TrkB signaling is required for postnatal survival of CNS neurons and protects hippocampal and motor neurons from axotomy-induced cell death - PubMed (original) (raw)

TrkB signaling is required for postnatal survival of CNS neurons and protects hippocampal and motor neurons from axotomy-induced cell death

S Alcántara et al. J Neurosci. 1997.

Abstract

Newborn mice carrying targeted mutations in genes encoding neurotrophins or their signaling Trk receptors display severe neuronal deficits in the peripheral nervous system but not in the CNS. In this study, we show that trkB (-/-) mice have a significant increase in apoptotic cell death in different regions of the brain during early postnatal life. The most affected region in the brain is the dentate gyrus of the hippocampus, although elevated levels of pyknotic nuclei were also detected in cortical layers II and III and V and VI, the striatum, and the thalamus. Furthermore, axotomized hippocampal and motor neurons of trkB (-/-) mice have significantly lower survival rates than those of wild-type littermates. These results suggest that neurotrophin signaling through TrkB receptors plays a role in the survival of CNS neurons during postnatal development. Moreover, they indicate that TrkB receptor signaling protects subpopulations of CNS neurons from injury- and axotomy-induced cell death.

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Figures

Fig. 1.

Fig. 1.

Cytoarchitecture of brain structures of postnatal_trk_B (−/−) mutant mice. A–D, Low-power micrographs of the brain of wild-type (A, C) and_trk_B (−/−) (B, D) mice. Notice the normal appearance of Nissl-stained brain sections from P13 (B) and P16 (D) _trk_B (−/−) mice compared with their age-mated control littermates (A, C, respectively). E, F, High-power magnification of the neocortex of P16 wild-type (E) and_trk_B (−/−) (F) mice. Notice the presence of empty areas devoid of neurons in _trk_B (−/−) mice (F) (from D, inset).G, H, Calretinin immunoreactivity in the neocortex of P10 wild-type (G) and trk_B (−/−) (H) mice. Cortical layers are denoted by_roman numerals. NC, Neocortex;H, hippocampus; T, thalamus. Magnification: A–D, 2.5×; E, F, 40×;G, H, 10×.

Fig. 2.

Fig. 2.

Increased cell death in _trk_B (−/−) mice. A–D, Cresyl violet-stained sections of the neocortex (A, B) and dentate gyrus (C, D) of P16 control (A, C) and_trk_B (−/−) (B, D) mice. Notice the presence of pyknotic nuclei in layers II and III of the neocortex (B, arrows) and the granular layer of the dentate gyrus (D) in the _trk_B (−/−) mutant mice.GL, Granular layer; H, hilus;ML, molecular layer. Magnification, 40×.

Fig. 3.

Fig. 3.

Neurons in _trk_B (−/−) mice die by apoptosis. A, B, Immunolabeling with c-Jun antibody reveals the presence of pyknotic cells in layers II and III (A) and degenerating neurons in the pyriform cortex (B) in P10 _trk_B (−/−) mice.C, Electron micrograph showing the characteristic morphology of an apoptotic cell in the dentate gyrus that is being engulfed by a neighboring cell in a _trk_B (−/−) mouse.D–G, TUNEL staining of neocortex (layers II and III) (D, E) and hippocampus (F, G); sections from P13 control (D, F) and _trk_B (−/−) (E, G) mice. Notice the increase in TUNEL-positive nuclei in the trk_B (−/−) mice.Arrows in A denote pyknotic nuclei. The_arrowhead denotes a pyknotic nucleus negative for c-Jun staining. Sections in A and B are counterstained with cresyl violet. GL, Granular layer;H, hilus; ML, molecular layer. Magnification: A, B, 30×; C, 15,000×;D–G, 40×.

Fig. 4.

Fig. 4.

Expression of calcium-binding proteins in degenerating neurons in _trk_B (−/−) mutant mice.A–D, Calretinin immunoreactivity in layers II and III of the neocortex (A, B, D) and the stratum oriens of the hippocampus (C) of P10 _trk_B (−/−) mutant mice. Presumably, degenerating neurons with atrophic somata and beaded dendrites are immunoreactive for calretinin staining (A, B, arrows). E, Calbindin expression in layers II and III of the neocortex of a P13 _trk_B (−/−) mice. Arrows in C–E denote pyknotic cells immunoreactive for different calcium-binding proteins.Arrowheads denote unlabeled pyknotic cells. Sections are counterstained with cresyl violet. Magnification: _A, B,_35×; C, D, 40×; E, 45×.

Fig. 5.

Fig. 5.

Number of pyknotic nuclei in different regions of the brain at several developmental stages (expressed as number per 100,000 μm2). A, P5–P8 mice (control mice, n = 4; _trk_B (−/−) mice,n = 4). B, P10–P12 mice (control mice, n = 6; _trk_B (−/−) mice,n = 7). C, P13–P18 mice (control mice, n = 10; _trk_B (−/−) mice,n = 11). Black bars, Control mice;_white bars, trk_B (−/−) mutant mice. _DG,_Dentate gyrus; CA3, CA3 hippocampal field;II–III and V–VI, cortical layers II and III and V and VI, respectively; VPM, ventroposterior medialis thalamic nucleus; RET, reticular thalamic nucleus; STR, striatum. Error bars indicate SEM;asterisks indicate that there are significant differences between control and _trk_B (−/−) mutant mice (*p < 0.01; **p < 0.001).

Fig. 6.

Fig. 6.

Increased cell death in hippocampal slice cultures from _trk_B (−/−) mice. A, B, Cresyl violet-stained sections show the normal structure of hippocampal slices in both control (A, C) and _trk_B (−/−) mice (B, D) after 3 d in culture. C, D, Higher-magnification micrographs show an increase in pyknotic nuclei in the CA3 region of _trk_B (−/−) mutant mice (D) when compared with control mice (C). CA1, Pyramidal layer CA1;CA3, pyramidal layer CA3; DG, dentate gyrus; SB, subiculum. Magnification: A, B, 10×; C, D, 40×.

Fig. 7.

Fig. 7.

Number of pyknotic nuclei in different regions of the hippocampus after 3 (A) and 5 (B) d in culture (expressed as number per 25,000 μm2).Black bars, Control mice (n = 8 in each group of cultures); white bars, trk_B (−/−) mutant mice (n = 8 in each group of cultures).DG, Dentate gyrus; CA1 and_CA3, CA1 and CA3 hippocampal fields, respectively;SB, subiculum. Error bars indicate SEM;asterisks indicate that there are significant differences between control and _trk_B (−/−) mutant mice (*p < 0.001; **p < 0.0001).

Fig. 8.

Fig. 8.

Number of pyknotic nuclei in different regions of the hippocampus after 3 d of treatment with neurotrophins in culture (expressed as number per 25,000 μm2).Black bars, Vehicle; dotted bars, BDNF;gray bars, NT-3; white bars, NT-4;striped bars, NGF (n = 8 or 9 cultures per group). DG, Dentate gyrus;CA1 and CA3, CA1 and CA3 hippocampal fields, respectively; SB, subiculum. Error bars indicate SEM; asterisks indicate that there are significant differences between the different treatments (*p < 0.01, ANOVA with Fisher’s protected least significant difference for_post hoc_ comparisons).

Fig. 9.

Fig. 9.

Decrease in the number of facial motor neurons after axotomy in _trk_B (−/−) mice. _A–F,_Cresyl violet-stained sections show the reduction in the size of the axotomized facial motor nucleus (B) but the normal appearance of the surviving motor neurons (E) in P10_trk_B (−/−) mice. A, C, D, Wild-type mice. _B, E, F, trk_B (−/−) mice. _G, H,_Acetylcholinesterase activity in P7 wild-type (G) and_trk_B (−/−) (H) mice. Similar activity was observed in the _trk_B (−/−) mutant mice when compared with wild-type mice. Magnification: A, B, G, H, 2.5×; C–F, 100×.

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