Synaptic mechanisms of sound intensity coding in the cochlea
by Nare Karagulyan
Date of Examination:2024-11-08
Date of issue:2025-10-02
Advisor:Prof. Dr. Tobias Moser
Referee:Prof. Dr. Erwin Neher
Referee:Prof. Dr. Dr. Oliver M. Schlueter
Persistent Address:
http://dx.doi.org/10.53846/goediss-11524
Files in this item
Name:Thesis.pdf
Size:28.4Mb
Format:PDF
The following license files are associated with this item:
Abstract
English
In the mammalian cochlea sensory inner hair cells (IHCs) convert the mechanical sound waves into receptor potential and transmit the signal to the spiral ganglion neurons (SGNs) via afferent synapses. SGNs receiving input from a single IHC display diverse spontaneous rates, sound thresholds and dynamic ranges, thereby enabling a wide dynamic range of sound intensity coding. The position dependent heterogeneity of the IHC active zones (AZs) is one of the candidate mechanisms for SGN diversity. In this thesis we explored the formation and function of IHC synapses by focusing on three key areas: the signaling mechanisms underlying IHC-SGN synaptic connectivity, the signaling mechanisms setting the position dependent heterogeneity of IHC AZs, and the relationship between presynaptic heterogeneity and SGN diversity. First, we investigated the signaling mechanisms that ensure proper IHC innervation by SGNs focusing on the Nogo receptor homolog NgR2 and signaling molecule Semaphorin3A (Sema3A). By employing serial block-face electron microscopy and immunohistochemistry, we found that NgR2 deletion in mice (NgR2-/-) resulted in a subset of “orphan” postsynaptic densities (PSDs) and type I SGN neurites that did not make a contact with IHCs. Furthermore, the presynaptic AZs displayed a depolarized operation range of Ca2+ channels and the PSDs seemed to lack GluA2/3 subunits of AMPA receptors. These synaptic changes led to impaired auditory responses. We therefore postulate a critical role of NgR2 in afferent synapse formation and normal hearing. In contrast to NgR2, mice with attenuated Sema3A signaling (Sema3AK108N) did not show any gross changes of IHC innervation pattern, but the acute exposure of Sema3A to developing cultured wild-type cochleae, resulted in decreased SGN activity and increased arborization, implying the role of Sema3A in activity dependent regulation of IHC innervation. Second, we examined the signaling mechanisms contributing to the position-dependent heterogeneity of IHC AZs by employing immunohistochemistry and patch-clamp combined with single synaptic Ca2+ imaging. We found that while Sema3A signaling resulted in altered postsynaptic gradients of GluA2 receptor patches and SGN fiber caliber, the presynaptic AZ spatial gradients, such as ribbon size, maximal Ca2+ influx, and voltage of half-maximal activation, remained intact. Furthermore, we investigated the role of synaptic activity in establishing position dependent heterogeneity of IHC AZs by studying mice with abolished glutamatergic signaling (Vglut3-/-) and evoked exocytosis (OtofTDA/TDA). Our findings indicated that the spatial gradients of AZ properties in IHCs were largely unaffected, implying that synaptic transmission is dispensable for establishing these gradients. Lastly, we explored the relationship between presynaptic heterogeneity and SGN diversity, particularly assessing the influence of presynaptic Cav1.3 gating on SGN physiological properties. We made use of mice harboring a point mutation in Cav1.3 channels (Cav1.3AG), which was previously shown to result in channel activation at lower voltages. By performing Ca2+ and glutamate imaging at single AZ level in IHCs, we showed approximately 15 mV hyperpolarized shift in voltage dependency of both Ca2+ influx and glutamate release. In-vivo single unit recordings from SGNs revealed increased spontaneous rates and the auditory thresholds assessed from auditory brainstem responses were reduced in the mutant mice. These results directly demonstrate the regulation of SGN firing by presynaptic Cav1.3 gating and imply that the heterogeneous voltage dependence of Ca2+ channel activation at IHCs AZs governs the firing diversity of SGNs.
Keywords: Inner hair cells; Spiral ganglion neurons; Ribbon synapses; Calcium channels; Synaptic adhesion molecules; Synaptic heterogeneity; Synapse formation; Intensity coding
