Biology subjects

Kusch, K.

Publications and source records attributed to Kusch, K..

10 recordsLinked to original sources

bioRxiv2026

Multichannel optical cochlear implants enable spectrally distinct auditory activity

When hearing fails, cochlear implants (CIs) restore auditory perception. Yet, coding of spectral information remains a bottleneck because each electrode broadly activates the auditory nerve. As light can be more conveniently confined, optical (o)CIs present an alternative. Here, we combined expression of the potent channelrhodopsin ChReef in spiral ganglion neurons (SGNs) with oCIs comprising 5-10 green LEDs in gerbils. This combination enabled systematic comparison of encoding intensity and spectral information by individual oCI channels to acoustic and electrical stimulation within a translationally feasible energy range. Recordings from the inferior colliculus (ICC) showed that ChReef aligned SGN light sensitivity with LED radiant fluxes: ICC activity had thresholds <200 nJ and reached a maximum equivalent to that achieved with 51 dB SPL pure tones. Multichannel oCIs enabled tonotopically ordered, spectrally distinct stimulation that more closely resembled acoustic stimulation than did electrical stimulation. Some LEDs elicited multiple spectral peaks at higher intensities. Linear discriminant analysis of ICC activity indicated improved channel discriminability for optical over electrical stimulation. In summary, ,micro-joule oCI-stimulation achieves improved spectral resolution over state-of-the-art electrical stimulation.

neuroscience↗

bioRxiv2025

CochleaNet: deep learning-based image analysis for cochlear connectomics and gene therapy

With the emergence of gene and optogenetic therapies targeting deafness, the comprehensive analysis of the molecular anatomy and physiology of the cochlea has become ever more important. Here, we introduce CochleaNet, a deep learning-based framework to analyze volumetric imaging data obtained by light-sheet microscopy of decalcified, cleared and fluorescently labeled cochleae. CochleaNet covers the workflow from reconstruction of the cochlea to segmentation of inner hair cells, spiral ganglion neurons and their afferent synapses, to analyzing the expression of gene therapy products. We validated CochleaNet by comparison to manual image analysis. Trained on high isotropic resolution mouse data, CochleaNet was also applicable to the cochlea of the gerbil, another relevant animal model, and lower-resolution mouse data from a commercially available microscope. We conclude that the combination of light-sheet microscopy and image analysis with CochleaNet paves the way for rapid and reliable quantification of cochlear molecular anatomy and preclinical gene therapy outcomes.

neuroscience↗

bioRxiv2025

Paralemmin-3 sustains the integrity of the lateral plasma membrane and subsurface cisternae of auditory hair cells

In the mammalian inner ear, cochlear inner hair cells (IHCs) enable accurate and faithful synaptic sound encoding, while outer hair cells (OHCs) perform frequency-specific sound amplification and fine-tuning through their intrinsic voltage-dependent somatic electromotility. This latter process is facilitated by the unique trilaminate structure of the OHC lateral wall, which consists of the plasma membrane that is densely occupied by the transmembrane motor protein Prestin, the submembrane actin- and spectrin-based cytoskeleton, and the endomembranous subsurface cisternae. This complex system provides mechanical resilience while allowing for cell expansion and contraction during electromotility. Whereas the ultrastructure of the lateral wall is well described, its molecular architecture remains largely elusive. Here, we identified Paralemmin-3 (Palm3) as a novel protein specifically localized to the lateral walls of auditory HCs to play a crucial role in connecting the plasma membrane to the underlying cytoskeleton and subsurface cisternae. Palm3-KO mice display early-onset and progressive hearing impairment that results from diminished cochlear amplification. Subsequent multiscale morphological analyses revealed structural collapse of OHCs that led to progressive and extensive OHC loss along the tonotopic axis. Palm3-KO OHCs exhibited disrupted expression and distribution of several membrane-associated proteins - including spectrin isoforms and Prestin - suggesting an essential role of Palm3 in plasma membrane scaffolding. Electron tomography of OHC lateral walls revealed significantly fewer and structurally perturbed subsurface cisternae. Finally, adeno-associated virus (AAV)-mediated rescue of Palm3 during early postnatal development partly restored hearing function, enhanced OHC survival, and restored OHC cell shape as well as membrane protein expression levels. In summary, Palm3 is a key component of the submembrane cytoskeleton in cochlear hair cells, playing a fundamental role in hair cell biology and hearing, and emerges as an attractive candidate for the long-elusive "pillar" component of the hair cell lateral wall ultrastructure.

cell biology↗

bioRxiv2025

A correlative workflow streamlines synaptic imaging by cryo-electron tomography

Despite decades of intense research, the molecular organization of the synapse is not well understood. To address this issue, we sought to develop a method for systematic imaging of synapses by cryo-electron tomography (cryo-ET), a technology capable of mapping cellular architecture at molecular resolution. Thinning of cellular samples by cryo-focused ion beam milling is a prerequisite for high-quality cryo-ET imaging, but this process needs to be guided to the structures of interest. To allow robust synaptic targeting, we established a correlative cryo-light/electron microscopy approach by which synapses are fluorescently labeled in a minimally invasive manner, using a synthetic binder of the postsynaptic scaffold PSD-95 and antibodies against the presynaptic protein Synaptotagmin-1. Cryo-ET imaging at sites of colocalization consistently revealed excitatory synapses. Our method allows structural studies of synaptic protein complexes in situ, facilitating investigations of the molecular architecture of synapses.

neuroscience↗

bioRxiv2024

Improved optogenetic modification of the spiral ganglion neurons for future optical cochlear implants

Optogenetic stimulation has become a promising approach for restoring lost body function. For example, partial restoration of vision has been achieved in a blind patient and proof-of-concept has been demonstrated for optogenetic hearing restoration in rodents. In order to prepare clinical translation of hearing restoration, efficient and safe optogenetic modification of spiral ganglion neurons (SGNs) in the mature cochlea remains to be developed. Here, we established microcatheter-based administration adeno-associated virus (AAV) to scala tympani of the cochlea of Mongolian gerbils and compared it to the previously developed AAV-injection into the spiral ganglion. We probed the potential AAV-PHP.S capsid to express channelrhodopsins (ChRs) under the control of the human synapsin promotor in mature SGNs in hearing and deafened gerbils. Using the microcatheter approach, but not with the AAV-modiolus injection, we achieved reliable ChR expression in SGN enabling optogenetic stimulation of the auditory pathway in 80% of the treated animals. Yet, the efficiency of SGN transduction was modest with only [~]30% ChR-expressing SGNs. Moreover, we encountered off-target expression in hair cells in hearing gerbils in both approaches, but not ChR expression in the central nervous system using microcatheter administration. Comparing optogenetic auditory brainstem responses of gerbils with and without hair cell transduction confirmed that SGNs were the primary site of optogenetic stimulation of the pathway.

neuroscience↗

bioRxiv2023

Efficient and sustained optogenetic control of nervous and cardiac systems

Optogenetic control is used to manipulate the activity of specific cell types in vivo for a variety of biological and clinical applications. Here we report ChReef, an improved variant of the channelrhodopsin ChRmine. ChReef offers minimal photocurrent desensitization, a unitary conductance of 80 fS and closing kinetics of 30 ms, which together enable reliable optogenetic control of cells at low light levels with good temporal fidelity and sustained stimulation. We demonstrate efficient and reliable red-light pacing and depolarization-block of ChReef-expressing cardiomyocyte clusters. We used AAV-based gene transfer to express ChReef in retinal ganglion cells, where it restores visual function in blind mice with light sources as weak as an iPad screen. Toward optogenetic hearing restoration, ChReef enables stimulation of the auditory pathway in rodents and non-human primates with nano-Joule threshold, enabling efficient and frequency-specific stimulation by LED-based optical cochlear implants.

biophysics↗

bioRxiv2023

CaBP1 and 2 enable sustained CaV1.3 calcium currents and synaptic transmission in inner hair cells

To encode continuous sound stimuli, the inner hair cell (IHC) ribbon synapses utilize calcium-binding proteins (CaBPs), which reduce the inactivation of their CaV1.3 calcium channels. Mutations in the CABP2 gene underlie non-syndromic autosomal recessive hearing loss DFNB93. Besides CaBP2, the structurally related CaBP1 is highly abundant in the IHCs. Here, we investigated how the two CaBPs cooperatively regulate IHC synaptic function. In Cabp1/2 double-knockout mice, we find strongly enhanced CaV1.3 inactivation, slowed recovery from inactivation and impaired sustained exocytosis. Already mild IHC activation further reduces the availability of channels to trigger synaptic transmission and may effectively silence synapses. Spontaneous and sound-evoked responses of spiral ganglion neurons in vivo are strikingly reduced and strongly depend on stimulation rates. Transgenic expression of CaBP2 leads to substantial recovery of IHC synaptic function and hearing sensitivity. We conclude that CaBP1 and 2 act together to suppress voltage and calcium-dependent inactivation of IHC CaV1.3 channels in order to support sufficient rate of exocytosis and enable fast, temporally precise and indefatigable sound encoding.

neuroscience↗

bioRxiv2022

Myelin lipids as nervous system energy reserves

Neuronal functions and impulse propagation depend on the continuous supply of glucose1,2. Surprisingly, the mammalian brain has no obvious energy stores, except for astroglial glycogen granules3. Oligodendrocytes make myelin for rapid axonal impulse conduction4 and also support axons metabolically with lactate5-7. Here, we show that myelin itself, a lipid-rich membrane compartment, becomes a local energy reserve when glucose is lacking. In the mouse optic nerve, a model white matter tract, oligodendrocytes survive glucose deprivation far better than astrocytes, by utilizing myelin lipids which requires oxygen and fatty acid beta-oxidation. Importantly, fatty acid oxidation also contributes to axonal ATP and basic conductivity. This metabolic support by fatty acids is an oligodendrocyte function, involving mitochondria and myelin-associated peroxisomes, as shown with mice lacking Mfp2. To study reduced glucose availability in vivo without physically starving mice, we deleted the Slc2a1 gene from mature oligodendrocytes. This caused a significant decline of the glucose transporter GLUT1 from the myelin compartment leading to myelin sheath thinning. We suggest a model in which myelin turnover under low glucose conditions can transiently buffer axonal energy metabolism. This model may explain the gradual loss of myelin in a range of neurodegenerative diseases8 with underlying hypometabolism9.

neuroscience↗

bioRxiv2020

White matter integrity requires continuous myelin synthesis at the inner tongue

Myelin, the electrically insulating axonal sheath, is composed of lipids and proteins with exceptionally long lifetime. This raises the question how myelin function is affected by myelin turnover. We have studied the integrity of myelinated tracts after experimentally preventing the formation of new myelin in the CNS of adult mice, using an inducible Mbp null allele. Oligodendrocytes survived recombination, continued expressing myelin genes, but failed to maintain compacted myelin sheaths. Using 3D electron microscopy and mass spectrometry imaging we visualized myelin-like membranes that failed to incorporate adaxonally, most prominently at juxta-paranodes. Myelinoid body formation indicated degradation of existing myelin at the abaxonal side and at the inner tongue of the sheath. Compacted myelin thinning and shortening of internodes, with about 50% myelin lost after 20 weeks (=5 months), ultimately led to axonal pathology and neurological disease. These data reveal that functional axon-myelin units require the continuous incorporation of new myelin membranes.

neuroscience↗

bioRxiv2019

A role of oligodendrocytes in information processing independent of conduction velocity

Myelinating oligodendrocytes enable fast impulse propagation along axons as revealed through studies of homogeneously myelinated white matter tracts. However, gray matter myelination patterns are different, with sparsely myelinated sections leaving large portions of the axons naked. The consequences of this patchy myelination for oligodendrocyte function are not understood but suggest other roles in information processing beyond the regulation of axonal conduction velocity. Here, we analyzed the contribution of myelin to auditory information processing using paradigms that are good predictors of speech understanding in humans. We compared mice with different degrees of dysmyelination using acute cortical multiunit recordings in combination with behavioral readouts. We identified complex alterations of neuronal responses that reflect fatigue and temporal acuity deficits. Partially discriminable but overall similar deficits were observed in mice with oligodendrocytes that can myelinate but cannot fully support axons metabolically. Thus, myelination contributes to sustained stimulus perception in temporally complex paradigms, revealing a role of oligodendrocytes in the CNS beyond the increase of axonal conduction velocity.

neuroscience↗