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Morphological and Phagocytic Profile of Microglia in the Developing Rat Cerebellum(1,2,3).

Perez-Pouchoulen M, VanRyzin JW, McCarthy MM - eNeuro (2015)

Bottom Line: We found that microglial morphology changed from amoeboid to ramified during the first 3 postnatal weeks in a region specific manner.At P17 males showed an approximately twofold increase in microglia with thin processes compared with females.Our findings indicate a continuous process of microglial maturation and a nonuniform distribution of microglia in the cerebellar cortex that implicates microglia as an important cellular component of the developing cerebellum.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Pharmacology, University of Maryland School of Medicine , Baltimore, Maryland 21201.

ABSTRACT
Microglia are being increasingly recognized as playing important roles in neurodevelopment. The cerebellum matures postnatally, undergoing major growth, but the role of microglia in the developing cerebellum is not well understood. Using the laboratory rat we quantified and morphologically categorized microglia throughout the vermis and across development using a design-based unbiased stereology method. We found that microglial morphology changed from amoeboid to ramified during the first 3 postnatal weeks in a region specific manner. These morphological changes were accompanied by the sudden appearance of phagocytic cups during the third postnatal week from P17 to P19, with an approximately fourfold increase compared with the first week, followed by a prompt decline at the end of the third week. The microglial phagocytic cups were significantly higher in the granular layer (∼69%) than in the molecular layer (ML; ∼31%) during a 3 d window, and present on ∼67% of microglia with thick processes and ∼33% of microglia with thin processes. Similar proportions of phagocytic cups associated to microglia with either thick or thin processes were found in the ML. We observed cell nuclei fragmentation and cleaved caspase-3 expression within some microglial phagocytic cups, presumably from dying granule neurons. At P17 males showed an approximately twofold increase in microglia with thin processes compared with females. Our findings indicate a continuous process of microglial maturation and a nonuniform distribution of microglia in the cerebellar cortex that implicates microglia as an important cellular component of the developing cerebellum.

No MeSH data available.


Related in: MedlinePlus

Microglial phagocytosis in the postnatal developing cerebellum. A, The highest density of phagocytic cups was observed during the third postnatal week at P17 (***p < 0.000 compared with P5, P7, P10, P12, P14, and P21; n = 4, 2 males + 2 females for each group). Data are expressed as mean ± SEM. B, Phagocytic cups exhibited by microglia (red arrows) in the developing cerebellum at P17. Scale bar, 100 µm. C, Microglia with phagocytic cups (top) or microglia without phagocytic cups (bottom row) at different time points during postnatal development. Scale bars, 25 µm.
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Figure 3: Microglial phagocytosis in the postnatal developing cerebellum. A, The highest density of phagocytic cups was observed during the third postnatal week at P17 (***p < 0.000 compared with P5, P7, P10, P12, P14, and P21; n = 4, 2 males + 2 females for each group). Data are expressed as mean ± SEM. B, Phagocytic cups exhibited by microglia (red arrows) in the developing cerebellum at P17. Scale bar, 100 µm. C, Microglia with phagocytic cups (top) or microglia without phagocytic cups (bottom row) at different time points during postnatal development. Scale bars, 25 µm.

Mentions: Microglia counts were performed on P5, P7, P10, P12, P14, P17, and P21 cerebellums from intact male and female rat pups (n = 4, 2 males + 2 females for each group). Microglia were morphologically characterized based on Lenz et al. (2013) with modifications into four categories; (1) round/amoeboid microglia, (2) stout microglia, (3) microglia with thick processes (short or long), and (4) microglia with thin processes (short or long), as also described by others (Wu et al., 1992; Gómez-Gonzalez and Escobar, 2010; Schwarz et al., 2012). For descriptive purposes we identified microglia with thick and thin processes as follows: microglia with thick processes are large cells with an amorphous cell body and with at least two ramified short, long, or both thick processes. Microglia with thin processes are large or small cells with a round and small cell body, with at least four ramified short, long, or both thin processes (more ramified than microglia with thick processes). The density of total microglia, i.e., regardless of morphology, was obtained by summing all four microglial morphologies described above. In addition, cup-shaped invaginations of the plasma membrane formed around cellular debris, infectious agents or dead cells, and called “phagocytic cups” (Swanson, 2008) were also counted in the same cerebellar sections stained with Iba1. To have a clear identification of phagocytic cups, only those located at the tip of microglia processes with a round morphology were counted (Fig. 3B,C). All counts were performed solely in the cerebellar cortex; the white matter was not included. Cellular layers (granular versus molecular) were not distinguished in this experiment, as they are not fully formed at the younger ages.


Morphological and Phagocytic Profile of Microglia in the Developing Rat Cerebellum(1,2,3).

Perez-Pouchoulen M, VanRyzin JW, McCarthy MM - eNeuro (2015)

Microglial phagocytosis in the postnatal developing cerebellum. A, The highest density of phagocytic cups was observed during the third postnatal week at P17 (***p < 0.000 compared with P5, P7, P10, P12, P14, and P21; n = 4, 2 males + 2 females for each group). Data are expressed as mean ± SEM. B, Phagocytic cups exhibited by microglia (red arrows) in the developing cerebellum at P17. Scale bar, 100 µm. C, Microglia with phagocytic cups (top) or microglia without phagocytic cups (bottom row) at different time points during postnatal development. Scale bars, 25 µm.
© Copyright Policy - open-access
Related In: Results  -  Collection

License
Show All Figures
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Figure 3: Microglial phagocytosis in the postnatal developing cerebellum. A, The highest density of phagocytic cups was observed during the third postnatal week at P17 (***p < 0.000 compared with P5, P7, P10, P12, P14, and P21; n = 4, 2 males + 2 females for each group). Data are expressed as mean ± SEM. B, Phagocytic cups exhibited by microglia (red arrows) in the developing cerebellum at P17. Scale bar, 100 µm. C, Microglia with phagocytic cups (top) or microglia without phagocytic cups (bottom row) at different time points during postnatal development. Scale bars, 25 µm.
Mentions: Microglia counts were performed on P5, P7, P10, P12, P14, P17, and P21 cerebellums from intact male and female rat pups (n = 4, 2 males + 2 females for each group). Microglia were morphologically characterized based on Lenz et al. (2013) with modifications into four categories; (1) round/amoeboid microglia, (2) stout microglia, (3) microglia with thick processes (short or long), and (4) microglia with thin processes (short or long), as also described by others (Wu et al., 1992; Gómez-Gonzalez and Escobar, 2010; Schwarz et al., 2012). For descriptive purposes we identified microglia with thick and thin processes as follows: microglia with thick processes are large cells with an amorphous cell body and with at least two ramified short, long, or both thick processes. Microglia with thin processes are large or small cells with a round and small cell body, with at least four ramified short, long, or both thin processes (more ramified than microglia with thick processes). The density of total microglia, i.e., regardless of morphology, was obtained by summing all four microglial morphologies described above. In addition, cup-shaped invaginations of the plasma membrane formed around cellular debris, infectious agents or dead cells, and called “phagocytic cups” (Swanson, 2008) were also counted in the same cerebellar sections stained with Iba1. To have a clear identification of phagocytic cups, only those located at the tip of microglia processes with a round morphology were counted (Fig. 3B,C). All counts were performed solely in the cerebellar cortex; the white matter was not included. Cellular layers (granular versus molecular) were not distinguished in this experiment, as they are not fully formed at the younger ages.

Bottom Line: We found that microglial morphology changed from amoeboid to ramified during the first 3 postnatal weeks in a region specific manner.At P17 males showed an approximately twofold increase in microglia with thin processes compared with females.Our findings indicate a continuous process of microglial maturation and a nonuniform distribution of microglia in the cerebellar cortex that implicates microglia as an important cellular component of the developing cerebellum.

View Article: PubMed Central - HTML - PubMed

Affiliation: Department of Pharmacology, University of Maryland School of Medicine , Baltimore, Maryland 21201.

ABSTRACT
Microglia are being increasingly recognized as playing important roles in neurodevelopment. The cerebellum matures postnatally, undergoing major growth, but the role of microglia in the developing cerebellum is not well understood. Using the laboratory rat we quantified and morphologically categorized microglia throughout the vermis and across development using a design-based unbiased stereology method. We found that microglial morphology changed from amoeboid to ramified during the first 3 postnatal weeks in a region specific manner. These morphological changes were accompanied by the sudden appearance of phagocytic cups during the third postnatal week from P17 to P19, with an approximately fourfold increase compared with the first week, followed by a prompt decline at the end of the third week. The microglial phagocytic cups were significantly higher in the granular layer (∼69%) than in the molecular layer (ML; ∼31%) during a 3 d window, and present on ∼67% of microglia with thick processes and ∼33% of microglia with thin processes. Similar proportions of phagocytic cups associated to microglia with either thick or thin processes were found in the ML. We observed cell nuclei fragmentation and cleaved caspase-3 expression within some microglial phagocytic cups, presumably from dying granule neurons. At P17 males showed an approximately twofold increase in microglia with thin processes compared with females. Our findings indicate a continuous process of microglial maturation and a nonuniform distribution of microglia in the cerebellar cortex that implicates microglia as an important cellular component of the developing cerebellum.

No MeSH data available.


Related in: MedlinePlus