Basic fibroblast growth factor (Fgf2) is necessary for cell proliferation and neurogenesis in the developing cerebral cortex - PubMed (original) (raw)

Basic fibroblast growth factor (Fgf2) is necessary for cell proliferation and neurogenesis in the developing cerebral cortex

R Raballo et al. J Neurosci. 2000.

Abstract

Little is known about regionally specific signals that control the number of neuronal progenitor cells in vivo. We have previously shown that the germline mutation of the basic fibroblast growth factor (Fgf2) gene results in a reduction in the number of cortical neurons in the adult. We show here that Fgf2 is expressed in the pseudostratified ventricular epithelium (PVE) in a dorsoventral gradient and that Fgf2 and its receptor, Fgfr-1, are downregulated by mid to late stages of neurogenesis. In Fgf2 knockout mice, the volume and cell number of the dorsal PVE (the cerebral cortical anlage) are substantially smaller, whereas the volume of the basal PVE is unchanged. The dorsal PVE of Fgf2 knockout mice has a 50% decrease in founder cells and a reduced expansion of the progenitor pool over the first portion of neurogenesis. Despite this reduction, the degree of apoptosis within the PVE is not changed in the Fgf2 knockouts. Cortical neuron number was decreased by 45% in Fgf2 knockout mice by the end of neurogenesis, whereas the number of neurons in the basal ganglia was unaffected. Microscopically, the frontal cerebral cortex of neonatal Fgf2 null mutant mice lacked large neurons in deep cortical layers. We suggest that Fgf2 is required for the generation of a specific class of cortical neurons arising from the dorsal PVE.

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Figures

Fig. 1.

Fgf2 and Fgfr-1 expression in the developing rat telencephalon. Immunocytochemistry for Fgf2 (a–c) and Fgfr-1 (d–g, i) in parasagittal frozen sections is shown; the embryonic age is indicated above each panel. The_inset_ in a shows a higher magnification of the dorsal PVE (dPVE). g shows that no reaction is present after preincubation of the Fgfr-1 antibody with an excess of Fgfr-1 peptide. h shows BrdU immunostaining (green) after BrdU incorporation in vivo for 3.5 hr, and i shows the same section double-immunostained with Fgfr-1. Arrows indicate proliferative cells that contain Fgfr-1 immunoreactivity. In_a_, d, and g, anterior is_left_, and in all sections the ventricular surface is at the bottom. lge, Lateral ganglionic eminence; mge, medial ganglionic eminence;dPVE, dorsal PVE. Scale bars: a, 400 μm; d, g, 200 μm; b,c, e, f, 20 μm;h, i, 50 μm.

Fig. 2.

Decrease in PVE size in Fgf2−/− mice. Estimation of dorsal and basal PVE volumes for Fgf2−/− mutant and wild-type littermate mice. Embryos were harvested at the indicated stages after the entire proliferative population was labeled with cumulative BrdU injections. Volumes were determined by the Cavalieri method using serial sections immunostained for BrdU and counterstained with cresyl violet.

Fig. 3.

The rate of increase in cell number within the PVE is lower in Fgf2 knockout embryos. Shown is a plot of cell number as a function of age in the dorsal portion of the PVE during the initial stages of neurogenesis. Cell number was assessed by stereological analyses in Fgf2 null mutant (Fgf2 − /−) and wild-type (wt) littermates. Curve fitting yielded the equations y_= 317 + 888 · x for wild type and_y = 78 + 630 · x for Fgf2−/−;_R_2 was 0.98 and 0.95, respectively. Cell number was significantly different between genotypes by ANOVA (F = 6.7; p < 0.01).

Fig. 4.

Cerebral cortical neurons in newborn FGF2−/− mice. Shown is NeuN immunostaining in the frontal cortex of wild-type (a) and FGF2−/− mice (b) at P1. The bar at left indicates the boundary between supragranular and infragranular layers. Note the prominent nuclear staining of large, pyramidal-shaped cells in layer 5/6 (arrows) in wild-type mice that is absent in FGF2−/− mice. No difference in neuronal number is apparent in upper layers. Pial side is at the top. Scale bar, 100 μm.

Fig. 5.

Apoptosis during forebrain development in wild-type and Fgf2−/− mice. Shown is TUNEL assay in parasagittal sections from wild-type (a, c,e, f) and Fgf2−/− (b, d) mice counterstained with methyl green. Apoptotic cells (brown) are indicated by_arrowheads_, and arrows point to migrating neurons in the intermediate zone. Age is indicated above the panels.e is a positive control treated with DNase, and_f_ shows the nasal cavity with apoptotic cells at the tips of the nasal turbinates and in the endothelium of the tongue. The ventricular surface is at the bottom.pve, Pseudostratified ventricular epithelium;iz, intermediate zone; cp, cortical plate; mz, marginal zone; e, epidermis. Scale bars:, a_–_e, 100 μm;f, 200 μm.

Fig. 6.

Model of the possible relationships between Fgf2 availability and cell cycle events. A, Salt-and-pepper appearance of Fgf2 mRNA in the early PVE (Vaccarino et al., 1999a), suggesting that cell-to-cell variability in internal sources of Fgf2 may represent a primary factor governing Fgf2 exposure within these cells and their neighbors. B, At later stages of development, Fgf2 is downregulated within the PVE except for the cells close to the apical surface. The interkinetic nuclear movement coupled to the cell cycle is shown in relation to the hypothesized action of Fgf2. In a subset of progenitor cells, Fgf2 is necessary in early G1 to promote commitment to a subsequent cell cycle (DeHamer et al., 1994). During early G1, cells are also close to the ventricular surface, which could be a source of Fgf2 protein. Fgf2 may be released into the CSF by PVE cells or by cells of the choroid plexus, which contain high amounts of Fgf2. The increased nuclear shading represents the association of Fgf ligands with the nucleus, also occurring during the G1 phase (Zhan et al., 1993; Prudovsky et al., 1994).

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Basic fibroblast growth factor (Fgf2) is necessary for cell proliferation and neurogenesis in the developing cerebral cortex - PubMed (original) (raw)