Olfactory enrichment modulates DCX and PSA-NCAM expression in new-generated olfactory interneurons.

<p><i>A,</i> Quantification of BrdU-positive PGCs double-labelled for DCX or PSA-NCAM among the total number of BrdU-positive cells counted in the GL at 10, 21 and 42 days survival in standard and enriched groups. Error bars indicate SEM. * p<0.05, ** p<0.01. <i>B,</i> Confocal analysis of olfactory bulb GL stained in green for BrdU and red for PSA-NCAM. <i>C,</i> Confocal analysis of olfactory bulb GL stained in green for BrdU and red for DCX. The arrows shows BrdU-positive neuron expressing PSA-NCAM (B) or DCX (C). Scale bar in C corresponds to 25 µm and applies to B. GL: glomerular layer.</p

Olfactory deprivation does not alter the relative proportion of new-generated PGCs expressing GAD67, CB and CR whereas it regulates TH expression in both new-generated and resident PGCs.

<p><i>A,</i> Experimental procedure. Adult mice were injected with BrdU 3 weeks after unilateral naris closure (four injections, 4-h interval, 50 mg/kg, i.p.). The quantification of neurogenesis (survival of BrdU-labelled cells) was done by counting BrdU-labelled cells in the GL at 21 days post-BrdU injection. <i>B,</i> Quantification of BrdU-positive PGCs double-labelled for GAD67, CB, CR and TH among the total number of BrdU-positive cells counted in the GL at 21 days survival in standard and deprived groups. Error bars indicate SEM. ** p<0.01. <i>C,</i> Confocal analysis of olfactory bulb GL stained in red for BrdU and green for TH, 21 days p.i. <i>D,</i> Total number of TH-positive cells in the GL of standard and deprived animals 42 days after unilateral naris closure. <i>E,</i> Quantification of TH-positive PGCs double-labelled for BrdU among the total number of TH-positive cells counted in the GL in standard and deprived groups 42 days after unilateral naris closure. Error bars indicate SEM. *** p<0.001. Scale bar in C corresponds to 25 µm.</p

BrdU-positive cell density increases in the GL and GCL of mice reared in enriched olfactory environment.

<p><i>A,</i> Experimental procedure. Adult mice were injected with BrdU 3 weeks after initial treatment (four injections, 4-h interval, 50 mg/kg, i.p.). The quantification of neurogenesis (survival of BrdU-labelled cells) was done by counting BrdU-labelled cells in the GL and GCL at 10, 21 and 42 days post-BrdU injection (p.i.). <i>B,</i> Mean number of BrdU-IR cells per millimetres cubed in the GL of standard and enriched mice at 10, 21 and 42 days p.i.. <i>C,</i> Mean number of BrdU-IR cells per millimetres cubed in the GCL of standard and enriched mice at 10, 21 and 42 days p.i.. Error bars indicate SEM. * p<0.05, ** p<0.01. St-1: standard at 10 days p.i.; St-2: standard at 21 days p.i.; St-3: standard at 42 days p.i.; EN-1: enriched at 10 days p.i.; EN-2: enriched at 21 days p.i.; EN-3a: enriched at 42 days p.i; EN-3b: animals enriched 21 days p.i. and returned to standard housing conditions for 21 more days.</p

Olfactory enrichment does not alter the relative proportion of new-generated PGCs expressing CB, CR and TH.

<p><i>A,</i> Quantification of BrdU-positive PGCs double-labelled for CB, CR and TH among the total number of BrdU-positive cells counted in the GL at 21 days survival in standard and enriched groups. Error bars indicate SEM. <i>(B–D)</i> Confocal analysis of olfactory bulb GL stained in red for BrdU and green for CB (B), CR (C) and TH (D) in standard mice. The arrows in C and D show a BrdU-positive neuron expressing CR (higher magnification in inset in C; single confocal plane) and TH (higher magnification in inset in D; single confocal plane). Scale bar in D corresponds to 25 µm in D, B and C and to 10 µm in insets in C and D. GL: glomerular layer.</p

Olfactory enrichment modulates GAD67 expression in new-generated olfactory interneurons.

<p><i>A,D</i> Quantification of BrdU-positive PGCs <i>(A)</i> and GCs <i>(D)</i> double-labelled for GAD67 among the total number of BrdU-positive cells counted in the GL and GCL at 10, 21 and 42 days survival in standard and enriched groups. <i>B,E</i> Confocal analysis of olfactory bulb GL <i>(B)</i> and GCL <i>(E)</i> stained in green for BrdU and red for GAD67 in standard conditions. The arrow shows a BrdU-positive neuron expressing GAD67 (higher magnification in inset; single confocal plane). <i>C,F</i> signal intensity of GAD67 expression in the GL <i>(C)</i> and GCL <i>(F)</i> of standard and olfactory enriched mice. Error bars indicate SEM. ** p<0.01, *** p<0.001. Scale bar in E corresponds to 25 µm and applies to B. Scale bar in insets corresponds to 7 µm. GL: glomerular layer; GCL: granule cell layer.</p

Newborn interneurons in the accessory olfactory bulb promote mate recognition in female mice

<p>In the olfactory bulb of adult rodents, local interneurons are constantly replaced by immature precursors derived from the subventricular zone. Whether any olfactory sensory process specifically relies on this cell renewal remains largely unclear. By using the well known model of mating-induced imprinting to avoid pregnancy block, which requires accessory olfactory bulb (AOB) function, we demonstrate that this olfactory memory formation critically depends on the presence of newborn granule neurons in this brain region. We show that, in adult female mice, exposure to the male urine compounds involved in mate recognition increases the number of new granule cells surviving in the AOB. This process is modulated by male signals sensed through the vomeronasal organ and, in turn, changes the activity of the downstream amygdaloid and hypothalamic nuclei involved in the pregnancy block response. Chemical depletion of newly generated bulbar interneurons causes strong impairment in mate recognition, thus resulting in a high pregnancy failure rate to familiar mating male odors. Taken together, our results indicate that adult neurogenesis is essential for specific brain functions such as persistent odor learning and mate recognition.</p