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Differential timing of granule cell production during cerebellum development underlies generation of the foliation pattern

View Article: PubMed Central - PubMed

ABSTRACT

Background: The mouse cerebellum (Cb) has a remarkably complex foliated three-dimensional (3D) structure, but a stereotypical cytoarchitecture and local circuitry. Little is known of the cellular behaviors and genes that function during development to determine the foliation pattern. In the anteroposterior axis the mammalian cerebellum is divided by lobules with distinct sizes, and the foliation pattern differs along the mediolateral axis defining a medial vermis and two lateral hemispheres. In the vermis, lobules are further grouped into four anteroposterior zones (anterior, central, posterior and nodular zones) based on genetic criteria, and each has distinct lobules. Since each cerebellar afferent group projects to particular lobules and zones, it is critical to understand how the 3D structure of the Cb is acquired. During cerebellar development, the production of granule cells (gcs), the most numerous cell type in the brain, is required for foliation. We hypothesized that the timing of gc accumulation is different in the four vermal zones during development and contributes to the distinct lobule morphologies.

Methods and results: In order to test this idea, we used genetic inducible fate mapping to quantify accumulation of gcs in each lobule during the first two postnatal weeks in mice. The timing of gc production was found to be particular to each lobule, and delayed in the central zone lobules relative to the other zones. Quantification of gc proliferation and differentiation at three time-points in lobules representing different zones, revealed the delay involves a later onset of maximum differentiation and prolonged proliferation of gc progenitors in the central zone. Similar experiments in Engrailed mutants (En1−/+;En2−/−), which have a smaller Cb and altered foliation pattern preferentially outside the central zone, showed that gc production, proliferation and differentiation are altered such that the differences between zones are attenuated compared to wild-type mice.

Conclusions: Our results reveal that gc production is differentially regulated in each zone of the cerebellar vermis, and our mutant analysis indicates that the dynamics of gc production plays a role in determining the 3D structure of the Cb.

Electronic supplementary material: The online version of this article (doi:10.1186/s13064-016-0072-z) contains supplementary material, which is available to authorized users.

No MeSH data available.


The kinetics of accumulation of gcs is distinct in each lobule from P2 to P14. Graph showing the accumulation of gcs over time in each lobule calculated as follows from the percentage of ML labeled (i.e., the % of gcs that remained to be produced) after Tm administration at the indicated times in P28 animals: % of gcs produced = 100 %–% of labeled ML (n = 3 animals per time-point, error bars indicate SD). A third order polynomial equation was fitted to the data; the curves have been color coded to indicate which of the four cerebellar zones the lobule is associated with (green: anterior zone, blue: central zone, purple: posterior zone, and yellow: nodular zone)
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Fig3: The kinetics of accumulation of gcs is distinct in each lobule from P2 to P14. Graph showing the accumulation of gcs over time in each lobule calculated as follows from the percentage of ML labeled (i.e., the % of gcs that remained to be produced) after Tm administration at the indicated times in P28 animals: % of gcs produced = 100 %–% of labeled ML (n = 3 animals per time-point, error bars indicate SD). A third order polynomial equation was fitted to the data; the curves have been color coded to indicate which of the four cerebellar zones the lobule is associated with (green: anterior zone, blue: central zone, purple: posterior zone, and yellow: nodular zone)

Mentions: Examination of midline sections of P28 Atoh1-Tau mice administered Tm at P10 and stained for X-gal showed a striking difference in the density of marked gcs in the lobules corresponding to different zones, with the AZ and PZ having the least gcs marked (Fig. 1a). Consistent with this observation, quantification of the proportion of labeled ML area and density of gcs labeled at P8 revealed differences between lobules in particular zones (Fig. 2c). Given that more cells should be produced after the time of Tm administration in lobules with a higher proportion of the ML labeled, our finding raised the possibility that the timing of gc production differs more between zones than between lobules within a zone. We tested this idea by calculating the percentage of gcs that had already been produced on the day of Tm administration in Atoh1-Tau mice as the proportion of the ML area that was not labeled at P28 and generated a cumulative graph of gc production in each lobule (n = 3 or 4 mice per time point) (Fig. 3). For statistical tests (Additional file 1: Table S1A-B), we compared lobules 3, 7 and 10 as representing the AZ, CZ and NZ.Fig. 3


Differential timing of granule cell production during cerebellum development underlies generation of the foliation pattern
The kinetics of accumulation of gcs is distinct in each lobule from P2 to P14. Graph showing the accumulation of gcs over time in each lobule calculated as follows from the percentage of ML labeled (i.e., the % of gcs that remained to be produced) after Tm administration at the indicated times in P28 animals: % of gcs produced = 100 %–% of labeled ML (n = 3 animals per time-point, error bars indicate SD). A third order polynomial equation was fitted to the data; the curves have been color coded to indicate which of the four cerebellar zones the lobule is associated with (green: anterior zone, blue: central zone, purple: posterior zone, and yellow: nodular zone)
© Copyright Policy - OpenAccess
Related In: Results  -  Collection

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getmorefigures.php?uid=PMC5017010&req=5

Fig3: The kinetics of accumulation of gcs is distinct in each lobule from P2 to P14. Graph showing the accumulation of gcs over time in each lobule calculated as follows from the percentage of ML labeled (i.e., the % of gcs that remained to be produced) after Tm administration at the indicated times in P28 animals: % of gcs produced = 100 %–% of labeled ML (n = 3 animals per time-point, error bars indicate SD). A third order polynomial equation was fitted to the data; the curves have been color coded to indicate which of the four cerebellar zones the lobule is associated with (green: anterior zone, blue: central zone, purple: posterior zone, and yellow: nodular zone)
Mentions: Examination of midline sections of P28 Atoh1-Tau mice administered Tm at P10 and stained for X-gal showed a striking difference in the density of marked gcs in the lobules corresponding to different zones, with the AZ and PZ having the least gcs marked (Fig. 1a). Consistent with this observation, quantification of the proportion of labeled ML area and density of gcs labeled at P8 revealed differences between lobules in particular zones (Fig. 2c). Given that more cells should be produced after the time of Tm administration in lobules with a higher proportion of the ML labeled, our finding raised the possibility that the timing of gc production differs more between zones than between lobules within a zone. We tested this idea by calculating the percentage of gcs that had already been produced on the day of Tm administration in Atoh1-Tau mice as the proportion of the ML area that was not labeled at P28 and generated a cumulative graph of gc production in each lobule (n = 3 or 4 mice per time point) (Fig. 3). For statistical tests (Additional file 1: Table S1A-B), we compared lobules 3, 7 and 10 as representing the AZ, CZ and NZ.Fig. 3

View Article: PubMed Central - PubMed

ABSTRACT

Background: The mouse cerebellum (Cb) has a remarkably complex foliated three-dimensional (3D) structure, but a stereotypical cytoarchitecture and local circuitry. Little is known of the cellular behaviors and genes that function during development to determine the foliation pattern. In the anteroposterior axis the mammalian cerebellum is divided by lobules with distinct sizes, and the foliation pattern differs along the mediolateral axis defining a medial vermis and two lateral hemispheres. In the vermis, lobules are further grouped into four anteroposterior zones (anterior, central, posterior and nodular zones) based on genetic criteria, and each has distinct lobules. Since each cerebellar afferent group projects to particular lobules and zones, it is critical to understand how the 3D structure of the Cb is acquired. During cerebellar development, the production of granule cells (gcs), the most numerous cell type in the brain, is required for foliation. We hypothesized that the timing of gc accumulation is different in the four vermal zones during development and contributes to the distinct lobule morphologies.

Methods and results: In order to test this idea, we used genetic inducible fate mapping to quantify accumulation of gcs in each lobule during the first two postnatal weeks in mice. The timing of gc production was found to be particular to each lobule, and delayed in the central zone lobules relative to the other zones. Quantification of gc proliferation and differentiation at three time-points in lobules representing different zones, revealed the delay involves a later onset of maximum differentiation and prolonged proliferation of gc progenitors in the central zone. Similar experiments in Engrailed mutants (En1−/+;En2−/−), which have a smaller Cb and altered foliation pattern preferentially outside the central zone, showed that gc production, proliferation and differentiation are altered such that the differences between zones are attenuated compared to wild-type mice.

Conclusions: Our results reveal that gc production is differentially regulated in each zone of the cerebellar vermis, and our mutant analysis indicates that the dynamics of gc production plays a role in determining the 3D structure of the Cb.

Electronic supplementary material: The online version of this article (doi:10.1186/s13064-016-0072-z) contains supplementary material, which is available to authorized users.

No MeSH data available.