Draxin, a repulsive axon guidance protein, is involved in hippocampal development
Introduction
The hippocampus is an archicortical structure located at the caudomedial edge of the neocortex in mice (Amaral and Witter, 1995). The adult hippocampus forms an anatomically unique structure, with the longitudinal axis adopting a ‘C’ shape that is divided into distinct fields of the dentate gyrus (DG) and the CA1, CA2, and CA3 fields of Ammon's horn. This structure is one of the best-analyzed parts of the brain due to its involvement in the higher functions of learning and memory and in various neural diseases (Kandel, 2000). The hippocampus is absolutely necessary for making new memories and is affected severely and before other parts of the cortex in Alzheimer's disease. The hippocampus also seems to be involved in severe mental illnesses; it is reduced in size in both cases of schizophrenia and depression (Sapolsky, 2001). It is particularly vulnerable to ischemia, which selectively affects CA1 pyramidal neurons and destroys hippocampal circuitry by inducing cell death (Pulsinelli et al., 1982). Abnormal development of the hippocampus gives rise to pathological conditions involving cognitive dysfunction and seizures.
Many factors underlying the proper assembly of the hippocampus have already been identified, such as Wnt signaling pathway (Lee et al., 2000, Galceran et al., 2000, Zhou et al., 2004); the homeodomain protein Emx2 (Tole et al., 2000); fibroblast growth factor receptor 1 (Fgfr1) (Ohkubo et al., 2004); the proneural protein neurogenin 2 (Ngn2) (Galichet et al., 2008). Thus, various molecules such as growth and transcription factors are known to control the development of the hippocampus; however, such molecules as axon guidance proteins (Hinck, 2004) also have the potential to be involved in this development.
Draxin is a chemorepulsive axon guidance molecule required for development of the spinal cord and all forebrain commissures: the corpus callosum, hippocampal commissures and anterior commissures (Islam et al., 2009). Mouse draxin expression has been observed in many regions of the brain, including the hippocampus. In the present study, we examined the effects of draxin gene deprivation in hippocampal development. We found that draxin is widely expressed in the hippocampus by different cell types and that draxin knockout mice show abnormal hippocampal morphology and a loss of dentate granule cells.
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Animal care and treatments
All experiments were carried out in accordance with the guidelines for the care and use of animals approved by the Animal Care and Use Committee (Kumamoto University). All efforts were made to minimize the number of animals used and their suffering. All of the mice used in this experiment were obtained either from a local company (wild type mice) or from a colony in an animal center in Kumamoto University (draxin knockout mice/β-galactosidase (β-gal) knockin and knockout mice). They were
Draxin is widely expressed in the developing hippocampus
To address the role of Draxin in development of the hippocampus, we first characterized the expression of draxin mRNA in the developing hippocampus of draxin/LacZ heterozygous mice. In situ hybridization and β-galactosidase staining was performed to detect expression from the draxin allele and revealed that draxin mRNA was widely expressed in the hippocampus, including the hippocampal ventricular zone (VZ), CA fields and the dentate gyrus at embryonic and early postnatal stages (Fig. 1A-D). The
Discussion
Formation of the hippocampal DG in mice begins at around E15 in the dorsomedial part of the telencephalic vesicles. The DG primordium is initially populated by Cajal–Retzius cells and radial glia that are likely to participate to its histogenesis (Rickmann et al., 1987, Del Rio et al., 1997, Alcantara et al., 1998, Borrell et al., 1999). The portion of the hippocampal neuroepithelium that constitutes the DG primordium, also called the primary matrix, contains stem/progenitor cells that give
Acknowledgements
We thank Dr. Kumimasa Ohta for comments on the manuscript. We thank the Developmental Studies Hybridoma Bank for antibodies. This work was supported by grants-in-aid from the Ministry of Education, Science, Sports and Culture of Japan, by the 21st Century COE Program and by the Global COE Program (Cell Fate Regulation Research and Education Unit), MEXT, Japan.
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