Age-induced disruption of selective olfactory bulb synaptic circuits

Abstract

Little is known about how normal aging affects the brain. Recent evidence suggests that neuronal loss is not ubiquitous in aging neocortex. Instead, subtle and still controversial, region- and layer-specific alterations of neuron morphology and synapses are reported during aging, leading to the notion that discrete changes in neural circuitry may underlie age-related cognitive deficits. Although deficits in sensory function suggest that primary sensory cortices are affected by aging, our understanding of the age-related cellular and molecular changes is sparse. To assess the effect of aging on the organization of olfactory bulb (OB) circuitry, we carried out quantitative morphometric analyses in the mouse OB at 2, 6, 12, 18, and 24 mo. Our data establish that the volumes of the major OB layers do not change during aging. Parallel to this, we are unique in demonstrating that the stereotypic glomerular convergence of M72-GFP OSN axons in the OB is preserved during aging. We then provide unique evidence of the stability of projection neurons and interneurons subpopulations in the aging mouse OB, arguing against the notion of an age-dependent widespread loss of neurons. Finally, we show ultrastructurally a significant layer-specific loss of synapses; synaptic density is reduced in the glomerular layer but not the external plexiform layer, leading to an imbalance in OB circuitry. These results suggest that reduction of afferent synaptic input and local modulatory circuit synapses in OB glomeruli may contribute to specific age-related alterations of the olfactory function.

Fig. 1.

Mouse OB volume is stable during aging. (A) OB layers delineation. (Scale bar, 500 μm.) (B) Total OB Volume. (C) Glomerular layer volume. (D) EPL volume. (E) GCL volume. (F) Contribution of each layer. (n = 3). GL, glomerular layer; ONL, olfactory nerve layer.

Fig. 2.

Broad OSN axon projections on the aging OB are stable. (A) Example of VGluT2 immunostaining. (Scale bar, 500 μm.) (B) Mean glomerular diameter. (C) Number of glomeruli per section. (D) Total number of glomeruli per OB. (n = 3).

Fig. 3.

Convergence of OSN axon subpopulations to the mouse OB is stable during aging. (A) Whole-mount OB of M72-GFP mice. Two M72-GFP glomeruli are located on the dorsolateral surface (arrows) and posterior ventromedial region (arrowheads). (Scale bar, 1 mm) (B) Confocal analysis from boxed areas in A. M72-GFP axons enter glomeruli at multiple entry sites (arrows) and some follow tortuous courses (arrowheads). (Scale bar, 100 μm.) (C and D) Angle and distance were used to map the position of the M72-GFP glomerulus. (n = 4–8; **, P < 0.01; ***, P < 0.001). A, anterior; L, lateral; M, medial; P, posterior.

Fig. 4.

Mouse OB neuronal populations are stable during aging. (A) Mitral cell, (B) granule cell, and (C) NeuN⁺ PG cell density (n = 3). (D) Partial colocalization of GAD67-GFP and NeuN: arrow and arrowhead designate GAD67-GFP⁺ NeuN⁺ and GAD67-GFP⁻ NeuN⁺ PG cells, respectively. (Scale bar, 50 μm.) (E) TH⁺ PG cell density.

Fig. 5.

Synaptic density decreases with aging in mouse glomeruli. (A–B') A:D and D:D synapses in the glomerular layer. Arrow designates the polarity of the synapse. (Scale bar in A, 100 nm.) (C) Total synapse density. (D) D:D synapse density. (E) A:D synapse density. (F) Ratio of A:D to D:D synapses. (n = 3; *, P < 0.05). Ax, OSN axon; Md, mitral cell dendrite; PGd, PG cell dendrite.

Fig. 6.

Synaptic density is stable during aging in the mouse EPL. (A–C') Asymmetrical, symmetrical, and reciprocal D:D synapses in the EPL. Arrow indicates the polarity of the synapse. (Scale bars: A–B', 100 nm; C, 300 nm; C', 200 nm.) (D) Total synapse density. (E) Asymmetrical synapse density. (F) Symmetrical synapse density. (G) Reciprocal synapse density. (n = 3). Grd, Granule cell dendrite; Md: mitral cell dendrite.