The results from the present study demonstrate the involvement of GDNF in several steps of the development of cortical GABAergic cells, including their maturation and morphological differentiation, their initial motility in the subpallium, and their subsequent migration toward and through the cortex. Although a variety of roles were initially anticipated for GDNF in mammalian brain development, the analyses of mutant mice carried out previously had not uncovered any defect caused by lack of GDNF signaling in the developing brain. The requirement of GDNF signaling via GFRα1 for cortical GABAergic cell development revealed in this study represents the first function to be demonstrated for this neurotrophic factor during brain development in vivo. The fact that neither of the known GDNF signaling receptors—RET and NCAM—appeared to be involved in the effects reported here suggests the existence of novel signaling pathways, including unidentified transmembrane effectors, mediating the activities of this factor. While this paper was under review,have argued that GFRα1 expression in cells lacking RET is dispensable for CNS development, and indicated that validation of the functional importance of RET-independent signaling will require the identification of deficits that are unequivocally present in gfra1 but not in ret mutant mice. Our present paper now provides that evidence.

GDNF Signaling in the Development of Cortical GABAergic Interneurons

The activities of GDNF in the development of cortical GABAergic cells displayed a relatively broad time window, from the early stages of cell differentiation in the MGE, to subsequent tangential migration and later maturation of GABAergic neurons in the cortex and hippocampus. This is in accordance with the early expression of GDNF and GFRα1 in the MGE, along the tangential pathways of GABAergic cell migration in the developing cortical plate, and later in the cortex, during the maturation and radial dispersion of cortical GABAergic interneurons. Several of our observations suggest a role for endogenous GDNF signaling in GABAergic cell differentiation and tangential migration from the MGE: (1) MGE cells lacking GFRα1 displayed a striking immature morphology in vitro, essentially lacking neuritic processes, (2) MGE cells derived from explants lacking GDNF had reduced motility, (3) cortical explants derived from mice lacking GDNF had reduced chemoattractant activity toward MGE cells, and (4) MGE-derived calbindin- and Lhx6-expressing cells from mice lacking GFRα1 presented an aberrant pattern of tangential migration in vivo. Together with the ability of exogenous GDNF to increase GABAergic cell number and neuritogenesis in cultured MGE cells, these results indicate that endogenous GDNF signaling through GFRα1 may contribute to several fundamental aspects of the development of these cells in the MGE, including (1) the acquisition of a GABAergic phenotype by MGE neuronal precursors, (2) the ability of GABAergic cells to elaborate neuritic processes, and (3) the overall motility of these cells in situ.

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Marin O. Short- and long-range attraction of cortical GABAergic interneurons by neuregulin-1. The ability of GDNF to increase the number of GABAergic cells in MGE cultures without affecting cell proliferation or survival suggests that GDNF is a differentiation factor for MGE precursors, stimulating their acquisition of a GABAergic phenotype at the expense of other fates. This notion is supported by similar effects of GDNF on secondary MGE precursors selectively amplified through neurosphere cultures. The requirement of GFRα1 for the development of neuritic processes by MGE-derived GABAergic cells, together with the ability of GDNF to stimulate neurite outgrowth in these cells, suggests a role for endogenous GDNF signaling in the morphological differentiation of GABAergic cells in the MGE. It is well known that migrating neuronal cells utilize a leading process not only to guide cell movement but also to help propell the cell body along the migratory path (). Thus, the ability of GABAergic MGE cells to elaborate neuritic processes is likely to be instrumental for their migration. Several of our observations suggest that MGE-derived GDNF may contribute to the overall motility of GABAergic cells within this structure. Directionality in MGE cell migration could arise as a result of the combined action of MGE-derived motogenic factors, such as GDNF, and repulsive activities located in areas surrounding the MGE (). While this paper was under review,identified Neuregulin-1 as a chemoattractant factor for MGE-derived GABAergic interneurons.

The reduction in the number of calbindin-expressing cells observed in the cortex and hippocampus of mutant mice deficient in GDNF or GFRα1 could be accounted for by deficits in both the differentiation and migration of GABAergic cells. The fact that lower numbers of cells expressing Gad and Lhx6 mRNA were also observed in the cortex of mice lacking GFRα1 suggests an overall decrease in the number of inhibitory interneurons and not simply a mere reduction in the expression of calbindin in those cells. Although the physiological consequences of a reduction in the number of inhibitory interneurons on adult brain function could not be assessed in this study, our results warrant future investigations on the possible roles of GDNF in the assembly and function of inhibitory circuits using tissue-specific mutants.