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. 2018 May;17(5):6465-6471.
doi: 10.3892/mmr.2018.8705. Epub 2018 Mar 8.

Age-dependent Changes in Vesicular Glutamate Transporter 1 and 2 Expression in the Gerbil Hippocampus

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Free PMC article

Age-dependent Changes in Vesicular Glutamate Transporter 1 and 2 Expression in the Gerbil Hippocampus

Hyo Young Jung et al. Mol Med Rep. .
Free PMC article

Abstract

Glutamate is a major excitatory neurotransmitter that is stored in vesicles located in the presynaptic terminal. Glutamate is transported into vesicles via the vesicular glutamate transporter (VGLUT). In the present study, the age‑associated changes of the major VGLUTs, VGLUT1 and VGLUT2, in the hippocampus were investigated, based on immunohistochemistry and western blot analysis at postnatal month 1 (PM1; adolescent), PM6, PM12 (adult group), PM18 and PM24 (the aged groups). VGLUT1 immunoreactivity was primarily detected in the mossy fibers, Schaffer collaterals and stratum lacunosum‑moleculare. By contrast, VGLUT2 immunoreactivity was observed in the granule cell layer and the outer molecular layer of the dentate gyrus, stratum pyramidale, Schaffer collaterals and stratum lacunosum‑moleculare in the hippocampal CA1‑3 regions. VGLUT1 immunoreactivity and protein levels remained constant across all age groups. However, VGLUT2 immunoreactivity and protein levels decreased in the PM3 group when compared with the PM1 group. VGLUT2 immunoreactivity and protein levels were not altered in the PM12 group; however, they increased in the PM18 group. In addition, in the PM18 group, highly immunoreactive VGLUT2 cells were also identified in the stratum radiatum and oriens of the hippocampal CA1 region. In the PM24 group, VGLUT2 immunoreactivity and protein levels were significantly decreased and were the lowest levels observed amongst the different groups. These results suggested that VGLUT1 may be less susceptible to the aging process; however, the increase of VGLUT2 in the non‑pyramidal cells in the PM18 group, and the consequent decrease in VGLUT2, may be closely linked to age‑associated memory impairment in the hippocampus.

Figures

Figure 1.
Figure 1.
Immunohistochemistry for (A) VGLUT1 and (B) VGLUT2 in the cerebellum and with pre-immune serum in (C) cerebellum and (D) hippocampus of adult PM6 gerbils. VGLUT1 and VGLUT2 immunoreactivity is found in the molecular and granular layer of cerebellum. Immunostaining with pre-immune serum does not show any immunoreactivities in cerebellum and hippocampus. Scale bar=100 µm.
Figure 2.
Figure 2.
Immunohistochemistry for VGLUT1 in the hippocampus of the (A) adolescent PM1, adult (B) PM6 and (C) PM12, and aged (D) PM18 and (E) PM24 groups. VGLUT1 immunoreactivity was observed in the mossy fibers (indicated by MF) of the dentate gyrus as well as the Schaffer collaterals (indicated by SC) and stratum lacunosum-moleculare (indicated by SLM). VGLUT1 immunoreactivity was detected in the inner molecular layer (indicated by iML) in the PM6 and PM24 groups. Scale bar=500 µm. (F) ROD are expressed as a percentage of the value of the VGLUT1 immunoreactivity in the PM1 group in the dentate gyrus, hippocampal CA1 and CA2/3 regions per section of the PM1, PM6, PM12, PM18 and PM24 groups (n=5/group). Data are presented as the mean ± standard error mean. *P<0.05 vs. PM1. GLUT, vesicular glutamate transporter; PM, postnatal month; ROD, relative optical densities.
Figure 3.
Figure 3.
Immunohistochemistry for VGLUT2 in the hippocampus of the (A) adolescent PM1, adult (B) PM6 and (C) PM12, and aged (D) PM18 and (E) PM24 groups. VGLUT2 immunoreactivity was observed in the granule cell layer (indicated by GCL) and the outer molecular layer (indicated by oML), stratum pyramidale (indicated by SP), Schaffer collaterals (indicated by SC) and stratum lacunosum-moleculare (indicated by SLM) of the hippocampus. VGLUT2 immunoreactivity as markedly increased in the stratum radiatum (indicated by SR) and oriens (indicated by SO) of the CA1 region in the PM18 group. VGLUT2 immunoreactivity decreased in all layers of the hippocampus in the PM24 group. Scale bar=500 µm. (F) ROD are expressed as a percentage of the value of the VGLUT2 immunoreactivity in the PM1 group in the dentate gyrus, hippocampal CA1 and CA2/3 regions per section of the PM1, PM6, PM12, PM18 and PM24 groups (n=5/group). Data are presented as the mean ± standard error mean. *P<0.05 vs. the PM1 group; #P<0.05, vs. the PM6 group; $P<0.05 vs. the PM12 group; &P<0.05 vs. the PM18 group. VGLUT, vesicular glutamate transporter; PM, postnatal month; ROD, relative optical densities.
Figure 4.
Figure 4.
Western blot analysis. Protein levels are expressed as a percentage of VGLUT1 and VGLUT2 protein levels normalized by their respective β-actin protein level in the hippocampal homogenates of the PM1, PM6, PM12, PM18 and PM24 groups (n=5/group). Data are presented as the mean ± standard error mean. *P<0.05 vs. PM1 group; #P<0.05 vs. PM18 group.

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