Aging results in reduced epidermal growth factor receptor signaling, diminished olfactory neurogenesis, and deficits in fine olfactory discrimination - PubMed (original) (raw)

Aging results in reduced epidermal growth factor receptor signaling, diminished olfactory neurogenesis, and deficits in fine olfactory discrimination

Emeka Enwere et al. J Neurosci. 2004.

Abstract

Previous studies demonstrating olfactory interneuron involvement in olfactory discrimination and decreased proliferation in the forebrain subventricular zone with age led us to ask whether olfactory neurogenesis and, consequently, olfactory discrimination were impaired in aged mice. Pulse labeling showed that aged mice (24 months of age) had fewer new interneurons in the olfactory bulb than did young adult (2 months of age) mice. However, the aged mice had more olfactory interneurons in total than their younger counterparts. Aged mice exhibited no differences from young adult mice in their ability to discriminate between two discrete odors but were significantly poorer at performing discriminations between similar odors (fine olfactory discrimination). Leukemia inhibitory factor receptor heterozygote mice, which have less neurogenesis and fewer olfactory interneurons than their wild-type counterparts, performed more poorly at fine olfactory discrimination than the wild types, suggesting that olfactory neurogenesis, rather than the total number of interneurons, was responsible for fine olfactory discrimination. Immunohistochemistry and Western blot analyses revealed a selective reduction in expression levels of epidermal growth factor (EGF) receptor (EGFR) signaling elements in the aged forebrain subventricular zone. Waved-1 mutant mice, which express reduced quantities of transforming growth factor-alpha, the predominant EGFR ligand in adulthood, phenocopy aged mice in olfactory neurogenesis and performance on fine olfactory discrimination tasks. These results suggest that the impairment in fine olfactory discrimination with age may result from a reduction in EGF-dependent olfactory neurogenesis.

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Figures

Figure 1.

Figure 1.

Olfactory discrimination generalization gradient, with male 2-month-old (2M) and 24-month-old (24M) C57BL/6 mice. Ratios along the abscissa represent COC:ALM concentration ratios used in [+] (top line) and in [–] (bottom line). The number of successful discriminations per trial session was expressed as a percentage of total trials per session.*p<0.05 and ***p < 0.001 versus the 2-month-old animals; Student's t test.

Figure 2.

Figure 2.

Aged mice have fewer newly generated olfactory interneurons than younger mice. A, B, In 2-month-old mice, 4 weeks after BrdU administration, a large number of BrdU-positive cells (red) was located throughout the OB, whereas a dramatic decrease in the number of BrdU-positive cells was observed in the OB of 24-month-old animals. C, D, An analysis of the interneuronal subpopulations of the OB revealed a decrease in the number of TH plus BrdU periglomerular neurons in 24-month-old versus 2-month-old mice; nevertheless, the glomerular layer was considerably larger in the 24-month-old mice than the 2-month-old mice. E–H, Decreases in the number of calretinin plus BrdU (E, F) and GABA plus BrdU (G, H) cells were also observed in the glomerular layer of aged versus young mice. I, J, This decrease in neurogenesis was not restricted to the glomerular layer, because a decrease in the number of GABA plus BrdU-expressing cells was observed in the granule cell layer of aged animals as well. For corresponding data, see Results and Table 1. Scale bars: A, C, 100 μm; (in E) E_–_J, 50 μm. Arrows indicate examples of double-labeled cells.

Figure 3.

Figure 3.

Lifr+/– mutant mice are impaired at fine olfactory discrimination. Six-month-old to 7-month-old Lifr+/+ and Lifr+/– mice were assessed for olfactory discrimination on a generalization gradient. *p < 0.05 with respect to the Lifr+/+ mice; Student's t test.

Figure 4.

Figure 4.

The number of proliferating cells in the SVZ and dorsolateral corner decreases with age. There are fewer proliferating cells (BrdU labeled) seen in 2-month-old (2M) SVZs than in those of 24-month-old (24M) SVZs. Similarly, the numbers of neuronal progenitors immunoreactive for PSA-NCAM and MASH1 in the dorsolateral corner of the SVZ, seen in 2-month-old mice, are significantly reduced in their 24-month-old counterparts. The corresponding quantitative data are in Table 2. Scale bars: (in bottom left) top left, bottom left, 100 μm; (in bottom right) middle and right, 50 μm.

Figure 5.

Figure 5.

Aging results in decreased levels of expression of the EGFR and the EGFR ligand, TGFα, within the adult SVZ. A, Immunohistochemical analysis of EGFR and CNTFRα expression in the SVZ of 2-month-old (2M) and 24-month-old (24M) mice. There are fewer cells expressing EGFR (red) in the SVZs of older mice (D, dorsal; M, medial). This becomes more apparent in a close-up of the dorsolateral corner (DLC) of the SVZ, where corresponding nuclear Hoechst staining (blue) illustrates the reduction in SVZ thickness. CNTFRα expression levels (red) were not different in the SVZs of 2-month-old and 24-month-old animals. Scale bars: SVZ panels (left), 100 μm; DLC panels (right), 50 μm. B, Western blot analysis of EGFR, TGFα, CNTFRα, and actin using protein isolated from the SVZs of 2- and 24-month-old mice. C, NIH imaging analysis (and normalization to the corresponding expression of actin) revealed that, in 24-month-old mice, EGFR expression levels were ∼50% lower than those in the younger mice (p < 0.05), TGFα levels were ∼70% lower (p < 0.05), and CNTFRα levels were unchanged. In all experiments, n = 3 animals per group.

Figure 6.

Figure 6.

Aged neural stem cells and their progeny display a decreased capacity to respond to EGF infusion and a complete but slower time course for repopulating the SVZ. A, After a 3 d infusion of EGF, the number of BrdU-expressing cells in the SVZ of 2-month-old (2M) mice increased by 82% [compared with corresponding vehicle (VEH) controls; p < 0.05], whereas BrdU cells in the SVZ of 24-month-old (24M) mice were increased by 38% (p < 0.05). B, After a 3 d AraC kill, which destroyed the entire BrdU population, 2-month-old mice repopulated after 8 d, whereas 24-month-old mice required 14 d for complete repopulation. *p < 0.0.5 compared with 2-month-old mice at the same time point. Cont., Control.

Figure 7.

Figure 7.

Waved-1 (Tgfawa1/wa1) mice exhibit NSC, OB, and behavioral characteristics similar to the 24-month-old mice. A, There were fewer NSC-derived neurospheres produced from the Tgfawa1/wa1 mouse forebrains (n = 4) than from those of the heterozygous littermates (Tgfawa1/+) (n = 4). B, The Tgfawa1/wa1 mice have more TH-positive periglomerular cells than their Tgfawa1/+ counterparts. C, Tgfawa1/wa1 mice have fewer BrdU and TH double-labeled neurons in the OB 4 weeks after BrdU administration than the Tgfawa1/+ mice. D, The Tgfawa1/wa1 mice demonstrated poorer performance at fine olfactory discrimination than the Tgfawa1/+ mice. *p < 0.05; **p < 0.01; ***p < 0.001; Student's t test.

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