During the development of the mammalian brain, most of the interneurons that will become part of cortical networks are born from progenitors in the subpallial medial ganglionic eminence (MGE). The resulting post-mitotic interneurons (IN) reach the developing cortex through a long tangential migration. We have previously shown that Nrg1, sensed trough ErbB4 receptor, is a potent cortical chemoattractant for migrating IN. Here, we have investigated the cellular and molecular mechanisms underlying the chemoattraction exerted by Nrg1 over migrating interneurons. Objectives: i) To describe the intracellular signaling pathways elicited by Nrg1 on MGE-derived interneurons, ii) To analyze the specific role of ErbB4 as the main Nrg1 molecular sensor, iii) To determine the cellular correlates of Nrg1/ErbB4 activation in migrating neurons. Methods: We prepared MGE-derived primary cultures from ICR or ErbB4 mutant mice, and challenged them with purified Nrg1. The phosphorylation status of AKT, ERK and other signalling cascades was analyzed by WB, and cross-linked experiments were carried out to investigate the formation of ErbB dimers. WB and RT-PCR analyses were also done to determine the pool of ErbB receptors expressed by interneurons. Cellular analyses were carried out with MGE explants or organotypic slices, and chemical inhibitors of the relevant signalling pathways added to the medium. Finally, slice electroporation experiments were performed to investigate the role of different ErbB4 isoforms transducing Nrg1 chemoattraction. Results: Several lines of evidence suggest that migrating interneurons primarily respond to Nrg1 through ErbB4 homodimer receptors. Nrg1 stimulates both AKT and ERK1/2 pathways, as shown by WB and pharmacological experiments. ErbB4 mutant explants displayed impaired Nrg1-mediated responses. Finally, we found that a special combination of ErbB4 isoforms –and hence a specific plethora of signals - is needed for the chemoattraction of the IN towards Nrg1.
Dissecting the molecular mechanisms involved in the ErbB4/Nrg1- directed chemoattraction of cortical interneurons
GAMBAROTTA, Giovanna;
2009-01-01
Abstract
During the development of the mammalian brain, most of the interneurons that will become part of cortical networks are born from progenitors in the subpallial medial ganglionic eminence (MGE). The resulting post-mitotic interneurons (IN) reach the developing cortex through a long tangential migration. We have previously shown that Nrg1, sensed trough ErbB4 receptor, is a potent cortical chemoattractant for migrating IN. Here, we have investigated the cellular and molecular mechanisms underlying the chemoattraction exerted by Nrg1 over migrating interneurons. Objectives: i) To describe the intracellular signaling pathways elicited by Nrg1 on MGE-derived interneurons, ii) To analyze the specific role of ErbB4 as the main Nrg1 molecular sensor, iii) To determine the cellular correlates of Nrg1/ErbB4 activation in migrating neurons. Methods: We prepared MGE-derived primary cultures from ICR or ErbB4 mutant mice, and challenged them with purified Nrg1. The phosphorylation status of AKT, ERK and other signalling cascades was analyzed by WB, and cross-linked experiments were carried out to investigate the formation of ErbB dimers. WB and RT-PCR analyses were also done to determine the pool of ErbB receptors expressed by interneurons. Cellular analyses were carried out with MGE explants or organotypic slices, and chemical inhibitors of the relevant signalling pathways added to the medium. Finally, slice electroporation experiments were performed to investigate the role of different ErbB4 isoforms transducing Nrg1 chemoattraction. Results: Several lines of evidence suggest that migrating interneurons primarily respond to Nrg1 through ErbB4 homodimer receptors. Nrg1 stimulates both AKT and ERK1/2 pathways, as shown by WB and pharmacological experiments. ErbB4 mutant explants displayed impaired Nrg1-mediated responses. Finally, we found that a special combination of ErbB4 isoforms –and hence a specific plethora of signals - is needed for the chemoattraction of the IN towards Nrg1.
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