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The area of the molecular cell layer and granule cell layer of the cerebellum were measured in thionin/fuchsin acid stained sections in 8 serial sections.
IHC analysis of GM2 ganglioside revealed prominent accumulation throughout the granule cell layer and occasional storage within the molecular cell layer of end-stage untreated Npc1−/− mice cerebella.
To investigate the effect of Pam3CSK4 administration on the neonatal cerebellum, molecular cell layer and granule cell layer volumes were measured and the density of Purkinje cells was counted (Fig. 7A).
There was a significant decrease in the molecular cell layer volume in Pam3CSK4-treated mice but not LPS treated mice compared with saline-treated pups (Fig. 7B), while there were no differences in the granule cell layer volume (Fig. 7C) or the number of Purkinje cells between the three groups (Fig. 7D).
IHC analysis of GM2 in the cerebellum of untreated Npc1−/− mice revealed prominent accumulation throughout the granule cell layer and occasional storage in the molecular cell layer, while age-matched CD-treated Npc1−/− mice exhibited less accumulation in both the granule and molecular cell layers (data not shown).
In our study we have found that in the mature mouse brain, the Purkinje cell layer and the molecular cell layer (where the dendritic processes of Purkinje cells end) present strong GSK3β labeling, while the granular cell layer was devoid of immunostaining and non-axonal staining was observed.
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Most striking was the Cre staining pattern in the developing granular, Purkinje and molecular cell layers.
No obviously morphological changes could be found in the Purkinje cell layer and the molecular cell layers between control (Figure 4C) and tg mice (Figure 4D), where Purkinje cell dendrites are observable.
Analysis of cerebellar cortex from 22-day old untreated Npc1−/− mice revealed that nearly every Purkinje cell, as well as presumptive neurons in both the granule and molecular cell layers, showed evidence of cholesterol accumulation by filipin labeling (Fig. 3E).
Approximately half (see Table 2) of all Cre-positive cells co-labeled with NeuN (Fig. 5E F, K L, Fig. S4G I), suggesting a large population of neurons within the granular, Purkinje and molecular cell layers in which the PLP promoter is active at P4.
The highest upregulation of GFAP was found in the molecular cell layers in Kv3.1 KO mice.
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