Neural activity patterns in response to interspecific and intraspecific variation in mating calls in the túngara frog

Abstract

Background: During mate choice, individuals must classify potential mates according to species identity and relative attractiveness. In many species, females do so by evaluating variation in the signals produced by males. Male túngara frogs (Physalaemus pustulosus) can produce single note calls (whines) and multi-note calls (whine-chucks). While the whine alone is sufficient for species recognition, females greatly prefer the whine-chuck when given a choice.

Methodology/principal findings: To better understand how the brain responds to variation in male mating signals, we mapped neural activity patterns evoked by interspecific and intraspecific variation in mating calls in túngara frogs by measuring expression of egr-1. We predicted that egr-1 responses to conspecific calls would identify brain regions that are potentially important for species recognition and that at least some of those brain regions would vary in their egr-1 responses to mating calls that vary in attractiveness. We measured egr-1 in the auditory brainstem and its forebrain targets and found that conspecific whine-chucks elicited greater egr-1 expression than heterospecific whines in all but three regions. We found no evidence that preferred whine-chuck calls elicited greater egr-1 expression than conspecific whines in any of eleven brain regions examined, in contrast to predictions that mating preferences in túngara frogs emerge from greater responses in the auditory system.

Conclusions: Although selectivity for species-specific signals is apparent throughout the túngara frog brain, further studies are necessary to elucidate how neural activity patterns vary with the attractiveness of conspecific mating calls.

Figure 1. Waveforms and spectrograms of the calls that we used to represent conspecific (Physalaemus pustulosus) and heterospecific (Physalaemus enesefae) mating calls in order to identify brain regions that contribute to species recognition.

Figure 2. Waveforms and spectrograms of the túngara frog (Physalaemus pustulosus) calls that we used to represent the whine and whine-chuck in order to examine responses of brain regions to mating calls that vary in their attractiveness.

Figure 3. Diagram of the major ascending and descending connections in the frog auditory system.

Abbreviations: A, anterior thalamus; AP, amphibian papilla; BP, basilar papilla; C, central thalamus; DMN, dorsal medullary nucleus; Ltor, laminar nucleus of the torus semicircularis; MCtor, magnocellular nucleus of the torus semicircularis; MP, medial pallium; P, posterior thalamus; POa, anterior preoptic nucleus; Ptor, principal nucleus of the torus semicircularis; S, septum; SON, superior olivary nucleus; Str, striatum; VH, ventral hypothalamus.

Figure 4. Effects of interspecific variation in mating calls on egr-1 mRNA expression in the auditory brainstem and its forebrain targets.

Data are shown as mean (± SE) fold change in silver grains per cell above background relative to the no sound group. Sample sizes are indicated for each group and letters above bars indicate groups that are statistically different (p<0.05). Abbreviations: A, anterior thalamus; C, central thalamus; DMN, dorsal medullary nucleus; dMP, dosal part of the medial pallium; Ltor, laminar nucleus of the torus semicircularis; MCtor, magnocellular nucleus of the torus semicircularis; P, posterior thalamus; POa, anterior preoptic nucleus; Ptor, principal nucleus of the torus semicircularis; Slv, ventrolateral septal nucleus; SON, superior olivary nucleus; VH, ventral hypothalamus; vStr, ventral striatum.

Figure 5. Brightfield images (left column) and inverted darkfield images of transverse sections showing egr-1 expression within sampling windows (boxes) in response to conspecific whine-chucks (middle column) and heterospecific whines (right column) in the auditory brainstem (A–B) and thalamus (C–E).

Scale bars represent 400 µm. Abbreviations: A, anterior thalamus; C, central thalamus; DMN, dorsal medullary nucleus; Fr, reticular formation; La, lateral thalamus; LH, lateral hypothalamus; Ltor, laminar nucleus of the torus semicircularis; OT, optic tectum; P, posterior thalamus; Ptor, principal nucleus of the torus semicircularis; SC, suprachiasmatic nucleus; SON, superior olivary nucleus; Teg, tegmentum; Tel, telencephalon; VH, ventral hypothalamus; VL, ventrolateral thalamus; VM, ventromedial thalamus.

Figure 6. Brightfield images (left column) and inverted darkfield images of transverse sections showing egr-1 expression within sampling windows (boxes) in response to conspecific whine-chucks (middle column) and heterospecific whines (right column) in the anterior preoptic nucleus (A), septum (B), striatum (C), and medial pallium (D).

Scale bars represent 400 µm. Abbreviations: Acc, nucleus accumbens; dMP, dorsal part of the medial pallium; DP, dorsal pallium; dStr, dorsal striatum; LP, lateral pallium; MP, medial pallium; POa, anterior preoptic nucleus; Sd, dorsal septal nucleus; Sl, lateral septal nucleus, Sld, dorsolateral septal nucleus; Slv, ventrolateral septal nucleus; Sm, medial septal nucleus; Str, striatum; vMP, ventral part of the medial pallium; VP, ventral pallium; vStr, ventral striatum.

Figure 7. Effects of intraspecific variation in mating calls on egr-1 mRNA expression in the auditory brainstem and its forebrain targets.

Data are shown as mean (± SE) fold change in silver grains per cell above background relative to the no sound group. Sample sizes are indicated for each group. The bars above the columns indicate statistical comparisons between females hearing conspecific calls (whine or whine +3 chucks) to those hearing no sound and between females hearing the preferred, whine +3 chucks to those hearing whines; p values are indicated as follows: asterisks indicate p<0.05, actual p values are given for those tests where 0.050.2. Abbreviations: A, anterior thalamus; C, central thalamus; dMP, dosal part of the medial pallium; Ltor, laminar nucleus of the torus semicircularis; MCtor, magnocellular nucleus of the torus semicircularis; P, posterior thalamus; POa, anterior preoptic nucleus; Ptor, principal nucleus of the torus semicircularis; Slv, ventrolateral septal nucleus; SON, superior olivary nucleus; vStr, ventral striatum.