In utero fate mapping reveals distinct migratory pathways and fates of neurons born in the mammalian basal forebrain
Hynek Wichterle1,*,
Daniel H. Turnbull2,
Susana Nery2,
Gord Fishell2 and
Arturo Alvarez-Buylla1,3,
1 The Rockefeller University, New York, NY 10021, USA
2 The Skirball Institute of Biomolecular Medicine, New York University School of Medicine, New York, NY 10016, USA
3 University of California, San Francisco, Department of Neurosurgery and Anatomy, San Francisco, CA 94143, USA
* Present address: Hynek Wichterle, Howard Hughes Medical Institute, Center for Neurobiology and Behavior, Columbia University, 1013 Hammer Health Sciences Center, 701 West 168th St, New York, NY 10032, USA
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Fig. 1. Distribution of MGE cells one (E14.5, A-C), two (E15.5, D-F) and four (E17.5, G-I) days after homotopic transplantation. Coronal sections of embryonic mouse brains containing MGE cells labeled with PKH26 fluorescent dye before transplantation. Graft-derived cells are black or dark blue. (A) One day after transplantation, most of the grafted cells were located at the site of transplantation in the MGE. No grafted cells were detected in the lateral ventricle (lv). The pipette insertion site is marked by an arrow. (B) At 1 day post transplantation migrating cells were observed in the LGE and at the cortical boundary (arrowhead). (C) Example of a migratory cell with a leading process and a growth cone. (D) Two days after transplantation, multiple cells migrated into the neocortex. Migration is directional as no cells migrate to the septum (sep). (E) The highest density of migrating MGE cells is in the cortical subventricular zone. (F) Tangential section through the anterior neocortex reveals migrating cells in the subventricular zone (central lighter region). No graft-derived cells migrated into the olfactory bulb (ob). (G) Four days after transplantation, most of the MGE-derived cells settled in the neocortical marginal zone. No labeled cells were found in the hypothalamus (ht) and thalamus (th). (H) Cortical subventricular zone contains fewer migrating cells than at 2 days and most of the grafted cells are found in the marginal zone. (I) Disappearance of migrating neurons from the subventricular zone is obvious in the tangentially cut anterior cortex. Olfactory bulb is devoid of labeled cells.
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Fig. 2. MGE cells in the subventricular and marginal zones at two (E15.5) and four (E17.5) days after transplantation. (A) Higher magnification of the boundary between the developing striatum and neocortex at 2 days after transplantation reveals the shape and orientation of tangentially migrating neurons. Most PKH26-labeled MGE neurons migrate close to the neocortical ventricular zone and only few cells are observed in the marginal zone. (B) Double labeling for BrdU reveals most of the tangentially oriented cells in the neocortical SVZ. (C) Tangentially migrating cells do not use corticofugal axons in the upper intermediate zone, which were labeled with antibody against neurofilament-145. (D) Confocal image of differentiating MGE-derived neurons in the marginal zone 4 days after transplantation. (E) Quantification of the density of graft-derived cells demonstrates the shift from the subventricular (SVZ) to marginal zone (MZ) between days 2 and 4. (F) Immunostaining of transplanted MGE cells expressing alkaline phosphatase in one day old pup (7 days after transplantation) reveals that some of the MGE-derived cells (green) express GABA (red). (G) Staining with antibody CR-50 against reelin (red) revealed that MGE-derived cells (green) are found close to clusters of CR-50 positive cells but they are not double-labeled.
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Fig. 3. MGE cells expressing alkaline phosphatase were grafted into the wild-type E13.5 MGE. The majority of graft-derived cells are found in the neocortex in 3-week-old (left, more posterior coronal section) and 4-month-old (right, more anterior coronal section) animals. Some cells in the posterior section are observed in the amygdala and CA1 field of the hippocampus. In contrast to the 4-day survival (Fig. 1G-I), no graft-derived cells are found in the marginal zone (the dark staining in the marginal zone are labeled projections of neurons located deeper in the cortical plate, arrow). A, amygdala; MZ, marginal zone; NCx, neocortex; Str, striatum.
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Fig. 4. Morphology of MGE-derived neurons stained for alkaline phosphatase. (A,D,E,H) Neurons in 4-month-old animal; (B,C,F,G,I) neurons in 3-week-old animal; dorsal is towards the top throughout. MGE cells in the neocortex differentiated into: aspiny multipolar neurons (A,C), bi-tufted neurons (B), neurons with a small cell body (D, arrow), basket cells (E) and chandelier cells (F). MGE cells were also observed in: the CA1 field of the hippocampus (basket cells) (G), the lateral globus pallidus (H) and the striatum (I). To capture some of the fine three dimensional processes, two to three photographs taken in different focal planes were fused to produce the final image.
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Fig. 5. Immunohistochemical characterization of grafted MGE (A-C) and LGE (D-F) cells in the neocortex (NCx; A,B,F) and striatum (Str; C,D,E). MGE-derived cells are from a 4-month-old animal, LGE-derived cells are from 3-week-old animal. The second column and green color in the third column represent staining for alkaline phosphatase expressed in transplanted cells. (A-C) Many MGE cell in the neocortex expressed parvalbumin (A) and somatostatin (B) but within striatum did not express striatal marker DARPP-32 (C). (D,E) The majority of LGE cells in the striatum expressed DARPP-32 (D) but did not express parvalbumin (E). (F) By contrast, some of the LGE-derived neurons that migrated into the neocortex expressed parvalbumin.
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Fig. 6. Homotopic transplants of PKH26-labeled LGE cells. (A) Coronal section of E17.5 embryonic brain four days after transplantation reveals accumulation of LGE cells in the striatum and absence of LGE cells in the neocortex. Some labeled cells leaked into brain ventricles during the transplantation (black dots in choroid plexus of the lateral ventricle). (B) More anterior view reveals selective dispersion of LGE-derived cells throughout the striatum (str). Notice the sharp boundary between the striatum and neocortex (ncx). (C) LGE-derived cells migrate also into the nucleus accumbens (na) and olfactory tubercle (ot) in the basal forebrain. (D) High magnification of labeled cells in the olfactory tubercle reveals multiple processes extending from individual cells. (E) In several animals, labeled LGE cells migrated into the core of the olfactory bulb (ob).
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Fig. 7. Alkaline phosphatase-expressing LGE cells in 3-week-old animals. (B,D,E) Two to three fused photographs taken in different focal planes. (A) LGE cells disperse throughout the striatum and migrate ventrally into the olfactory tubercle (ot). Many labeled axons crisscross the ventral pallidum (vp). Notice that this distribution is complementary to the distribution of MGE cells (Fig. 3). No cells are detected in the septum (sep) and only rarely in the neocortex. (B) LGE cells in the striatum differentiated into medium spiny neurons. (C) Projecting axons were found among others in the medial globus pallidus. (D) The few LGE-derived neurons that migrated into the neocortex differentiated into aspiny neurons. LGE cells migrated to the olfactory bulb and differentiated into interneurons in the granule cell layer (gcl) and glomerular layer (gl). (E) Immature neurons were observed in the rostral migratory stream (rms). (F) LGE cells in the olfactory tubercle differentiated into bi-tufted spiny neurons with processes reaching the surface of the brain.
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Fig. 8. Model of tangential neuronal migratory pathways in the developing mouse brain. Longitudinal view of the developing telencephalic vesicle with the anterior end pointing to the right. Gray arrows depict the large scale migration of MGE cells into the developing neocortex (ncx). Black arrows depict the migratory pathways from different parts of the LGE. Cells from the dorsoanterior LGE migrate preferentially into the olfactory bulb (ob), while more posteriorly grafted LGE cells migrate preferentially into the nucleus accumbens (na). LGE cells disperse through the striatum and migrate ventrally to the olfactory tubercle (ot). Hypothetical migratory routes from the caudal ganglionic eminence (CGE) are depicted by broken arrows.
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