Spontaneous and Acetylcholine Evoked Calcium Transients in the Developing Mouse Utricle

Spontaneous calcium transients are present during early postnatal development in the mouse retina and cochlea, and play an important role in maturation of the sensory organs and neural circuits in the central nervous system (CNS). It is not known whether similar calcium transients occur during postnatal development in the vestibular sensory organs. Here we demonstrate spontaneous intracellular calcium transients in sensory hair cells (HCs) and supporting cells (SCs) in the murine utricular macula during the first two postnatal weeks. Calcium transients were monitored using a genetically encoded calcium indicator, GCaMP5G (G5), at 100 ms-frame−1 in excised utricle sensory epithelia, including HCs, SCs, and neurons. The reporter line expressed G5 and tdTomato (tdT) in a Gad2-Cre dependent manner within a subset of utricular HCs, SCs and neurons. Kinetics of the G5 reporter limited temporal resolution to calcium events lasting longer than 200 ms. Spontaneous calcium transients lasting 1-2 s were observed in the expressing population of HCs at birth and slower spontaneous transients lasting 10-30 s appeared in SCs by P3. Beginning at P5, calcium transients could be modulated by application of the efferent neurotransmitter acetylcholine (ACh). In mature mice, calcium transients in the utricular macula occurred spontaneously, had a duration 1-2 s, and could be modulated by the exogenous application of acetylcholine (ACh) or muscarine. Long-lasting calcium transients evoked by ACh in mature mice were blocked by atropine, consistent with previous reports describing the role of muscarinic receptors expressed in calyx bearing afferents in efferent control of vestibular sensation. Large spontaneous and ACh evoked transients were reversibly blocked by the inositol trisphosphate receptor (IP3R) antagonist aminoethoxydiphenyl borate (2-APB). Results demonstrate long-lasting calcium transients are present in the utricular macula during the first postnatal week, and that responses to ACh mature over this same time period

Ionic composition of endolymph and perilymph in the inner ear of the oyster toadfish, Opsanus tau

Author Posting. © Marine Biological Laboratory, 2008. This article is posted here by permission of Marine Biological Laboratory for personal use, not for redistribution. The definitive version was published in Biological Bulletin 214 (2008): 83-90.The concentrations of free Na+, K+, Ca2+, and Cl-in endolymph and perilymph from the inner ear of the oyster toadfish, Opsanus tau, were measured in vivo using double-barreled ion-selective electrodes. Perilymph concentrations were similar to those measured in other species, while endolymph concentrations were similar to those measured previously in elasmobranch fish, though significantly different from concentrations reported in mammals. Perilymph concentrations (mean ± std. dev.) were as follows: Na+, 129 mmol l-1 ± 20; K+, 4.96 mmol l-1 ± 2.67; Ca2+, 1.83 mmol l-1 ± 0.27; and Cl-, 171 mmol l-1 ± 20. Saccular endolymph concentrations were Na+, 166 mmol l-1 ± 22; K+, 51.4 mmol l-1 ± 16.7; Ca2+, 2.88 mmol l-1 ± 0.27; and Cl-, 170 mmol l-1 ± 12; and semicircular canal (utricular vestibule) endolymph concentrations were Na+, 122 mmol l-1 ± 15; K+, 47.7 mmol l-1 ± 13.2; Ca2+, 1.78 mmol l-1 ± 0.48; Cl-, 176 mmol l-1 ± 27. The relatively high concentrations of Ca2+ and Na+ in the endolymph may have significant implications for the physiological function of the mechanoelectrical transduction channels in the vestibular hair cells of fish compared to those of their mammalian counterparts.This work was supported by the National Institute of Deafness and Other Communications Disorders P01 DC01837, R01 DC06685, R01 DC04928, NASA NNA-04CK67H, and NSF Igert DGE9987616

A Mathematical Derivative Performed by Convergence of GABA and Glutamate in the Vestibular Periphery

Closed-loop neuromuscular control systems rely upon the nervous system to generate, in real time, motor outputs in response to sensory inputs. This typically requires the central nervous system to perform calculations analogous to the fundamental operations of mathematical calculus. A classic example of this is the vestibular ocular reflex (VOR), in which angular-velocity encoding afferent signals are input to the brainstem, mathematically integrated by the central nervous system in real time, and output by motor neurons to control the angular position of the eyes. Mathematical differentiation by the nervous system has received less attention than integration, but differentiation is just as fundamental and perhaps much more ubiquitous. For example, step stimuli often evoke in single neurons an initial increase in firing rate followed by a period of adaptation. The initial increase in discharge rate and subsequent adaptation can be approximated in some cases by a fractional mathematical derivative. For the most rapidly adapting neurons, the fractional exponent approaches one and the neural response approaches a Dirac delta function. A mathematical derivative similar to the fractional model is present in the vestibular periphery, and is evidenced by responses of first-order semicircular canal afferent neurons that respond with discharge rates proportional to the rate of change of sensory hair cell intracellular voltag

A mathematical model for mechanical activation and compound action potential generation by the utricle in response to sound and vibration

IntroductionCalyx bearing vestibular afferent neurons innervating type I hair cells in the striolar region of the utricle are exquisitely sensitive to auditory-frequency air conducted sound (ACS) and bone conducted vibration (BCV). Here, we present experimental data and a mathematical model of utricular mechanics and vestibular compound action potential generation (vCAP) in response to clinically relevant levels of ACS and BCV. Vibration of the otoconial layer relative to the sensory epithelium was simulated using a Newtonian two-degree-of-freedom spring-mass-damper system, action potential timing was simulated using an empirical model, and vCAPs were simulated by convolving responses of the population of sensitive neurons with an empirical extracellular voltage kernel. The model was validated by comparison to macular vibration and vCAPs recorded in the guinea pig, in vivo.ResultsTransient stimuli evoked short-latency vCAPs that scaled in magnitude and timing with hair bundle mechanical shear rate for both ACS and BCV. For pulse BCV stimuli with durations <0.8 ms, the vCAP magnitude increased in proportion to temporal bone acceleration, but for pulse durations >0.9 ms the magnitude increased in proportion to temporal bone jerk. Once validated using ACS and BCV data, the model was applied to predict blast-induced hair bundle shear, with results predicting acute mechanical damage to bundles immediately upon exposure.DiscussionResults demonstrate the switch from linear acceleration to linear jerk as the adequate stimulus arises entirely from mechanical factors controlling the dynamics of sensory hair bundle deflection. The model describes the switch in terms of the mechanical natural frequencies of vibration, which vary between species based on morphology and mechanical factors

Cooperativity of Kv7.4 channels confers ultrafast electromechanical sensitivity and emergent properties in cochlear outer hair cells.

The mammalian cochlea relies on active electromotility of outer hair cells (OHCs) to resolve sound frequencies. OHCs use ionic channels and somatic electromotility to achieve the process. It is unclear, though, how the kinetics of voltage-gated ionic channels operate to overcome extrinsic viscous drag on OHCs at high frequency. Here, we report ultrafast electromechanical gating of clustered Kv7.4 in OHCs. Increases in kinetics and sensitivity resulting from cooperativity among clustered-Kv7.4 were revealed, using optogenetics strategies. Upon clustering, the half-activation voltage shifted negative, and the speed of activation increased relative to solitary channels. Clustering also rendered Kv7.4 channels mechanically sensitive, confirmed in consolidated Kv7.4 channels at the base of OHCs. Kv7.4 clusters provide OHCs with ultrafast electromechanical channel gating, varying in magnitude and speed along the cochlea axis. Ultrafast Kv7.4 gating provides OHCs with a feedback mechanism that enables the cochlea to overcome viscous drag and resolve sounds at auditory frequencies

Dynamic displacement of normal and detached semicircular canal cupula

© 2009 The Authors. This is an open-access article distributed under the terms of the Creative Commons Attribution Noncommercial License. The definitive version was published in JARO - Journal of the Association for Research in Otolaryngology 10 (2009): 497-509, doi:10.1007/s10162-009-0174-y.The dynamic displacement of the semicircular canal cupula and modulation of afferent nerve discharge were measured simultaneously in response to physiological stimuli in vivo. The adaptation time constant(s) of normal cupulae in response to step stimuli averaged 36 s, corresponding to a mechanical lower corner frequency for sinusoidal stimuli of 0.0044 Hz. For stimuli equivalent to 40–200 deg/s of angular head velocity, the displacement gain of the central region of the cupula averaged 53 nm per deg/s. Afferents adapted more rapidly than the cupula, demonstrating the presence of a relaxation process that contributes significantly to the neural representation of angular head motions by the discharge patterns of canal afferent neurons. We also investigated changes in time constants of the cupula and afferents following detachment of the cupula at its apex—mechanical detachment that occurs in response to excessive transcupular endolymph pressure. Detached cupulae exhibited sharply reduced adaptation time constants (300 ms–3 s, n = 3) and can be explained by endolymph flowing rapidly over the apex of the cupula. Partially detached cupulae reattached and normal afferent discharge patterns were recovered 5–7 h following detachment. This regeneration process may have relevance to the recovery of semicircular canal function following head trauma.Financial support was provided by the NIDCD R01 DC06685 (Rabbitt) and NASA GSRP 56000135 & NSF IGERT DGE- 9987616 (Breneman)

Monitoring Voltage-Dependent Charge Displacement of Shaker B-IR K+ Ion Channels Using Radio Frequency Interrogation

Here we introduce a new technique that probes voltage-dependent charge displacements of excitable membrane-bound proteins using extracellularly applied radio frequency (RF, 500 kHz) electric fields. Xenopus oocytes were used as a model cell for these experiments, and were injected with cRNA encoding Shaker B-IR (ShB-IR) K+ ion channels to express large densities of this protein in the oocyte membranes. Two-electrode voltage clamp (TEVC) was applied to command whole-cell membrane potential and to measure channel-dependent membrane currents. Simultaneously, RF electric fields were applied to perturb the membrane potential about the TEVC level and to measure voltage-dependent RF displacement currents. ShB-IR expressing oocytes showed significantly larger changes in RF displacement currents upon membrane depolarization than control oocytes. Voltage-dependent changes in RF displacement currents further increased in ShB-IR expressing oocytes after ∼120 µM Cu2+ addition to the external bath. Cu2+ is known to bind to the ShB-IR ion channel and inhibit Shaker K+ conductance, indicating that changes in the RF displacement current reported here were associated with RF vibration of the Cu2+-linked mobile domain of the ShB-IR protein. Results demonstrate the use of extracellular RF electrodes to interrogate voltage-dependent movement of charged mobile protein domains — capabilities that might enable detection of small changes in charge distribution associated with integral membrane protein conformation and/or drug–protein interactions

Hair Cell Bundles: Flexoelectric Motors of the Inner Ear

Microvilli (stereocilia) projecting from the apex of hair cells in the inner ear are actively motile structures that feed energy into the vibration of the inner ear and enhance sensitivity to sound. The biophysical mechanism underlying the hair bundle motor is unknown. In this study, we examined a membrane flexoelectric origin for active movements in stereocilia and conclude that it is likely to be an important contributor to mechanical power output by hair bundles. We formulated a realistic biophysical model of stereocilia incorporating stereocilia dimensions, the known flexoelectric coefficient of lipid membranes, mechanical compliance, and fluid drag. Electrical power enters the stereocilia through displacement sensitive ion channels and, due to the small diameter of stereocilia, is converted to useful mechanical power output by flexoelectricity. This motor augments molecular motors associated with the mechanosensitive apparatus itself that have been described previously. The model reveals stereocilia to be highly efficient and fast flexoelectric motors that capture the energy in the extracellular electro-chemical potential of the inner ear to generate mechanical power output. The power analysis provides an explanation for the correlation between stereocilia height and the tonotopic organization of hearing organs. Further, results suggest that flexoelectricity may be essential to the exquisite sensitivity and frequency selectivity of non-mammalian hearing organs at high auditory frequencies, and may contribute to the “cochlear amplifier” in mammals

Conscious perception of errors and its relation to the anterior insula

To detect erroneous action outcomes is necessary for flexible adjustments and therefore a prerequisite of adaptive, goal-directed behavior. While performance monitoring has been studied intensively over two decades and a vast amount of knowledge on its functional neuroanatomy has been gathered, much less is known about conscious error perception, often referred to as error awareness. Here, we review and discuss the conditions under which error awareness occurs, its neural correlates and underlying functional neuroanatomy. We focus specifically on the anterior insula, which has been shown to be (a) reliably activated during performance monitoring and (b) modulated by error awareness. Anterior insular activity appears to be closely related to autonomic responses associated with consciously perceived errors, although the causality and directions of these relationships still needs to be unraveled. We discuss the role of the anterior insula in generating versus perceiving autonomic responses and as a key player in balancing effortful task-related and resting-state activity. We suggest that errors elicit reactions highly reminiscent of an orienting response and may thus induce the autonomic arousal needed to recruit the required mental and physical resources. We discuss the role of norepinephrine activity in eliciting sufficiently strong central and autonomic nervous responses enabling the necessary adaptation as well as conscious error perception

Power efficiency of outer hair cell somatic electromotility

© 2009 Rabbitt et al. This article is distributed under the terms of the Creative Commons Attribution License. The definitive version was published in PLoS Computational Biology 5 (2009): e1000444, doi:10.1371/journal.pcbi.1000444.Cochlear outer hair cells (OHCs) are fast biological motors that serve to enhance the vibration of the organ of Corti and increase the sensitivity of the inner ear to sound. Exactly how OHCs produce useful mechanical power at auditory frequencies, given their intrinsic biophysical properties, has been a subject of considerable debate. To address this we formulated a mathematical model of the OHC based on first principles and analyzed the power conversion efficiency in the frequency domain. The model includes a mixture-composite constitutive model of the active lateral wall and spatially distributed electro-mechanical fields. The analysis predicts that: 1) the peak power efficiency is likely to be tuned to a specific frequency, dependent upon OHC length, and this tuning may contribute to the place principle and frequency selectivity in the cochlea; 2) the OHC power output can be detuned and attenuated by increasing the basal conductance of the cell, a parameter likely controlled by the brain via the efferent system; and 3) power output efficiency is limited by mechanical properties of the load, thus suggesting that impedance of the organ of Corti may be matched regionally to the OHC. The high power efficiency, tuning, and efferent control of outer hair cells are the direct result of biophysical properties of the cells, thus providing the physical basis for the remarkable sensitivity and selectivity of hearing.This work was supported by NIDCD R01 DC04928 (Rabbitt), NIDCD R01 DC00384 (Brownell) and NASA Ames GSRA56000135 (Breneman)