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Expression of TrkC receptors in the developing brain of the Monodelphis opossum and its effect on the development of cortical cells.

Bartkowska K, Gajerska M, Turlejski K, Djavadian RL - PLoS ONE (2013)

Bottom Line: We found that in different neocortical areas located both at the rostral and caudal regions of the cortex, up to 98% of BrdU-labeled cells forming cortical layers (II-VI) had prominently expressed TrkC.The shRNAs were individually tested in transfected cortical progenitor cells grown on culture plates for 2 days.The effects of the shRNA-TrkC constructs were similar: blockade of TrkC receptors decreased the number of Ki67-positive and apoptotic cells, and it did not change the number of TUJ-positive neurons in vitro.

View Article: PubMed Central - PubMed

Affiliation: Department of Molecular and Cellular Neurobiology, Nencki Institute of Experimental Biology, Warsaw, Poland.

ABSTRACT
In this study, we investigated the distribution, localization and several various functions of TrkC receptors during development of the Monodelphisopossum brain. Western blotting analysis showed that two different forms of the TrkC receptor, the full-length receptor and one of its truncated forms, are abundantly expressed in the opossum brain. The expression of TrkC receptors was barely detected in the brain of newborn opossums. At postnatal day (P) 3, the expression of full-length TrkC remained at low levels, while moderate expression of the TrkC truncated form was detected. The expression levels of both forms of this protein gradually increased throughout development, peaking at P35. We found that in different neocortical areas located both at the rostral and caudal regions of the cortex, up to 98% of BrdU-labeled cells forming cortical layers (II-VI) had prominently expressed TrkC. To assess which developmental processes of cortical cells are regulated by TrkC receptors, three different shRNAs were constructed. The shRNAs were individually tested in transfected cortical progenitor cells grown on culture plates for 2 days. The effects of the shRNA-TrkC constructs were similar: blockade of TrkC receptors decreased the number of Ki67-positive and apoptotic cells, and it did not change the number of TUJ-positive neurons in vitro. Thus, the lack of TrkC receptors in cultured progenitor cells provided insight on the potential role of these receptors in the regulation of proliferation and cell survival but not in the differentiation of cortical cells.

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Expression of TrkC in newly generated BrdU-labeled cells, which form layers in the cerebral cortex.A-F - Location of the BrdU-labeled cell nuclei on coronal brain sections in 3-month-old opossums injected with BrdU on days P3 (A, D), P7 (B, E) and P12 (C, F). Two sections per brain are illustrated. A-C – Sections were obtained from the rostral part of the opossum brains. D-F - Sections were obtained from the caudal part of the opossum brain. Animals were injected with BrdU at 3 (D), 7 (E) and 12 (F) days after birth. G-L – confocal images of double staining for BrdU and TrkC coexpression in the same neurons. A coronal section from the caudal part of the opossum brain that was injected 12 days after birth and perfused at 3 months.
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pone-0074346-g004: Expression of TrkC in newly generated BrdU-labeled cells, which form layers in the cerebral cortex.A-F - Location of the BrdU-labeled cell nuclei on coronal brain sections in 3-month-old opossums injected with BrdU on days P3 (A, D), P7 (B, E) and P12 (C, F). Two sections per brain are illustrated. A-C – Sections were obtained from the rostral part of the opossum brains. D-F - Sections were obtained from the caudal part of the opossum brain. Animals were injected with BrdU at 3 (D), 7 (E) and 12 (F) days after birth. G-L – confocal images of double staining for BrdU and TrkC coexpression in the same neurons. A coronal section from the caudal part of the opossum brain that was injected 12 days after birth and perfused at 3 months.

Mentions: To examine the birthdates of cells in different cortical layers, the opossums were injected with BrdU at various developmental time points, and then perfused at the age of 3 months. The long survival period after BrdU injections determine the final location of the labeled cells in the brain at specific developmental time points. We found that at P3, a clear rostroventral-to-caudodorsal gradient of BrdU-immunoreactivity was present in the cortical layers (Figure 4 A, D). At P3, many BrdU-labeled nuclei were found in the infragranular layers VI and V (Figure 4 A) and in layer IV of the rostroventral region of the neocortex, where the frontal, cingulate and somatosensory areas are located. However, the immunoreactive BrdU cells were restricted only to layer VI and partially to layer V of the caudodorsal neocortex, which largely corresponded to visual area 17 (Figure 4 D). The emergence of layers IV and III of the rostroventral cortex (Figure 4 B) and layers V and IV of the caudodorsal cortex occurred at P7 (Figure 4 E). The last cortical layer of the rostrodorsal cortex was established at P12 (Figure 4 F), while generation of cortical neurons in the caudodorsal part of the cortex ended at P17 (data not shown). In addition, the vast majority of BrdU-labeled cells expressed TrkC and NeuN (Figure 4 G–L).


Expression of TrkC receptors in the developing brain of the Monodelphis opossum and its effect on the development of cortical cells.

Bartkowska K, Gajerska M, Turlejski K, Djavadian RL - PLoS ONE (2013)

Expression of TrkC in newly generated BrdU-labeled cells, which form layers in the cerebral cortex.A-F - Location of the BrdU-labeled cell nuclei on coronal brain sections in 3-month-old opossums injected with BrdU on days P3 (A, D), P7 (B, E) and P12 (C, F). Two sections per brain are illustrated. A-C – Sections were obtained from the rostral part of the opossum brains. D-F - Sections were obtained from the caudal part of the opossum brain. Animals were injected with BrdU at 3 (D), 7 (E) and 12 (F) days after birth. G-L – confocal images of double staining for BrdU and TrkC coexpression in the same neurons. A coronal section from the caudal part of the opossum brain that was injected 12 days after birth and perfused at 3 months.
© Copyright Policy
Related In: Results  -  Collection

Show All Figures
getmorefigures.php?uid=PMC3760877&req=5

pone-0074346-g004: Expression of TrkC in newly generated BrdU-labeled cells, which form layers in the cerebral cortex.A-F - Location of the BrdU-labeled cell nuclei on coronal brain sections in 3-month-old opossums injected with BrdU on days P3 (A, D), P7 (B, E) and P12 (C, F). Two sections per brain are illustrated. A-C – Sections were obtained from the rostral part of the opossum brains. D-F - Sections were obtained from the caudal part of the opossum brain. Animals were injected with BrdU at 3 (D), 7 (E) and 12 (F) days after birth. G-L – confocal images of double staining for BrdU and TrkC coexpression in the same neurons. A coronal section from the caudal part of the opossum brain that was injected 12 days after birth and perfused at 3 months.
Mentions: To examine the birthdates of cells in different cortical layers, the opossums were injected with BrdU at various developmental time points, and then perfused at the age of 3 months. The long survival period after BrdU injections determine the final location of the labeled cells in the brain at specific developmental time points. We found that at P3, a clear rostroventral-to-caudodorsal gradient of BrdU-immunoreactivity was present in the cortical layers (Figure 4 A, D). At P3, many BrdU-labeled nuclei were found in the infragranular layers VI and V (Figure 4 A) and in layer IV of the rostroventral region of the neocortex, where the frontal, cingulate and somatosensory areas are located. However, the immunoreactive BrdU cells were restricted only to layer VI and partially to layer V of the caudodorsal neocortex, which largely corresponded to visual area 17 (Figure 4 D). The emergence of layers IV and III of the rostroventral cortex (Figure 4 B) and layers V and IV of the caudodorsal cortex occurred at P7 (Figure 4 E). The last cortical layer of the rostrodorsal cortex was established at P12 (Figure 4 F), while generation of cortical neurons in the caudodorsal part of the cortex ended at P17 (data not shown). In addition, the vast majority of BrdU-labeled cells expressed TrkC and NeuN (Figure 4 G–L).

Bottom Line: We found that in different neocortical areas located both at the rostral and caudal regions of the cortex, up to 98% of BrdU-labeled cells forming cortical layers (II-VI) had prominently expressed TrkC.The shRNAs were individually tested in transfected cortical progenitor cells grown on culture plates for 2 days.The effects of the shRNA-TrkC constructs were similar: blockade of TrkC receptors decreased the number of Ki67-positive and apoptotic cells, and it did not change the number of TUJ-positive neurons in vitro.

View Article: PubMed Central - PubMed

Affiliation: Department of Molecular and Cellular Neurobiology, Nencki Institute of Experimental Biology, Warsaw, Poland.

ABSTRACT
In this study, we investigated the distribution, localization and several various functions of TrkC receptors during development of the Monodelphisopossum brain. Western blotting analysis showed that two different forms of the TrkC receptor, the full-length receptor and one of its truncated forms, are abundantly expressed in the opossum brain. The expression of TrkC receptors was barely detected in the brain of newborn opossums. At postnatal day (P) 3, the expression of full-length TrkC remained at low levels, while moderate expression of the TrkC truncated form was detected. The expression levels of both forms of this protein gradually increased throughout development, peaking at P35. We found that in different neocortical areas located both at the rostral and caudal regions of the cortex, up to 98% of BrdU-labeled cells forming cortical layers (II-VI) had prominently expressed TrkC. To assess which developmental processes of cortical cells are regulated by TrkC receptors, three different shRNAs were constructed. The shRNAs were individually tested in transfected cortical progenitor cells grown on culture plates for 2 days. The effects of the shRNA-TrkC constructs were similar: blockade of TrkC receptors decreased the number of Ki67-positive and apoptotic cells, and it did not change the number of TUJ-positive neurons in vitro. Thus, the lack of TrkC receptors in cultured progenitor cells provided insight on the potential role of these receptors in the regulation of proliferation and cell survival but not in the differentiation of cortical cells.

Show MeSH
Related in: MedlinePlus