Adult Born Periglomerular Cells of Odorant Receptor Specific Glomeruli

Abstract

The OR37 subsystem is characterized by a variety of unique features. The odorant receptors (ORs) of this subfamily are selectively tuned to specific ligands which are supposed to play a role in social communication. OR37 expressing sensory neurons project their axons to a single receptor specific glomerulus per bulb which have been shown to be unusually stable in size and to possess a distinct repertoire of periglomerular cells. Since the neuronal network surrounding glomeruli is typically modified by the integration of adult born neurons, in this study it was investigated whether the number of adult born cells might be different for OR37 glomeruli compared to other OR-specific glomeruli. Towards this goal, 23 days after BrdU injection, BrdU labeled cells in the proximity of OR37A glomeruli as well as around OR18-2 and OR256-17 glomeruli were determined. It was found that the number of BrdU labeled cells in the periglomerular region of OR37A glomeruli was significantly lower compared to glomeruli of the other OR types. This finding was in line with a lower number of neuroblasts visualized by the marker protein doublecortin. Double labeling experiments for BrdU and marker proteins revealed that despite a relatively high number of calretinin expressing cells at the OR37A glomeruli, the number of cells co-stained with BrdU was quite low compared to other glomeruli, which may point to an individual turnover rate of this cell type for different glomeruli. Together, the results of the present study support the notion that the neuronal network at the OR37 glomeruli is less dynamic than that of other glomerulus types. This indicates a specific processing of social information in OR37 glomerular networks.

Keywords: BrdU; OR37; adult neurogenesis; doublecortin; glomeruli; olfactory bulb; periglomerular cells.

Figure 1

BrdU labeling in the olfactory bulb at different time points after injection. (A) BrdU immunoreactive cells (red) and DAPI staining (blue) in the granule cell layer of the olfactory bulb at different time intervals after BrdU injections. The number of BrdU signals visible in the granule cell layer increases from day 12 to 23 days after the last injection and declines again at day 26. Scale bar: 100 μm. (B) At 23 days after the last injection, a substantial number of BrdU immunoreactive cells (red) was also visible in the glomerular layer. DAPI staining is shown in blue. Scale bar: 100 μm.

Figure 2

BrdU positive cells in the vicinity of odorant receptor (OR) specific glomeruli. (A) Representative images of glomeruli for OR18-2, OR256-17, OR37A, OR37B and OR37C, respectively (green), with BrdU immunoreactive cells (magenta) and DAPI staining (blue). Scale bar: 50 μm. (B) The number of BrdU positive cells per 1000 DAPI stained cells in the periglomerular region of OR256-17, OR18-2 and OR37 glomeruli in mean ± SD with significantly lower values for OR37 glomeruli with p = 0.0023 compared to OR256-17 and p = 0.0256 compared to OR18-2 glomeruli. n.s. p > 0.05; *p < 0.05; **p < 0.01.

Figure 3

Doublecortin positive cells in the periglomerular region of specific glomeruli. (A) Exemplary confocal image of BrdU (magenta, white arrowheads) and doublecortin (red, yellow arrows) double staining around an OR256-17 glomerulus (green). No co-labeling of cells was observed. Scale bar: 50 μm. (B) Scatter plot depiction of values for individual glomeruli. The values obtained for individual glomeruli vary notably, with a tendency to more doublecortin positive cells around OR256-17 glomeruli, fewer around OR18-2 glomeruli and still less doublecortin positive cells around OR37A glomeruli. (C) The number of doublecortin positive cells does not correlate with the number of BrdU positive cells at 23 days after BrdU injection for individual glomeruli. n.s. p > 0.05.

Figure 4

Subtype analyses of adult-born cells in the periglomerular region. (A) Scatterplot depicting the total number of NeuN and BrdU double stained cells around individual glomeruli of OR256-17, OR18-2 and OR37A, respectively. Black boxes represent mean values. The total number of NeuN immunoreactive cells in mean ± SD around the different glomerulus types is shown in the bar graph. (B) Scatterplot depicting the total number of GAD65 and BrdU double stained cells around individual glomeruli of OR256-17, OR18-2 and OR37A, respectively. Black boxes represent mean values. The total number of GAD65 positive cells in mean ± SD around the different glomerulus types is shown in the bar graph. (C) Scatterplot depicting the total number of calbindin and BrdU double stained cells around individual glomeruli of OR256-17, OR18-2 and OR37A, respectively. Black boxes represent mean values. The total number of calbindin immunoreactive cells in mean ± SD around the different glomerulus types is shown in the bar graph. (D) Scatterplot depicting the total number of calretinin and BrdU double stained cells around individual glomeruli of OR256-17, OR18-2 and OR37A, respectively. Black boxes represent mean values. The total number of calretinin immunoreactive cells in mean ± SD around the different glomerulus types is shown in the bar graph. n.s. p > 0.05; *p < 0.05; ***p < 0.001.