Supernumerary Formation of Olfactory Glomeruli Induced by Chronic Odorant Exposure: A Constructivist Expression of Neural Plasticity

It is accepted that sensory experience instructs the remodelling of neuronal circuits during postnatal development, after their specification has occurred. The story is less clear with regard to the role of experience during the initial formation of neuronal circuits, whether prenatal or postnatal, since this process is now supposed to be primarily influenced by genetic determinants and spontaneous neuronal firing. Here we evaluated this last issue by examining the effect that postnatal chronic exposure to cognate odorants has on the formation of I7 and M72 glomeruli, iterated olfactory circuits that are formed before and after birth, respectively. We took advantage of double knock-in mice whose I7 and M72 primary afferents express green fluorescent protein and β-galactosidase, correspondingly. Our results revealed that postnatal odorant chronic exposure led to the formation of permanent supernumerary I7 and M72 glomeruli in a dose and time dependent manner. Glomeruli in exposed mice were formed within the same regions of olfactory bulb and occupy small space volumes compared to the corresponding single circuits in non-exposed mice. We suggest that local reorganization of the primary afferents could participate in the process of formation of supernumerary glomeruli. Overall, our results support that sensory experience indeed instructs the permanent formation of specific glomeruli in the mouse olfactory bulb by means of constructivist processes

The largest glomeruli displayed primary afferent reorganization within a restricted space during chronic odorant exposure.

<p>The figure shows the expression of I7tauGFP glomeruli in non-exposed mice and mice exposed to heptaldehyde. Notice that primary afferents redistribute in variable ways between the largest and supernumerary glomeruli (arrows) in exposed mice. The effect was seen in all half bulbs analyzed (n = 10 for each condition) when mice were exposed from postnatal day (PD) 0 to PD5. In mice exposed from PD10 to PD15 the effect was seen in 8 out of 10 total half bulbs analyzed. Scale bar 100 µm; inset square is 2× magnification.</p

Adult mice were unable to form supernumerary glomeruli in response to chronic odorant exposure.

<p>Figures A and B show representative I7tauGFP glomeruli from non-exposed mice and from those exposed to 10% heptaldehyde, respectively; periglomerular neuron autofluorescence (arrows) is commonly seen in adults. Scale bar 100 µm. The bar graph in C depicts the average number of I7 glomeruli per half bulb in mice exposed during early postnatal stage (PD0–PD20) and in those exposed during adulthood (PD50–PD70). Mann-Whitney U test: ***, P<0.001.</p

Chronic odorant exposure led to the redistribution of the glomerular volume.

<p>The graph in A shows a volume histogram of the total population of I7 glomeruli in 20 half bulbs of non-exposed mice and in those exposed to 10% heptaldehyde. The bar graph in B depicts the average volume of the largest I7 glomerulus per half bulb and the average sum of the volume of I7 supernumerary glomeruli per half bulb in non-exposed mice and in those exposed to 10% heptaldehyde. Kruskal-Wallis sum rank test: p<0.01 followed by Dunn multiple comparison test: ***, P<0.001.</p

Supernumerary glomeruli grew in size after interrupting chronic odorant exposure.

<p>Figures A and B show representative I7tauGFP supernumerary glomeruli at postnatal day (PD) 20 and at PD50, respectively, from 10% heptaldehyde exposed mice. Scale bar 100 µm. Bar graph in C depicts the average I7 glomerular volume per half bulb, at PD20 and PD50, of the largest glomerulus in non-exposed mice, and the largest glomerulus and the sum of supernumerary glomeruli in mice exposed to 10% heptaldehyde (10% Hep). Non paired t-test: **, P<0.01; ***, P<0.001.</p

Supernumerary glomeruli persisted after interrupting chronic odorant exposure.

<p>Figures A and B show representative I7tauGFP glomeruli (green) in coronal brain slices stained with DAPI (red) from non-exposed mice and from those exposed to 10% heptaldehyde and later raised without the odorant until postnatal day 50. Arrowheads indicate supernumerary glomeruli. Scale bar 100 µm. Figure in C shows a schematic side view representation of the olfactory bulb, indicating the anterior ventrolateral position of I7 glomeruli in the coronal plane in non-exposed mice and in those exposed to 10% heptaldehyde, according to the location across brain slices. Bar graph in D shows the average number of I7 glomeruli per half bulb 30 days after the end of exposure. Kruskal-Wallis sum rank test: p<0.01 followed by Dunn multiple comparison test: ***, P<0.001.</p