Albert Kao

Magnetic nanoparticles for ultrafast mechanical control of inner ear hair cells.

We introduce cubic magnetic nanoparticles as an effective tool for precise and ultrafast control of mechanosensitive cells. The temporal resolution of our system is ∼1000 times faster than previously used magnetic switches and is comparable to the current state-of-the-art optogenetic tools. The use of a magnetism-gated switch reported here can address the key challenges of studying mechanotransduction in biological systems. The cube-shaped magnetic nanoparticles are designed to bind to components of cellular membranes and can be controlled with an electromagnet to exert pico-Newtons of mechanical force on the cells. The cubic nanoparticles can thus be used for noncontact mechanical control of the position of the stereocilia of an inner ear hair cell, yielding displacements of tens of nanometers, with sub-millisecond temporal resolution. We also prove that such mechanical stimulus leads to the influx of ions into the hair cell. Our study demonstrates that a magnetic switch can yield ultrafast temporal resolution, and has capabilities for remote manipulation and biological specificity, and that such magnetic system can be used for the study of mechanotransduction processes of a wide range of sensory systems.

Multiple-Timescale Dynamics Underlying Spontaneous Oscillations of Saccular Hair Bundles

Spontaneous oscillations displayed by hair bundles of the bullfrog sacculus have complex temporal profiles, not fully captured by single limit-cycle descriptions. Quiescent intervals are typically interspersed with oscillations, leading to a bursting-type behavior. Temporal characteristics of the oscillation are strongly affected by imposing a mechanical load or by the application of a steady-state deflection to the resting position of the bundle. Separate spectral components of the spontaneous motility are differently affected by increases in the external calcium concentration. We use numerical modeling to explore the effects of internal parameters on the oscillatory profiles, and to reproduce the experimental modulation induced by mechanical or ionic manipulation.

Dynamic state and evoked motility in coupled hair bundles of the bullfrog sacculus.

Spontaneous oscillations, one of the signatures of the active process in non-mammalian hair cells, have been shown to occur in individual hair bundles that have been fully decoupled from the overlying membrane. Here we use semi-intact preparations of the bullfrog sacculus to demonstrate that under more natural loading conditions, innate oscillations are suppressed by the presence of the overlying otolithic membrane, indicating that hair bundles lie in the quiescent rather than the unstable regime. Transepithelial electrical stimulation was then used to test the effect of evoking entrained hair bundle movement with an external stimulus. Firstly, we used a preparation in which the otolithic membrane has been partially detached, coupling only hair bundles of comparable orientations. Secondly, we deposited artificial polymer membranes on top of the epithelium so as to connect to only 10-20 cells. In both of these systems, hair bundle motion phase-locked by the electrical signal was found to induce movement in the overlying structures.

Low Frequency Entrainment of Oscillatory Bursts in Hair Cells

Sensitivity of mechanical detection by the inner ear is dependent upon a highly nonlinear response to the applied stimulus. Here we show that a system of differential equations that support a subcritical Hopf bifurcation, with a feedback mechanism that tunes an internal control parameter, captures a wide range of experimental results. The proposed model reproduces the regime in which spontaneous hair bundle oscillations are bistable, with sporadic transitions between the oscillatory and the quiescent state. Furthermore, it is shown, both experimentally and theoretically, that the application of a high-amplitude stimulus to the bistable system can temporarily render it quiescent before recovery of the limit cycle oscillations. Finally, we demonstrate that the application of low-amplitude stimuli can entrain bundle motility either by mode-locking to the spontaneous oscillation or by mode-locking the transition between the quiescent and oscillatory states.

Mechanical Overstimulation of Hair Bundles: Suppression and Recovery of Active Motility

We explore the effects of high-amplitude mechanical stimuli on hair bundles of the bullfrog sacculus. Under in vitro conditions, these bundles exhibit spontaneous limit cycle oscillations. Prolonged deflection exerted two effects. First, it induced an offset in the position of the bundle. Recovery to the original position displayed two distinct time scales, suggesting the existence of two adaptive mechanisms. Second, the stimulus suppressed spontaneous oscillations, indicating a change in the hair bundle’s dynamic state. After cessation of the stimulus, active bundle motility recovered with time. Both effects were dependent on the duration of the imposed stimulus. External calcium concentration also affected the recovery to the oscillatory state. Our results indicate that both offset in the bundle position and calcium concentration control the dynamic state of the bundle.

Self-Tuning of Hair Cells in the Bullfrog Sacculus

Spontaneous oscillations of the stereociliary bundle of a hair cell - the mechanosensory cell in auditory and vestibular systems - is considered to be a signature of an active amplification mechanism. We study whether an internal self-tuning process governs the active motility, by mimicking the effects of loud sound on the spontaneous oscillation. After the application of high-amplitude stimuli, with deflections on the order of micrometers applied to the hair bundle, the active oscillatory motion of the hair bundle was suppressed for hundreds of milliseconds, indicating a change in the dynamic state of the hair cell. Here we observe the recovery profile of an oscillating hair bundle after cessation of deflection. Data is compared to mathematical models which include a feedback equation to capture the temporal changes in the profile of the limit cycle oscillations.

Effects of Electrical and Mechanical Overstimulus on Spontaneous Oscillations in Hair Bundles

Spontaneous oscillations constitute one of the manifestations of the active process operant in hair cells and provides a sensitive probe for their internal dynamics. The influx of ions into the stereocilia can be modulated by applying an electrical current across the epithelium and has been previously shown to strongly affect the oscillatory profiles. We applied strong transient stimuli and demonstrated that they can induce a transition from the oscillatory to the quiescent state, an effect that can last over several seconds post stimulus cessation. The dynamics of recovery to the oscillatory state was found to be dependent on the amplitude and the duration of the stimulus. Similar dynamics were observed after high‐amplitude mechanical stimulus, which mimics the effects of loud sound on an individual bundle.

Numerical Study of the Complex Temporal Pattern of Spontaneous Oscillation in Bullfrog Saccular Hair Cells

Hair bundles of the bullfrog sacculus display spontaneous oscillations that show complex temporal profiles. Quiescent intervals are typically interspersed with oscillations, analogous to bursting behavior observed in neural systems. By introducing slow calcium dynamics into the theoretical model of bundle mechanics, we reproduce numerically the multi‐mode oscillations and explore the effects of internal parameters on the temporal profiles and the frequency tuning of their linear response functions. We also study the effects of mechanical overstimulation on the oscillatory behavior.

Exploring the Electrical Resonance's Affect on the Mechanical Oscillations of Hair Cells in the Bullfrog Sacculus

Under in vitro conditions, uncoupled hair bundles of the bullfrog (Rana catesbeiana) sacculus have been shown to exhibit spontaneous oscillations. We used a high-speed complementary metal oxide semiconductor camera to track the movements of hundreds of cells in parallel from dozens of preparations. This work revealed that the spontaneous oscillations exhibit multiple timescales with a slow modulation on a rapid oscillation. Experiments inhibiting the electrical resonance in the cell body show a strong effect on the mechanical oscillations of the hair bundles. This indicates that the electrical oscillation is coupled with the mechanical oscillations of the hair bundles.