Development of functional sex differences in the larynx of Xenopus laevis

Abstract

Three laryngeal properties associated with the production of masculine song--laryngeal muscle tension, fiber twitch type, and fiber recruitment--are markedly sexually dimorphic in adult Xenopus laevis frogs. To elucidate the pattern of sexual differentiation, tension and fiber recruitment in male and female larynges and fiber twitch type in male larynges were examined throughout postmetamorphic development. Masculinization of male laryngeal properties begins early in postmetamorphic development and continues until adulthood. In contrast, tension and fiber recruitment in females do not change after the end of metamorphosis. Laryngeal muscle tension and fiber type are gradually and progressively masculinized; the temporal pattern of masculinization is very similar for these properties. Fiber recruitment, on the other hand, appears to masculinize in a stepwise manner. Masculinization of all three properties is highly correlated with larynx weight in males. We have used this relation to divide postmetamorphic development into seven stages associated with key events in sexual differentiation. This staging scheme provides an important experimental tool for studying the hormonal regulation of sexual differentiation, the subject of the accompanying paper.

Fig. 1

Laryngeal weight as a function of body weight for males and females throughout postmetamorphic development. Although larynx weight is highly correlated with body weight in both sexes, males increase larynx weight more rapidly than females. Note that both body weights and laryngeal weights are sexually dimorphic in adults.

Fig. 2

Tension records from developing male and female larynges. Adult males typically produce 100% transient tension, while females typically produce mostly maintained tension with small transients superimposed. Tension (t) is sexually monomorphic at the end of metamorphosis and during early postmetamorphic development. The percentage of transient tension then increases in males but remains unchanged in females. Calibration bar: 50 ms. [Table: see text]

Fig. 3

The percentage of transient tension as a function of larynx weight in males and females throughout postmetamorphic development. The percentage of transient tension is sexually monomorphic early in development and then increases rapidly in males but remains unchanged in females.

Fig. 4

Photomicrographs of sections reacted for ATPase activity in male laryngeal muscle at three developmental stages. An example of a small dark muscle fiber (slow twitch) is indicated (arrow) at early developmental stages. A larger light muscle fiber (fast twitch) is also indicated (arrowhead) at each stage. At the end of metamorphosis (top, 9 mg larynx), male laryngeal muscle is composed of a heterogeneous population of fibers; 51% are slow twitch. During masculinization (middle, 102 mg larynx), male laryngeal muscle has 21% slow twitch fibers. In adulthood (bottom, 517 mg larynx), male laryngeal muscle is composed only of fast twitch fibers, no slow twitch fibers are present. Calibration bar: 10 μm.

Fig. 5

The percentage of slow twitch fibers as a function of larynx weight in males throughout postmetamorphic development. The percentage of slow twitch fibers begins to decline early in development and decreases rapidly. The loss of slow twitch fibers closely parallels the increase in the percentage of transient tension (Fig. 3).

Fig. 6

Electromyogram recordings from developing male and female larynges. In adulthood (bottom panels), male EMGs increase in amplitude throughout the stimulus train, while female EMGs have a nearly constant amplitude. At the end of metamorphosis (top panels), both sexes display a decrease in EMG amplitude in response to a stimulus train (EMG potentiation <1.0). Both sexes then increase EMG potentiation values somewhat (EMG potentiation ~1.0; middle panels). Males then further increase EMG potentiation during development while females do not. Calibration bar: 50 ms. [Table: see text]

Fig. 7

EMG potentiation as a function of larynx weight in males and females during postmetamorphic development. EMG potentiation is low in both sexes early in development. During postmetamorphic development, values increase in males and are not fully masculinized until adulthood. Female values remain low throughout postmetamorphic development.

Fig. 8

Average values for male percentage of slow twitch fibers, percentage of transient tension, and EMG potentiation at each stage in postmetamorphic development. Female values (mean of PM1 through PM6) are shown for comparison in percentage of transient tension and EMG potentiation graphs; adult female values (from Sassoon et al., 1987) are shown in percentage of slow twitch fibers graph. Bars represent mean ± SD. The number of animals included in each bar is indicated above the bar. All properties show a marked increase in masculinization between PM2 and PM3.