Rapid, hierarchical modulation of vocal patterning by steroid hormones

Abstract

Vocal control systems have been identified in all major groups of jawed vertebrates. Although steroid hormones are instrumental in the long-term development and maintenance of neural structures underlying vocalization, it is unknown whether steroids rapidly modulate the neural activity of vocal motor systems. The midshipman fish generates advertisement and agonistic calls that mainly differ in duration. A descending midbrain pathway activates a hindbrain-spinal vocal circuit that directly establishes the discharge frequency and duration of the rhythmic vocal motor volley. This vocal motor output, which can be monitored from occipital nerve roots, directly determines the rate and duration of contraction of a pair of sonic muscles and, in turn, the fundamental frequency and duration of vocalizations. Here, we demonstrate that the duration of the vocal motor volley, or fictive vocalization, is rapidly responsive to steroid hormones, including androgens, estrogens, and glucocorticoids. These responses are consistent, in part, with a nongenomic mechanism and are steroid specific at the receptor level, suggesting the possibility of multiple membrane-bound receptor populations. We also show, using intact and semi-intact preparations, that steroids hierarchically modulate fictive vocalizations; whereas the hindbrain-spinal region is both necessary and sufficient for rapid (within 5 min) effects on duration, descending midbrain input is necessary for maintenance (up to 120 min) of these effects. The conserved nature of vertebrate vocal motor systems suggests that the neuroendocrine principles outlined in this study may be a fundamental feature of all vocal vertebrates.

Figure 1.

A, B, Sounds recorded from type I male midshipman in their natural habitat. Males emit short-duration grunts (A) during aggressive encounters and long-duration hums (B) to attract females. C, Sagittal view of the midshipman brain, showing forebrain, midbrain, and hindbrain-spinal vocal motor regions. Arrows indicate rostral and caudal transection sites used in the hindbrain-spinal isolation experiments; fVAC, mVAC, hVAC are forebrain, midbrain, and hindbrain vocal acoustic complexes, respectively (Goodson and Bass, 2002). D-G, Oscillograms of fictive vocalizations evoked by electrical stimulation in two experiments on type I male midshipman. Stimulus artifact is presented for reference on left side of each trace. Recordings shown are under baseline conditions (D, F) and either 15 min after 17β-estradiol injection (E; 0.02 mg/kg) or 120 min after 11KT injection (G; 0.2 mg/kg). Note in each case the rapid change in burst duration but neither discharge frequency nor latency after steroid administration.

Figure 2.

Rapid and sustained steroid modulation of vocal motor output during midbrain stimulation. Data are presented as mean ± SEM for Figures 2, 3, 4. Control experiments (oil injection; n = 6 animals) are plotted in each figure for comparison. A, Cortisol, the major circulating teleost glucocorticoid, rapidly increases vocalization duration after intramuscular injection. This effect is both rapid (within 5 min) and sustained (up to 60 min) for both low (n = 4) and high (n = 8) doses. B, 11KT, the major circulating teleost androgen, rapidly increases vocalization duration after intramuscular injection. This effect is both rapid (within 5 min) and sustained (up to 120 min) for low (n = 5), medium (n = 5), and high (n = 5) doses. C, 17β-Estradiol, a neuroactive steroid hormone, rapidly increases vocalization duration after intramuscular injection. This effect is rapid (within 5 min) but not sustained beyond 30 min for both low (n = 5) and high (n = 3) doses. D, Testosterone, the synthesis precursor to both 17β-estradiol and 11KT, causes no change in vocalization duration after intramuscular injection at either low (n = 3) or high (n = 5) dose.

Figure 3.

The effects of 11KT and cortisol on vocal motor activity are eliminated in the presence of specific steroid receptor antagonists. A, The anti-androgen CA blocks the effects of the androgen 11KT (n = 3), whereas the response to cortisol (CORT) remains unchanged in the presence of CA (n = 3). B, The anti-glucocorticoid mifepristone (RU486) blocks the effects of CORT (n = 3), whereas the response to 11KT remains unchanged in the presence of RU486 (n = 3). Either CA (n = 2) or RU486 (n = 2) alone did not significantly affect vocal motor activity.

Figure 4.

The hindbrain-spinal vocal center is both necessary and sufficient for rapid steroid modulation. After hindbrain-midbrain transection (A), both 11KT (0.04 mg/kg) and cortisol (0.05 mg/kg) rapidly increase the duration of vocal bursts after intramuscular injection. These effects are rapid (5, 15, and 30 min) but not sustained for both 11KT (n = 5) and cortisol (n = 3). Oil injection (n = 3) produced no change in vocal motor activity. B, 11KT hierarchically modulates the duration of vocal bursts elicited from the midbrain and hindbrain in the same intact preparation (n = 4). After injection, 11KT rapidly increases burst duration in the hindbrain (5, 15, and 30 min), but this effect is not sustained beyond 30 min. In contrast, parallel stimulation in the midbrain evokes responses to 11KT that are sustained through 45, 60, 90, and 120 min after injection.

Figure 5.

Summary figure depicting rapid, hierarchical modulation of midbrain and hindbrain-spinal vocal motor activity by steroid hormones. The hindbrain-spinal vocal circuit is both necessary and sufficient for the rapid effect of steroids (up to 30 min), whereas descending input from the midbrain is necessary for sustained effects of steroids (>30 min). The rapid and sustained effects of steroid hormones on vocal motor circuitry are represented by neurophysiological recordings shown in the center of the figure. Also shown is the temporal specificity that different steroid hormones exert on the midbrain-hindbrain vocal motor system, suggesting the presence of multiple receptor populations in the vocal motor system of midshipman.