2008) - et al.
Learning during motherhood: a resistance to stress
Horm. Behav.
(2006) - et al.
Arrested neuronal proliferation and impaired hippocampal function following fractionated brain irradiation in the adult rat
Neuroscience
(2003)
Memory impairment associated with a dysfunction of the hippocampal cholinergic system induced by prenatal and neonatal exposures to bisphenol-A
Neurosci. Lett.
Corticosterone is required for the prolactin receptor gene expression in the late pregnant mouse mammary gland
Mol. Cell. Endocrinol.
Radiation-induced impairment of hippocampal neurogenesis is associated with cognitive deficits in young mice
Exp. Neurol.
Fluoro-Jade B: a high affinity fluorescent marker for the localization of neuronal degeneration
Brain Res
Serotonin mediates oestrogen stimulation of cell proliferation in the adult dentate gyrus
Eur. J. Neurosci.
Transient expression of doublecortin during adult neurogenesis
J. Comp. Neurol.
Maternal exposure to low levels of corticosterone during lactation protects the adult offspring against ischemic brain damage
J. Neurosci.
Monoclonal antibodies against recombinant parts of the Ki-67 antigen (MIB 1 and MIB 3) detect proliferating cells in microwave-processed formalin-fixed paraffin sections
J. Pathol.
Cited by (13)
Less can be more: Fine tuning the maternal brain
2022, Neuroscience and Biobehavioral ReviewsCitation Excerpt :Galea and McEwen (1999) reported that wild pregnant meadow voles captured during the breeding season had decreased levels of hippocampal cell proliferation compared to non-pregnant female meadow voles captured during the non-breeding season (Galea and McEwen, 1999). In addition, late pregnancy in the rat (day 21) and mouse (day 16.5) also decreased cell proliferation when measured using an endogenous marker, Ki67, compared to non-pregnant females (Kim et al., 2010c; Pawluski et al., 2015, 2020). However, early pregnancy (day 1 or 7) in the rat has not been associated with any changes in cell proliferation in the hippocampus (Pawluski et al., 2010, 2011; Shingo et al., 2003) suggesting an effect of pregnancy on hippocampal plasticity.
The long and short term effects of motherhood on the brain
2019, Frontiers in NeuroendocrinologyCitation Excerpt :In the mid-postpartum, primiparous female rats have lower brain weight and hippocampal volume compared to nulliparous females (Hillerer et al., 2014). Studies have also examined changes in neuroplasticity markers during pregnancy and postpartum in the hippocampus (Banasr et al., 2001; Eid et al., 2019; S.K. Kim et al., 2010; Leuner et al., 2010; Leuner and Sabihi, 2016; Pawluski et al., 2010; Pawluski and Galea, 2007; Rolls et al., 2008; Table 1), the dorsal raphe nucleus (Holschbach and Lonstein, 2017), and subventricular zone (SVZ; Furuta and Bridges, 2005; Shingo et al., 2003). In the hippocampus during gestation (GD13) and postpartum (PD8 and PD30), the number of immature neurons (doublecortin (DCX) expression) is lower in primiparous compared to nulliparous female rats and there is a parallel reduction in cell proliferation during gestation and early postpartum (Eid et al., 2019).
The birth of new neurons in the maternal brain: Hormonal regulation and functional implications
2016, Frontiers in NeuroendocrinologyCitation Excerpt :In accordance with the rat studies, Shingo et al. (2003) also showed no change in hippocampal cell proliferation when mice were injected with BrdU during early pregnancy (GD7). However, other work (Kim et al., 2010) using the cell proliferation marker Ki67 did uncover fewer proliferating cells in the mouse DG at later gestational time points (GD14.5, GD16.5 and GD18.5) suggesting that alterations in cell proliferation during pregnancy occur in a species- and time-dependent manner. Effects of pregnancy on cell survival and differentiation have also been demonstrated.
Parenting Behavior
2015, Knobil and Neill's Physiology of Reproduction: Two-Volume Set