Transplantation of Ears Provides Insights into Inner Ear Afferent Pathfinding Properties

Abstract

Numerous tissue transplantations have demonstrated that otocysts can develop into normal ears in any location in all vertebrates tested thus far, though the pattern of innervation of these transplanted ears has largely been understudied. Here, expanding on previous findings that transplanted ears demonstrate capability of local brainstem innervation and can also be innervated themselves by efferents, we show that inner ear afferents grow toward the spinal cord mostly along existing afferent and efferent fibers and preferentially enter the dorsal spinal cord. Once in the dorsal funiculus of the spinal cord, they can grow toward the hindbrain and can diverge into vestibular nuclei. Inner ear afferents can also project along lateral line afferents. Likewise, lateral line afferents can navigate along inner ear afferents to reach hair cells in the ear. In addition, transplanted ears near the heart show growth of inner ear afferents along epibranchial placode-derived vagus afferents. Our data indicate that inner ear afferents can navigate in foreign locations, likely devoid of any local ear-specific guidance cues, along existing nerves, possibly using the nerve-associated Schwann cells as substrate to grow along. However, within the spinal cord and hindbrain, inner ear afferents can navigate to vestibular targets, likely using gradients of diffusible factors that define the dorso-ventral axis to guide them. Finally, afferents of transplanted ears functionally connect to native hindbrain vestibular circuitry, indicated by eliciting a startle behavior response, and providing excitatory input to specific sets of extraocular motoneurons.

Topologically correct central projections of tetrapod inner ear afferents require Fzd3

“…If this is the case, Wnt ligands released by the hindbrain could directly bind to inner ear afferents through Fzd3 receptors, facilitating their targeting. Wnt ligands are dorsally expressed in the hindbrain and spinal cord across species 66,67 and previous ear transplantation experiments support the targeting of inner ear afferents to a dorsal source of Wnt 58,64 . Verifying this hypothesis will entail creating neuron specific knockouts of Fzd3 along with identification of the Wnt ligand(s) and its specific source.…”

“…Our previous data indicated that diffusible factors play a substantial role in guidance of central projections of inner ear afferent neurons 58,64,65 . That data, combined with the necessity of Fzd3 and Prickle1 for proper inner ear afferent targeting, suggests Wnt/PCP signaling may be mediating this process.…”

“…To three-dimensionally reconstruct individual afferents from X. laevis mutants, the LIF files obtained from the Leica SP5 confocal microscope were loaded into Amira version 5.4 software for manual segmentation as described previously 74 . Fibers were individually traced and reconstructed as previously described for dendrites 75 and central projections 64 . Fibers for inner ear afferents were reconstructed separately from those of the lateral line and trigeminal by segmenting each independently.…”

Primary sensory map formations reflect unique needs and molecular cues specific to each sensory system

“…The ability to form functional connections with the hindbrain does not depend on the entry point of vestibular ganglion processes into the hindbrain. Functional rerouting to the vestibular nucleus of afferents from transplanted ears that entered into the spinal cord has been demonstrated 138 . Transplantation and rotation of a third ear to generate non-matching stimulation relative to the native ear’s sensory epithelia result in “vestibular dominance columns” 139 .…”

Vestibular Influence on Vertebrate Skeletal Symmetry and Body Shape

“…This suggests the presence of symmetric levels of bilateral utricular afferent activity despite the absence of otoconia with potential, yet so far unknown, consequences for body shape and skeletal organization. Addition of ectopic third ears, either rostrally onto the head (Elliott et al, 2015) or caudally along the tail (Gordy et al, 2018), which imparts asymmetry through supernumerary input, does not appear to produce visible body deformations. This suggests that body symmetry might be correctly established as long as bilateral otolithic signals are available and the signal difference between vestibular neurons on both sides is sufficiently small to be centrally offset through commissural connections (Straka, 2020).…”