Multiple mechanosensory modalities influence development of auditory function

Abstract

Sensory development can be dependent on input from multiple modalities. During metamorphic development, ranid frogs exhibit rapid reorganization of pathways mediating auditory, vestibular, and lateral line modalities as the animal transforms from an aquatic to an amphibious form. Here we show that neural sensitivity to the underwater particle motion component of sound follows a different developmental trajectory than that of the pressure component. Throughout larval stages, cells in the medial vestibular nucleus show best frequencies to particle motion in the range from 15 to 65 Hz, with displacement thresholds of <10 mum. During metamorphic climax, best frequencies significantly increase, and sensitivity to lower-frequency (<25 Hz) stimuli tends to decline. These findings suggest that continued sensitivity to particle motion may compensate for the considerable loss of sensitivity to pressure waves observed during the developmental deaf period. Transport of a lipophilic dye from peripheral end organs to the dorsal medulla shows that fibers from the saccule in the inner ear and from the anterior lateral line both terminate in the medial vestibular nucleus. Saccular projections remain stable across larval development, whereas lateral line projections degenerate during metamorphic climax. Sensitivity to particle motion may be based on multimodal input early in development and on saccular input alone during the transition to amphibious life.

Figure 1.

Sample calibration curve for z-axis acceleration based on input from the accelerometers (102 mV/m/s²) expressed in decibels of dB attenuation (94 dB sound pressure level = 102 mV rms). Solid horizontal line represents voltage reference level (−25.6 dB). The system is essentially flat (±4 dB) across the frequency range of 15–65 Hz.

Figure 2.

Schematics of the medulla (caudal to rostral, from top to bottom) based on sections from a stage 36 tadpole. Left side of each schematic shows cresyl violet-stained material, and the right side shows lesion sites (black circles, reflected to the right for clarity). Nuclear boundaries are based on published materials and our own in-house atlases. IX, Ninth cranial nerve nucleus; RG, reticular gray; SON, superior olivary nucleus. Scale bar, 500 μm.

Figure 3.

Developmental changes in auditory tuning. a, Tuning curves from representative animals in each developmental group showing threshold in displacement (micrometers) at best frequency (hertz). Data are derived from tadpoles at stages 30 (multiple-unit recording), 36 (multiple-unit recording), 41 (single-unit recording), and 42 (multiple-unit recording). b, Differences in best frequency (hertz) by developmental group for the entire dataset (n = 24). One-way ANOVA indicates that BFs for the three groups are significantly different (p < 0.05). Post hoc tests (Tukey's HSD test) indicate that the larval and climax groups show the greatest difference. c, Changes in displacement thresholds by developmental group for the entire dataset. Although thresholds are absolutely lower in climax compared with larval groups, the difference is not statistically significant. Error bars are SD.

Figure 4.

Period histograms from representative late larval (a–c, multiple-unit recording, BF of 40.6 Hz, threshold at BF of 1.52 μm), deaf period (d–f, single-unit recording, BF of 46.1 Hz, threshold at BF of 9.6 μm), and climax (g–i, multiple-unit recording, BF of 45 Hz, threshold at BF of 6.2 μm) tadpoles in response to low-frequency (a, d, g), middle-frequency (b, e, h), and high-frequency (c, f, i) stimuli. Stimulus frequency, displacement values, and VS are listed by each histogram. Asterisk indicates significant VS based on the Rayleigh's test. Bin width for period histograms is 1 ms. The vertical axis shows the number of spikes per bin, and the horizontal axis shows one stimulus period in milliseconds.

Figure 5.

Changes in best synchronized frequency by Gosner stage for all animals. Best synchronized frequency is determined based on highest vector strength for any stimulus frequency that is significant based on the Rayleigh's test. Open circles show single-unit recordings, and filled circles show multiple-unit recordings. The black line is a first-order regression line (r² = 0.259) through the entire dataset; vertical dashed lines demarcate the deaf period.

Figure 6.

Trichrome staining of saccule and medulla. Trichrome-stained coronal sections, 10 μm thick, of the inner ear of stage 29 (a) and stage 44 (b) tadpole showing the saccule. Black arrows indicate the hair-cell layer of the sensory macula. Lateral is to the right, and dorsal is up. c, Coronal section, 10 μm thick, of the medulla of a stage 25 bullfrog tadpole at the level of the insertion of the anterior (saccular) branch of nVIII. Fibers above the white dotted line are typically saccular. White arrows indicate saccular fibers traversing the medulla toward the DMN and MVN but dorsal to the LLa. Higher magnification shows terminations in and near the MVN region. Scale bars: a, b, 200 μm; c, 100 μm.

Figure 7.

Medullary termination of the saccular branch of nVIII across development as shown by DiI-Ph tracing (bright red). a, Fiber projection pattern from the saccular branch of nVIII in a stage 28 (early larval) tadpole. Fibers course through the DMN and dorsal MVN and, to a lesser extent, through the LLnp. Terminations are seen in the DMN and MVN but not in the LLnp or LLa. Similar patterns are observed in stage 38 (deaf period) (b) and stage 42 (metamorphic climax) (c) tadpoles. Note that the position of the DMN changes across larval development from a more lateral position in early larval animals to a more medial and dorsal position in metamorphic climax. The area of the LLnp is demarcated by the white polygon. This region becomes relatively more compressed as the DMN moves more dorsally and medially. d, Terminal and puncta label in the MVN of a stage 45 tadpole. White arrowheads indicate putative terminals. e, Label of the saccular end organ in a stage 45 tadpole, showing label of supporting and hair cells (HC, white arrows). Scale bars: a–c, 200 μm; d, 10 μm; e, 20 μm.

Figure 8.

Label of the dorsal medulla resulting from DiI-Ph deposition in the anterior lateral line nerve of a stage 30 tadpole. a, Filled white arrows indicate the course of labeled fibers from the entry of nVIII (to the right; data not shown) through the LLnp. Open arrows indicate filled cells in and around the LLa. b, Higher-power image showing puncta and terminal label in both the MVN and LLnp. Dotted lines indicate the MVN. Lateral is to the right, and dorsal is up in all figures. Scale bars, 100 μm.