The synaptic ribbon is critical for sound encoding at high rates and with temporal precision

Abstract

We studied the role of the synaptic ribbon for sound encoding at the synapses between inner hair cells (IHCs) and spiral ganglion neurons (SGNs) in mice lacking RIBEYE (RBEKO/KO). Electron and immunofluorescence microscopy revealed a lack of synaptic ribbons and an assembly of several small active zones (AZs) at each synaptic contact. Spontaneous and sound-evoked firing rates of SGNs and their compound action potential were reduced, indicating impaired transmission at ribbonless IHC-SGN synapses. The temporal precision of sound encoding was impaired and the recovery of SGN-firing from adaptation indicated slowed synaptic vesicle (SV) replenishment. Activation of Ca2+-channels was shifted to more depolarized potentials and exocytosis was reduced for weak depolarizations. Presynaptic Ca2+-signals showed a broader spread, compatible with the altered Ca2+-channel clustering observed by super-resolution immunofluorescence microscopy. We postulate that RIBEYE disruption is partially compensated by multi-AZ organization. The remaining synaptic deficit indicates ribbon function in SV-replenishment and Ca2+-channel regulation.

Balancing presynaptic release and endocytic membrane retrieval at hair cell ribbon synapses

“…In contrast, genetic deletion of RIBEYE‐A in mice resulted in the complete loss of presynaptic dense bodies from retinal photoreceptors, rod bipolar cells, and cochlear hair cells . Interestingly, the functional consequences differed substantially between the latter two systems: while retinal neurotransmission was majorly impaired due to the mislocalization of presynaptic Ca 2+ channels, loss of membrane‐proximal SVs, and subsequently strongly reduced EPSC amplitudes , the cochlear phenotype of these mice was surprisingly mild.…”

“…This structural reorganization in the mutants led to a minor exocytosis deficit in response to mild depolarizations that was lost upon stronger stimulation. However, single unit recordings from auditory nerve fibers revealed that, while firing thresholds were increased, peak as well as adapted firing rates were significantly reduced . Moreover, the temporal jitter of firing and the time course of adaptation were increased and the recovery from adaptation was slowed.…”

“…Interestingly, the functional consequences differed substantially between the latter two systems: while retinal neurotransmission was majorly impaired due to the mislocalization of presynaptic Ca 2+ channels, loss of membrane‐proximal SVs, and subsequently strongly reduced EPSC amplitudes , the cochlear phenotype of these mice was surprisingly mild. Here, two independent studies thoroughly characterized the presynaptic morphology and function of cochlear IHCs. Apart from unchanged synapse counts, Jean and colleagues reported extensive presynaptic developmental compensation that produced multiple ‘ribbonless’ AZs at each individual synaptic contact, thereby maintaining or even increasing the presynaptic complement of releasable SVs .…”

“…Here, two independent studies thoroughly characterized the presynaptic morphology and function of cochlear IHCs. Apart from unchanged synapse counts, Jean and colleagues reported extensive presynaptic developmental compensation that produced multiple ‘ribbonless’ AZs at each individual synaptic contact, thereby maintaining or even increasing the presynaptic complement of releasable SVs . Additionally, both studies consistently reported an increased size of postsynaptic densities and GluA3 receptor clusters , indicative of extensive pre‐ and postsynaptic homeostatic plasticity mechanisms aiming to compensate the loss of presynaptic release capacity.…”

“…Consistent with the kiss‐and‐run hypothesis and based on the results from mathematical modeling of these events, one study argued that these bizarre, multiphasic EPSCs, which are usually characterized by a lower amplitude, but similar charge to that of simple EPSCs, might stem from fusion‐pore‐flickering of a transiently fusing single SV . Although compelling, this is difficult to prove experimentally: SVs of cochlear IHCs are small in size, measuring ~ 30–50 nm in diameter, depending on the fixation method , which equates to approximately 45 aF in membrane capacitance . Monitoring the flickering of such small SVs requires cell‐attached patch‐clamp recordings, amperometry or in vivo fluorescent imaging of single SVs, which are all technically extremely challenging methods – in particular when applied within a tightly‐structured and dense tissue such as the organ of Corti.…”