Percentage of birds that displayed at least one mount attempt (A) or one cloacal contact movement (B) and cumulative frequencies of these two behaviors in the 5 experimental groups (C–D).

<p>Data were analyzed by appropriate analyses of variance or χ2 tests that were followed by post-hoc tests specifically comparing the 4 experimental groups to the controls (see text). Results of these post-hoc comparisons are shown at the top of the bars as follows: * =  p<0.05, ** =  p<0.01 and *** =  p<0.001.</p

Photomicrographs illustrating the aromatase-immunoreactive perikarya (A) and the vasotocin-immunoreactive fibers (B, C) present within the medial preoptic nucleus (POM) that were quantified in the present study.

<p>Panel A illustrates the dense group of aromatase-immunoreactive neurons that outline the entire POM. The dotted line marks the limits of the POM as they were defined for quantification. Panel B shows the accumulation of vasotocin-immunoreactive fibers in the POM at the level of the anterior commissure. The rectangle drawn with a solid line indicates the area where quantification was performed that is illustrated at higher magnification in panel C. The dotted rectangle indicates how the camera field was originally placed before being moved to its final location (see text). Note that quantification of fibers concerned the steroid-sensitive network located in the POM, not the denser network located more ventrally that originates from the magnocellular neurons. CA: commissural anterior, LFB: latera forebrain bundle. Magnification bar =  500 µm in A–B, 100 µm in C.</p

Sex differences in the percentage of DAPI cells that were immunoreactive for parvalbumin (%PV-ir cells) or were surrounded by perineuronal nets (%PNN cells), in the percentage of PV-ir cells surrounded by PNN (%PV with PNN) and in the percentage of PNN surrounding PV-ir cells (%PNN with PV) in 3 auditory areas (Field L; the dorsal lateral mesencephalic nucleus, MLd; the medial nucleus of the dorsolateral thalamus, DLM), and 3 visual areas (the nucleus rotondus, ROT; the nucleus spiriformis lateralis, SpL; the nucleus subpretectalis, SP).

<p>The figure shows the mean ± SEM of data in males (black bars) and females (open bars). Numbers of data in each case are listed in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0123199#pone.0123199.t001" target="_blank">Table 1</a>. * = p<0.05 and (*) = 0.10</p

Aromatase activity (mean±SEM) measured in different regions of gonadally intact male and female Japanese quail.

<p><b>A</b>. A mixed-design ANOVA with all regions considered individually revealed a significant effect of sex (F_1,17 = 9.284; p = 0.007) and brain region (repeated factor; F_5,85 = 20.640; p<0.001) as well as an interaction between these two factors (F_5,85 = 2.473; p = 0.038). <b>B</b>. A mixed-design ANOVA of results after pooling data for POM and mBST as well as for VMN and tuber (MBH) revealed significant effects of the sex (F_1,17 = 9.407; p = 0.007) and regions (F_3,51 = 43.392; p<0.001) as well as a significant interaction (F_3,51 = 4.776; p = 0.005). Individual means were compared by the Fisher Protected Least Significance Difference test. Comparisons between males and females within a given region are reported at the top of the columns as follows: (*) 0.05 </p

Representative photomicrographs illustrating the distribution of chrondroitin sulfate immunoreactivity (Green signal) and of parvalbumin-immunoreactive cells (red signal) as observed in in parasagittal sections of male (C, C) and female (B, D) zebra finch brains.

<p>Panels A_B illustrate differences in HVC, panels C-D concern RA. The magnification bar in D (20 μm) also applies to all other panels.</p

Dose-response curve of labelling index after incubation of HEK293T cells with BrdU.

<p>(A–F) Cultured cells stained for BrdU, after incubation with BrdU at different concentrations (as indicated at the bottom of each image). Images were consistently modified to enhance contrast and remove colour, and are all shown to the same scale. (A) At very low concentrations, very few cells were labelled even weakly. (B–F) As BrdU concentration increased, the proportion of visible cells that were weakly-labelled (w), moderately-labelled (m), or strongly-labelled (s) increased. Every culture plate also contained some unlabelled cells (arrowhead). (G) Total numbers of labelled cells (weakly+moderately+strongly) reached a plateau with BrdU concentrations above 50 µg/mL; note the break in the x-axis scale<b>.</b> At lower concentrations, however, (inset) there was a strong linear relationship between BrdU concentration and the total number of labelled cells (Y = 1.651 X, line forced through [0,0]; confidence interval for slope = 1.403 to 1.898, r = 0.9606, p<0.0001). Dashed lines show 95% confidence interval around the (solid) line of best fit.</p

Sex differences in the percentage of DAPI cells that were immunoreactive for parvalbumin (%PV-ir cells) or were surrounded by perineuronal nets (%PNN cells), in the percentage of PV-ir cells surrounded by PNN (%PV with PNN) and in the percentage of PNN surrounding PV-ir cells (%PNN with PV) in 4 song control nuclei, HVC (used as a proper name), RA (robust nucleus of the arcopallium), Area X of the basal ganglia and LMAN (lateral magnocellular nucleus of the anterior nidopallium).

<p>The figure shows the mean ± SEM of data in males (black bars) and females (open bars). Numbers of data in each case are listed in <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0123199#pone.0123199.t001" target="_blank">Table 1</a>. ** = p<0.01, * = p<0.05 and (*) = 0.10</p

Labelling indices and deduced BrdU concentrations after incubation of HEK293T cells with serum from BrdU-injected animals.

<p>(A). Percentage (means ± SD) of the total numbers of cells visible in culture that were immunoreactive for BrdU after incubation with serum collected from adult canaries at various times after BrdU injection. All individual male (black spots) and female (open circles) data points are represented in the figure. (B) Average concentrations (means ± SD) of BrdU in these canary samples based on the calibration curve illustrated in the inset of <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0063692#pone-0063692-g001" target="_blank">Figure 1G</a>. (C–D) Concentrations of BrdU in blood samples collected in two female Japanese quail (C) or one male mouse (D) at various times after a single BrdU injection.</p

Cloacal gland area (in mm²) at the end of the experiment (A), percentage of birds that displayed at least one female-induced rhythmic cloacal sphincter movement (RCSM)(B) and cumulative frequencies of these RCSM in the 5 experimental groups (C).

<p>Data were analyzed by appropriate analyses of variance or c2 tests that were followed by post-hoc tests specifically comparing the 4 experimental groups to the controls (see text). Results of these post-hoc comparisons are shown at the top of the bars as follows: * =  p<0.05, ** =  p<0.01 and *** =  p<0.001.</p

Linear regression of number of EdU-positive cells versus number of BrdU-positive cells in the VZ of the combined sections of birds injected with EdU only 4 and 24 hours before brain collection.

<p>Linear regression of number of EdU-positive cells versus number of BrdU-positive cells in the VZ of the combined sections of birds injected with EdU only 4 and 24 hours before brain collection.</p