Anat Cell Biol.  2010 Sep;43(3):241-251. 10.5115/acb.2010.43.3.241.

Migratory defect of mesencephalic dopaminergic neurons in developing reeler mice

• Kang WY
• Kim SS
• Cho SK
• Kim S
• Suh-Kim H
• Lee YD

• KMID: 2168879
• DOI: http://doi.org/10.5115/acb.2010.43.3.241

Abstract

Reelin, an extracellular glycoprotein has an important role in the proper migration and positioning of neurons during brain development. Lack of reelin causes not only disorganized lamination of the cerebral and cerebellar cortex but also malpositioning of mesencephalic dopaminergic (mDA) neurons. However, the accurate role of reelin in the migration and positioning of mDA neurons is not fully elucidated. In this study, reelin-deficient reeler mice exhibited a significant loss of mDA neurons in the substantia nigra pars compacta (SNc) and a severe alteration of cell distribution in the retrorubal field (RRF). This abnormality was also found in Dab1-deficinet, yotari mice. Stereological analysis revealed that total number of mDA neurons was not changed compared to wild type, suggesting that the loss of mDA neurons in reeler may not be due to the neurogenesis of mDA neurons. We also found that formation of PSA-NCAM-positive tangential nerve fibers rather than radial glial fibers was greatly reduced in the early developmental stage (E14.5) of reeler. These findings provide direct evidence that the alteration in distribution pattern of mDA neurons in the reeler mesencephalon mainly results from the defect of the lateral migration using tangential fibers as a scaffold.

Keyword

Reelin; dopaminergic neurons; radial glia; tangential fibers; neuronal migration

MeSH Terms

Animals
Brain
Cerebellar Cortex
Dopaminergic Neurons
Glycoproteins
Mesencephalon
Mice
Mice, Neurologic Mutants
Nerve Fibers
Neurogenesis
Neurons
Substantia Nigra
Glycoproteins

Figure

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  Fig. 1 Abnormal positioning of mDA neurons in reeler and yotari mice at P0. (A) DA cells labeled with TH-immunoractivity are significantly reduced in the substantia nigra pars compacta (SNc) and lateralis (SNL) of reeler and yotari mouse (arrows in b and c). In retrorubral field (RRF), unlike to wild type, mDA neurons are diffusely distributed in the region between the vental tegmental area (VTA) and the RRF of reeler and yotari mice (asterisks in e and f). (B) Characteristically, the ectopic cluster of mDA neurons in the lateral border of the VTA is observed in reeler and yotari (arrows in e and f). Scale bar=50 µm (a, d in A, d in B), 500 µm (a in B).

• 

  Fig. 2 Stereologic analysis of the numbers of mDA neurons in the VTA, SNc, and RRF at P7. (A) Borders of the VTA, SNc, and RRF were designated based on the mouse brain atlas of Paxinos & Franklin (2001). To encompass the full rostrocaudal extent of the midbrain DA neurons, 12 sections from 1 : 3 series were analyzed for each brain. (B) In reeler mice, the number of DA neurons is significantly reduced in the SNC and RRF, but rather VTA cells are increased. It is noticed that the total number of mDA neurons in these major cell groups of reeler is almost same as that in wild type. (C) The pie charts show the decrease in the percentage of mDA neurons in the SNc and RRF of reeler. VTA, vental tegmental area; SNc, substantia nigra pars compacta; RRF, retrorubral field. Data were obtained from three experiments; values are means±SEM. 12 sections from a 1 in 3 series were analyzed per brain. Statistical analysis was performed using t-test compared with wild type. n.s, not significant.

• 

  Fig. 3 Immunofluorescence staining for TH (red) and RC2 (green) in the ventral mesencephalon at E14.5 (A, B) and E16.5 (C, D). In the ventral mesencephalon at E14.5, RC2-positive cells and fibers in upper (a) and lower (b) regions of the midline and the lateral region (c) do not show a distinct difference between wild type and reeler. However, at E16.5, RC2 immunoreactivity is markedly reduced in reeler compared to E14.5. Especially, a poor formation of RC2-positive radial glial fibers extending to the pial surface is noticed in reeler mutant (c in D). The lack of distribution of mDA neurons in the lateral region of the ventral mesencephalon at E16.5 is noted (arrows). Scale bar=500 µm (A) and 50 µm (a, b, c in A).

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  Fig. 4 Immunofluorescence staining for BLBP (green) in the ventral mesencephalon of E14.5 (A, B) and E16.5 (C, D). At E14.5, BLBP-positive radial fibers in reeler do not show a difference compared to wild type. However, at E16.5, the number of BLBP-positive radial glial fibers in reeler is greatly reduced, parallel to RC2-positive radial glial fibers (c of D in Fig. 3). Scale bar=500 µm (A, low magnification image) and 50 µm (enlarged image of A).

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  Fig. 5 Immunofluorescence staining for TH (red) and PSA-NCAM (green) in the RRF, caudal regions of the mesencephalon at E14.5. This figure shows different two levels of the RRF (A, C and B, D). Although, formation of RC2- or BLBP-positive radial fibers in reeler at the same stage was not different from wild type (as shown in Fig. 3 and Fig. 4), PSA-NCAM-positive tangential nerve fibers in the RRF are weakly organized compared to wild type. Scale bar: 500 µm (A, low magnification image) and 50 µm (enlarged image of A).

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  Fig. 6 Immunostaining of radial glial fibers and tangential fibers in E16.5 reeler and yotari mice. The reduction of radial glial fibers formation in reeler is also seen in yotari mice (B, C and E, F). Furthermore, poor development of PSA-NCAM-positive tangential nerve fibers is noticed in both reeler and yotari mutants (arrows in H, I). Scale bar=50 µm (A, D, G).

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