Intrinsic and spontaneous neurogenesis in the postnatal slice culture of rat hippocampus

Abstract

Organotypic slice culture preserves the morphological and physiological features of the hippocampus of live animals for a certain time. The hippocampus is one of exceptional regions where neurons are generated intrinsically and spontaneously throughout postnatal life. We investigated the possibility that neurons are generated continuously at the dentate granule cell layer (GCL) in slice culture of the rat hippocampus. Using 5-bromodeoxyuridine (BrdU) labelling and retrovirus vector transduction methods, the phenotypes of the newly generated cells were identified immunohistochemically. At 4 weeks after BrdU exposure, BrdU-labelled cells were found in the GCL and were immunoreactive with a neuronal marker, anti-NeuN. There were fibrils immunoreactive with anti-glial fibrillary acidic protein (GFAP), an astrocyte marker, in the layer covering the GCL and occasionally encapsulated BrdU-labelled nuclei. When the newly divided cells were marked with the enhanced green fluorescent protein (EGFP) using a retrovirus vector, these cells had proliferative abilities throughout the following 4-week cultivation period. Four weeks after the inoculation, the EGFP-expressing cells consisted of various phenotypes of both early and late stages of differentiation; some were NeuN-positive cells with appearances of neurons in the GCL and some were immunoreactive with anti-Tuj1, a marker of immature neurons. Some EGFP-expressing cells were immunoreactive with anti-GFAP or anti-nestin, a marker of neural progenitors. The present study suggests that slice cultures intrinsically retain spontaneous neurogenic abilities for their cultivation period. The combination of slice culture and retrovirus transduction methods enable the newly divided cells to be followed up for a long period.

Human Immunodeficiency Virus Type 1 Vpr Inhibits Axonal Outgrowth through Induction of Mitochondrial Dysfunction

“…Not only the extracellular architect but also the synaptodendritic interactions are reconstituted in slice cul- tures (18). In addition, the presence of neurogenesis has been reported to happen in hippocampal slice cultures (28,45). For these reasons, slice cultures have been used widely to study the morphology and plasticity of the hippocampus.…”

“…Intrinsic and spontaneous neurogenesis are reported to take place at the DG in organotypic hippocampal slice cultures (28,45). To investigate the dysfunction of neuronal cell development in hippocampal slice cultures exposed to HIV-MDM or rVpr, we labeled endogenous precursor cells in the slices using an EGFP-expressing MLV vector (SR␣-EGFP).…”

“…To investigate the dysfunction of neuronal cell development in hippocampal slice cultures exposed to HIV-MDM or rVpr, we labeled endogenous precursor cells in the slices using an EGFP-expressing MLV vector (SR␣-EGFP). The virus vector was microinjected into the suprapyramidal region of the GCL as described before (28). Since MLV only infects dividing cells and its DNA is incorporated into the host DNA, only newly divided cells express EGFP.…”

“…Organotypic hippocampal slices were prepared from postnatal 7-day-old Wistar Hannover GALAS rats (CLEA Japan, Inc.) as previously described (28). The slices were cultured on a porous translucent membrane (Millicell-CM; Millipore) at 34°C for 14 days.…”

Voluntary Running Enhances Hippocampal Proliferation by Increasing Hippocampal NGF, BDNF, and IGF-1

“…Many studies have confirmed that neurogenesis continues in discrete regions of the adult brain, especially in the subgranular zone (SGZ) of the dentate gyrus (DG) and the subventricular zone (SVZ) of the lateral ventricles [1,2]. During adult hippocampal neurogenesis, newly generated cells can proliferate, differentiate, mature, and integrate into functional neural networks [3].…”