Abstract
Neurons are added throughout life to the dentate gyrus of the hippocampus of the mammalian brain. Progenitors residing in the dentate gyrus progress through three distinct stages of adult neurogenesis: proliferation, survival and differentiation. One of the most potent factors which regulates adult neurogenesis is adrenal-derived glucocorticoids. Raised levels of glucocorticoids suppress progenitor division, while removal of glucocorticoids by adrenalectomy stimulates proliferation of these cells in the dentate gyrus. We have recently reported that both pre- and post-mitotic corticoid environments powerfully regulate survival of progenitor cells in a time-dependent manner. However, it is unknown if glucocorticoids alter the process of neuronal differentiation, since not all of the newly-formed cells acquire a neuronal fate during development. Here we employ triple immuno-fluorescence staining techniques to phenotype surviving progenitor cells 28 days after labeling. Results show that high levels of corticosterone (the major glucocorticoid in rodents) either before or after progenitor labeling discouraged the acquisition of neuronal fate. Similar to its effect on survival, post-mitotic corticosterone also regulates neuronal differentiation in a time-dependent fashion, but this action is most prominent from around 19–27 days after the cells were born. In contrast, a corticoid-free environment either before or after progenitor proliferation did not affect neuronal differentiation. Combining these data with previous survival data obtained from the same animals allowed us to estimate the total number of neurons formed resulting from different corticoid treatments. Raised corticosterone significantly reduced neuronal production while adrenalectomy resulted in significantly higher number of neurons in the adult male rat hippocampus.
Highlights
Neurogenesis in the adult brain is well established since its first report 40 years ago (Altman and Das, 1966)
Corticosterone had no effect on differentiation into GFAPstained cells (GFAP: F1,11ϭ0.06, Pϭ0.82)
To reveal whether this represented a delay in maturation, day 28 sections were triple-stained for BrdU/DCX/GFAP
Summary
Animals All procedures were carried out under Home Office (UK) license and guidelines. These stipulate the minimal use of animals and procedures required to minimize pain and suffering. Animals were given a choice of 0.9% saline and water to replenish salt loss. 5-bromodeoxyuridine (BrdU) (Sigma), a thymidine analog incorporated into dividing cells during DNA synthetic phase (S-phase) of the cell cycle, was dissolved in 0.9% saline and injected i.p. Basal levels of corticosterone were replaced with s.c. implants of 1:1 corticosterone/cholesterol (Sigma) melted and molded into pellets weighing 200 –220 mg. To increase plasma corticosterone above ‘basal’ levels, animals were given additional s.c. injections of corticosterone suspended in peanut oil at 40 mg/kg/day, a dose shown to reduce neurogenesis. Reference plasma corticosterone levels of these different paradigms can be obtained in our recent publication (Wong and Herbert, 2005)
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