patch clamp recordings

COCHLEAR SIGNALING MEDIATED BY HENSEN’S CELLS

PROJECT SUMMARY/ABSTRACT The organ of Corti has two types of auditory sensory cells (inner and outer hair cells) surrounded by nearly a dozen different types of supporting cells organized in a very meticulous pattern. Hair cells mediate the mechano-electrical transduction process of the organ of Corti and thus most cochlear auditory research has focused on these sensory cells. In contrast, much less is known about the different types of cochlear supporting cells, even though they likely impact hair cell function. Hensen’s cells are located laterally to the outer hair cell rows and appear to be the only cell type in the cochlear epithelium that expresses TRPA1 channels. These channels are widely known for their role as sensors of tissue damage and inflammation in nociceptive neurons. Not surprisingly, we recently found that Hensen’s cells are main sensors of tissue damage in the cochlear epithelium via the activation of TRPA1 channels (Velez-Ortega et al., Nat Commn, 2023). Additionally, our preliminary data also supports the role of Hensen’s cells in signaling pathways important for the proper innervation of the organ of Corti (aim 1), for the transmission of cochlear damage signals to the brain (aim 2), and for the regulation of hearing sensitivity after acoustic trauma (aim 3). Thus, here we will explore the hypothesis that TRPA1- mediated signaling pathways in the Hensen’s cells are required for the proper innervation and auditory function of the organ of Corti. In Aim 1 we will perform a detailed comparison of the morphology and synapses of afferent cochlear neurons of wild-type and Trpa1-/- mice at several developmental stages (using immunolabeling, confocal microscopy, STED microscopy, and electron microscopy) to assess the role of TRPA1 activity on the postnatal refinement of the cochlear innervation. Aim 2 will evaluate whether the afferent type II spiral ganglion neurons (SGN) can be activated downstream of TRPA1 channel gating in Hensen’s cells by testing responses of neonate and adult type II SGN to TRPA1 agonists (via live-cell time-lapse calcium imaging and patch clamp recordings of type II SGN dendrites). Aim 3 will test the impact of TRPA1 signaling in Hensen’s cells to the operating point of the cochlear transducer (via the recording of cochlear microphonics) and to cochlear tuning (via the recording of ABR tuning curves). This study is significant because it will contribute to our understanding of the cellular (Hensen’s cells plus type II SGN) and molecular (TRPA1 channels) mechanisms of the elusive cochlear nociceptive pathway. In addition, given that the loss of TRPA1 channels does not affect hearing thresholds in mice, we believe that undiagnosed deficits in TRPA1-dependent responses in the human population could represent a hidden susceptibility for cochlear damage after noise exposure or other insults.

NMC4 Short Talk: Resilience through diversity: Loss of neuronal heterogeneity in epileptogenic human tissue impairs network resilience to sudden changes in synchrony

A myriad of pathological changes associated with epilepsy, including the loss of specific cell types, improper expression of individual ion channels, and synaptic sprouting, can be recast as decreases in cell and circuit heterogeneity. In recent experimental work, we demonstrated that biophysical diversity is a key characteristic of human cortical pyramidal cells, and past theoretical work has shown that neuronal heterogeneity improves a neural circuit’s ability to encode information. Viewed alongside the fact that seizure is an information-poor brain state, these findings motivate the hypothesis that epileptogenesis can be recontextualized as a process where reduction in cellular heterogeneity renders neural circuits less resilient to seizure onset. By comparing whole-cell patch clamp recordings from layer 5 (L5) human cortical pyramidal neurons from epileptogenic and non-epileptogenic tissue, we present the first direct experimental evidence that a significant reduction in neural heterogeneity accompanies epilepsy. We directly implement experimentally-obtained heterogeneity levels in cortical excitatory-inhibitory (E-I) stochastic spiking network models. Low heterogeneity networks display unique dynamics typified by a sudden transition into a hyper-active and synchronous state paralleling ictogenesis. Mean-field analysis reveals a distinct mathematical structure in these networks distinguished by multi-stability. Furthermore, the mathematically characterized linearizing effect of heterogeneity on input-output response functions explains the counter-intuitive experimentally observed reduction in single-cell excitability in epileptogenic neurons. This joint experimental, computational, and mathematical study showcases that decreased neuronal heterogeneity exists in epileptogenic human cortical tissue, that this difference yields dynamical changes in neural networks paralleling ictogenesis, and that there is a fundamental explanation for these dynamics based in mathematically characterized effects of heterogeneity. These interdisciplinary results provide convincing evidence that biophysical diversity imbues neural circuits with resilience to seizure and a new lens through which to view epilepsy, the most common serious neurological disorder in the world, that could reveal new targets for clinical treatment.

Neocortex saves energy by reducing coding precision during food scarcity

Information processing is energetically expensive. In the mammalian brain, it is unclear how information coding and energy usage are regulated during food scarcity. We addressed this in the visual cortex of awake mice using whole-cell patch clamp recordings and two-photon imaging to monitor layer 2/3 neuronal activity and ATP usage. We found that food restriction resulted in energy savings through a decrease in AMPA receptor conductance, reducing synaptic ATP usage by 29%. Neuronal excitability was nonetheless preserved by a compensatory increase in input resistance and a depolarized resting membrane potential. Consequently, neurons spiked at similar rates as controls, but spent less ATP on underlying excitatory currents. This energy-saving strategy had a cost since it amplified the variability of visually-evoked subthreshold responses, leading to a 32% broadening in orientation tuning and impaired fine visual discrimination. These findings reveal novel mechanisms that dynamically regulate energy usage and coding precision in neocortex.

Nicotine effects on anxiety behavior and brain biochemical markers in adult male mice and in rat hippocampal slices: patch clamp recordings

Novel method for reliably measuring miniature and spontaneous postsynaptic potentials/currents in whole-cell patch clamp recordings