Elsevier

Neuroscience

Volume 136, Issue 1, 2005, Pages 343-355
Neuroscience

Systems neuroscience
Identification and characterization of two neurogenic zones in interface organotypic hippocampal slice cultures

https://doi.org/10.1016/j.neuroscience.2005.07.058Get rights and content

Abstract

Neurogenesis plays a role in many physiological (memory formation) and pathological (stroke, depression) processes. However the mechanisms of postnatal stem cell proliferation and neurogenesis are still poorly understood. We characterized early neurogenesis in vitro in rat organotypic hippocampal slice cultures. Proliferation was assessed by bromodeoxyuridine incorporation, neurogenesis by bromodeoxyuridine-double labeling with doublecortin or β-III tubulin. We showed for the first time that in addition to the dentate gyrus organotypic hippocampal slice cultures include a second neurogenic zone: the posterior periventricle, which is a part of the lateral ventricle wall. This structure lining the stratum oriens contained Nestin+ precursors. We could identify morphological and functional differences between dentate gyrus and posterior periventricle precursor populations. Our data demonstrate that basic fibroblast growth factor treatment induced a fast but short-lasting neurogenic response in the dentate gyrus while the posterior periventricle showed a more pronounced and long lasting neurogenic effect of basic fibroblast growth factor. Thus two neurogenic zones with different neurogenic properties were identified in organotypic hippocampal slice cultures.

Section snippets

OHC

Organotypic hippocampal cultures were prepared according to the standard interface method (Stoppini et al., 1991). Hippocampal slices were prepared from postnatal days 7–9 (P7-9) Wistar rats (Harlan Winkelmann GmbH, Borchen, Germany). After decapitation, the brain was removed, hippocampi were dissected and transversely sliced into a 350μm thickness on a McIlwain tissue chopper (The Mickle Laboratory Engineering Co., Guildford, UK). The slices were then carefully separated with two pairs of fine

Identification and in situ distribution of CNS cell types in OHC

OHC were prepared from P7-9 Wistar rats and cultivated for two weeks before experiments were performed. For the general experimental setup see Fig. 1A. To characterize the three-dimensional structure of OHC, different cell types were detected by immunohistochemistry in unsectioned (“whole mounts”) and cross-sectioned 14 DIV slices. Astrocytes (GFAP+), precursor cells (Nestin+), migrating neuroblasts (DCX+), young and mature neurons (β-III tubulin+), mature neurons (NeuN+) and microglia (IB4+)

Discussion

The aim of this work was to investigate the early neurogenesis in vitro in interface OHC. Hippocampal slices were prepared from P7-9 rats and maintained in vitro for 14 d before the proliferating cells were labeled with BrdU.

The initial trauma during preparation and subsequent cultivation stimulated astrocytes to proliferate and form a glial barrier ensheathing the slice (GE). The presence of activated astrocytes expressing Nestin and activated microglia indicated a protective function of the

Acknowledgments

Work was supported by graduate program “Biologische Grundlagen von Erkrankungen des Nervensystems,” Medizinische Fakultät, Otto-von Guericke-Universität, Magdeburg and European Grant QLK3-CT-2001-00407.

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