Sex-specific modulation of cell proliferation by socially relevant stimuli in the adult green treefrog brain (Hyla cinerea)

Abstract

Social experience plays an important role in regulating the neural, physiological and hormonal changes that accompany the expression of reproductive behavior in vertebrates. This suite of functions is sexually dimorphic, with different neural control areas preeminent in males and females. In anuran amphibians, social experience comes in the form of acoustic communication, which is central to their reproductive behavior. We sought to determine whether acoustic cues regulate cell proliferation in the brain of adult green treefrogs (Hyla cinerea). Our results show that both male and female treefrogs that heard a conspecific chorus during the breeding season exhibited increased brain cell proliferation compared to animals that heard random tones. Increased cell proliferation, as assessed by the number of 5-bromo-2'-deoxyuridine-immunoreactive (BrdU+) cells, were found near the ventricles of acoustically sensitive brain regions such as the preoptic area (POA) and the infundibular hypothalamus (IF). Sex differences emerged in the location of this socially modulated cell proliferation: increases occurred primarily in the male POA and the female IF. In addition, gonadal steroid hormones might have played a role in the social modulation of cell proliferation: by statistically control- ling for hormone level, we revealed that androgens might influence socially induced increases in BrdU+ cells in the male POA, but estrogen did not contribute to socially induced increases in the female IF. These results indicate that the reception of social cues increases cell proliferation in brain regions mediating sexual behavior and endocrine regulation, and moreover that social modulation of cell proliferation occurs in a sexually differentiated fashion.

Fig. 1

Schematics (top rows) and photomicrographs (bottom rows) of brain sections showing adult cell proliferation in Hyla cinerea. Sections (B, C) are from a representative male in the Tone condition and (E, F) from a representative female in the Tone condition. A top-view drawing of the H. cinerea brain is oriented with anterior to the left. The lines indicate the level of the sections through the POA (A) and IF (D). Photomicrographs of transverse sections through the brain show the distribution of proliferating cells (i.e., dark brown cells) labeled with BrdU in the POA (B, C) and the IF (E, F); sections are Nissl-stained with toluidine blue, which labels non-proliferating cells blue. Photomicrographs are shown in increasing magnification down the left and right panels. The dotted line boxes represent the location of the subsequent photomicrograph. Scale bar = 500 μm for A, D and 50 μm for B, C, E, F. See list of abbreviations.

Fig. 2

The total number of BrdU+ cells in the POA (left side) and IF (right side) of Chorus- and Tone-stimulated Hyla cinerea. Chorus-stimulated males and females exhibited increased cell proliferation in acoustically sensitive brain regions, the POA and IF, compared to Tone controls. Shown are the mean number of cells counted [± the standard error of the mean (SEM)]. Asterisks (*) represent significant between-subjects tests at p < 0.05.

Fig. 3

Proliferating cells, as labeled by BrdU, were socially modulated in a sex- and region-specific manner. (A) In male H. cinerea, hearing their conspecific chorus increased BrdU+ cell numbers in the POA compared to Tone controls. (B) In females, hearing their conspecific chorus increased BrdU+ cell numbers in the IF compared to Tone controls. Shown are means ± SEM; asterisks (*) represent significant post hoc tests at p < 0.05.

Fig. 4

The relationship between gonadal steroid level and BrdU+ cells in male and female Hyla cinerea. (A) There was a significant positive correlation between androgen level and BrdU+ cells in the POA of males (r = 0.659, p = 0.005). (B) In females, there was a significant negative correlation between BrdU+ cells in the IF and estrogen level (r = −0.466, p = 0.033).

Fig. 5

The influence of acoustic stimulation and calling behavior on cell proliferation in the POA of male Hyla cinerea. (A) Callers (i.e., those that called back to stimulation) in response to Chorus stimulation exhibited increased cell proliferation in the POA compared to Non-callers. Shown are means ± SEM. The asterisk (*) represents a significant post hoc test at p < 0.05. (B) The number of evoked calls was positively correlated with cell proliferation in the male POA (r = 0.659, p = 0.003).