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Hair Cell Transduction, Tuning, and Synaptic Transmission in the Mammalian Cochlea

Robert Fettiplace

10.1002/cphy.c160049

Source: Volume 7, Issue 4, October 2017

Published online: September 2017

Full Article on Wiley Online Library



ABSTRACT

Sound pressure fluctuations striking the ear are conveyed to the cochlea, where they vibrate the basilar membrane on which sit hair cells, the mechanoreceptors of the inner ear. Recordings of hair cell electrical responses have shown that they transduce sound via submicrometer deflections of their hair bundles, which are arrays of interconnected stereocilia containing the mechanoelectrical transducer (MET) channels. MET channels are activated by tension in extracellular tip links bridging adjacent stereocilia, and they can respond within microseconds to nanometer displacements of the bundle, facilitated by multiple processes of Ca2+‐dependent adaptation. Studies of mouse mutants have produced much detail about the molecular organization of the stereocilia, the tip links and their attachment sites, and the MET channels localized to the lower end of each tip link. The mammalian cochlea contains two categories of hair cells. Inner hair cells relay acoustic information via multiple ribbon synapses that transmit rapidly without rundown. Outer hair cells are important for amplifying sound‐evoked vibrations. The amplification mechanism primarily involves contractions of the outer hair cells, which are driven by changes in membrane potential and mediated by prestin, a motor protein in the outer hair cell lateral membrane. Different sound frequencies are separated along the cochlea, with each hair cell being tuned to a narrow frequency range; amplification sharpens the frequency resolution and augments sensitivity 100‐fold around the cell's characteristic frequency. Genetic mutations and environmental factors such as acoustic overstimulation cause hearing loss through irreversible damage to the hair cells or degeneration of inner hair cell synapses. © 2017 American Physiological Society. Compr Physiol 7:1197‐1227, 2017.

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Figure 1. Figure 1. Schematic of the sound transmission pathway from the eardrum to the cochlea. Sound stimuli impinge on the tympanum (t), or eardrum, at the end of the ear canal and the vibrations (denoted by red arrows) are transmitted through the three bones of the middle ear: malleus (m), incus (i), and stapes (s). The footplate of the stapes behaves like a piston in the oval window and initiates pressure waves in the cochlear fluids so setting in vibration the basilar membrane. The pressure is relieved at the round window (rw). The cochlea, here depicted as straight, is in situ coiled like a snail's shell and embedded in the petrous temporal bone. It is subdivided into three compartments containing perilymph or endolymph fluid, the two outer compartment being connected by the helicotrema. The total length of the cochlea is 35 mm (humans), 26 mm (cat), 18 mm (guinea pig), and 6 mm (mice).
Figure 2. Figure 2. Cross section though the cochlear duct showing the cellular structure. The scala media is delimited by Reissner's membrane, the spiral ligament, and the basilar membrane, which is surmounted by the organ of Corti. The width of the basilar membrane ranges from approximately 100 to 500 μm in humans. The scala media is filled with a K+‐based endolymph, here colored pink. The organ of Corti contains the sensory hair cells embedded in assorted supporting cells of distinct shape. The hair‐cell stereociliary bundles are covered in an acellular tectorial sheet and the cells are innervated by the cochlear branch of the VIIIth cranial nerve. Inner hair cells are contacted by afferents (orange) whereas outer hair cells are innervated mainly by efferent fibers (yellow). The stria vascularis is an epithelial strip on the lateral wall that is specialized for secreting endolymph.
Figure 3. Figure 3. Schematic of the stria vascularis. The stria comprises two cellular layers separated by an intrastrial space. Marginal cells face the endolymph and intermediate/basal cells, interconnected by gap junctions (blue pairs of lines), are exposed to fibrocytes of the strial ligament and perilymph; adjacent cells in each layer are linked by tight junctions (purple). (Note that the orientation is reversed with regard to that shown in Fig. 2.) Flow of K+ ions is facilitated by the inwardly rectifying KCNJ10 K+ channel on intermediate cells and the KCNQ1/KCNE1 K+ channel on the endolymphatic aspect of the marginal cells. Ionic balance is maintained by Na/K ATPase, Na‐2Cl‐K and Cl transporters. The voltages given (+90, +100, and +10 mV) refer to the static potentials of the extracellular spaces with respect to the scala tympani. The endolymphatic potential of +90 mV is attributable to a Nernst K+ equilibrium potential of ∼100 mV across the highly K+ selective apical membrane of intermediate cells. The intrastrial space has low K+ due to uptake of the ion by the Na‐2Cl‐K cotransporter and the Na/K ATPase and K+ is then secreted into endolymph across the K+‐selective membrane of marginal cell.
Figure 4. Figure 4. Stereociliary bundles and the transduction apparatus. Scanning electron micrographs of stereociliary bundles of (A) an outer hair cell and (B) an inner hair cell, showing the staircase in heights of the rows. (C) Transmission electron micrograph of an outer hair cell showing a tip link connecting two stereocilia; the insertion sites of the tip link (TL) are heavily electron dense suggesting dense protein densities. (D) Schematic of the molecular structure of the tip link apparatus deduced from various mutations. USH‐1 and USH‐2 denote different Usher type 1 and type 2 mutations. The association between the N‐termini of protocadherin‐15 and cadherin‐23 is Ca2+ dependent. Two MET channels (red) are situated at the lower end of the tip link and are present as complexes with TMIE, LHFPL5, TMC1, and possibly other proteins. Modified, with permission, from (80).
Figure 5. Figure 5. Mechanoelectrical transducer (MET) currents in outer hair cells. (A) Schematic of the stimulating and recording techniques. OHCs are patch clamped and the stereociliary bundle is deflected either by a glass probe attached to a piezoelectric device or by a fluid jet. Displacement of the bundle are calibrated by projection of image onto a photodiode array (55,243). (B) MET currents for family of step displacements, X, of a hair bundle, displaying rapid rise to peak and then adaptive decline to a steady level. (C) Plot of peak MET current against bundle displacement with an operating range of ∼0.25 μm. (D) Expanded scale of MET current onset showing that it develops as quickly as the displacement step (shown above) but then adapts with a time constant, τA, of 100 μs. (E) MET currents in OHCs from the apex and base of the cochlea for sinusoidal modulation of hair bundle position (top). Bundle motion was calibrated by projecting its image on to a pair of photodiodes, the noisy grey trace denoting the photocurrent. (F) MET current increases from apex to base of cochlea; current amplitude was 50% larger in the reduced Ca2+ of the endolymph solution‐bathing bundle. All currents measured at a holding potential of −84 mV. Modified, with permission, from (80,146).
Figure 6. Figure 6. Single MET channels in mouse hair cells. (A) Apical outer hair cell: four representative single channel records for 150 nm hair bundle displacement steps; middle, ensemble average of 10 responses; bottom amplitude histograms giving mean single‐channel current of 6.2 pA. (B) Basal outer hair cell: four representative single channel records for 150 nm hair bundle stimuli; middle, ensemble average of 10 responses; bottom, amplitude histograms giving mean single‐channel current of 12 pA. (C) Single‐channel current and conductance (mean ± 1 SD) as a function of position in the cochlea, expressed as relative distance from the apical end. Total length of cochlea is 6 mm. All measurements made at room temperature and −84 mV holding potential. Modified, with permission, from (23).
Figure 7. Figure 7. Adaptation assayed with two‐pulse experiment. (A) MET currents for two series of brief bundle displacements, the first are control steps and the second are test steps, which are preceded by a long adapting step. Note the current decay during the adapting step. (B) Current‐displacement relationships for first (control) pulse and for second (test) pulse after adapting step. The current I is scaled to its maximum value, Imax. Note the positive shift, ΔX0.5, in the current‐displacement relationship. (C) Schematic of experiment where the amplitude of the adapting step was varied. (D) Plot of shift in current‐displacement relation, ΔX0.5, as a function of the size of the adapting step. The slope is typically 0.5–0.6. All currents measured in outer hair cells at a holding potential of −84 mV. Results, with permission, from reference (18).
Figure 8. Figure 8. Tonotopic variations in membrane properties of rodent outer hair cells. (A) Principal membrane currents determining potential of outer hair cell. MET current, IMT, carried mainly by K+ ions, flows in through MET channels down a potential gradient determined by the positive endolymphatic potential (EP, 90 mV) and the resting potential (VR, ∼ −50 mV); the K+ current exits mainly via GK,n channels in lateral wall, down a K+ concentration gradient into the perilymph. (B) MET conductance, GMT, increases with the characteristic frequency at the location of the hair cell. (C) Voltage‐dependent K+ conductance, GK,n, increases with hair‐cell characteristic frequency. (D) Membrane capacitance decreases with hair‐cell characteristic frequency, signifying a progressive decrease in the size, mainly the length, of the outer hair cell. Combining results in B, C, and D, implies a significant reduction in the membrane time constant determined by C/(GMET + GKn). Results are combined measurements from gerbils (filled circles) and rats (filled squares) and were taken, with permission, from (129). (E) OHC length (and hence membrane area and electrical capacitance) decreases with increase in characteristic frequency in different mammals: (a) chinchilla, human; (b) guinea pig; (c) chinchilla, gerbil; (d) guinea pig, chinchilla; (e) gerbil, rat; (f) chinchilla, mouse, rat; (g) guinea pig, rat, human; (h) rat, bat; (i) mouse; (j) bat. Data, with permission, from [(60); rat, bat, guinea pig, and gerbil], [(25); chinchilla], [(239,235); human], and from author's laboratory (rat, mouse, and gerbil).
Figure 9. Figure 9. Filtering of receptor potentials by inner hair cell. (A) Changes in IHC membrane potential elicited by current pulses of magnitudes given next to each trace in isolated guinea pig inner hair cell. Note the voltage inactivation for larger responses. (B) Schematic of organ of Corti showing the IHC and innervation by multiple afferents. The medial and lateral sides of the IHC are often referred to as “modiolar” and “pillar,” the orientation of which is shown beneath the schematic. (C) Receptor potentials in an inner hair cell of an anesthetized guinea pig for tones of different frequencies, given in Hz alongside the traces. At low frequencies, the response is purely sinusoidal, reflecting the sound stimulus. At frequencies above 1000 Hz, the periodic (AC) component is filtered by the membrane time constant leaving a sustained depolarizing (DC) component. (D) Synchronization index, indicating phase‐locking in auditory nerve discharge, as a function of the frequency of the sound stimulus in auditory nerve fibers of cats (crosses) and guinea pigs (filled and open squares). An index of 1.0 denotes perfect synchronization of the spikes to a specific phase on every cycle of the tone, whereas an index of 0 denotes no relationship between the spike firing and the sound cycle. Records in (A) modified, with permission, from (153) and (C) and (D), with permission, from (222). See also Figure 14 for examples of phase locking.
Figure 10. Figure 10. Tonotopic organization of the turtle auditory papilla. Left, medial view with the hair‐cell papilla on the right‐hand side of the basilar membrane; scale bar = 100 μm. Right, examples of electrical resonance in hair cells at different positions along the epithelium. Resonant frequency, given beside traces, increases from apex to base. Each record is the voltage response to a small depolarizing current step, the timing of which is shown at top; cells had resting potentials in the range −44 to −51 mV. Figure taken, with permission, from (246).
Figure 11. Figure 11. Mechanical and electrical tuning curves in the mammalian cochlea. (A) Solid curves are frequency‐threshold tuning curves for two auditory nerve fibers in the chinchilla cochlea, with characteristic frequencies of 0.4 and 9.5 kHz. Superimposed on each nerve‐fiber tuning curve at similar locations are the basilar membrane vibrations: iso‐displacement response (dotted curves, 1‐nm left and 2.7 nm right) and isovelocity response (dashed curves, 2.5 μm/s left, and 164 μm/s right). The results indicate almost all of the frequency tuning is present in the basilar membrane vibrations, with isovelocity responses giving better fits to the nerve fiber frequency‐threshold curve; from (254). (B) Schematic of auditory nerve fiber tuning curves for the cat cochlea based on results in references (165,125). Similar sets of tuning curves are also available for other mammals including the Mongolian gerbil (215) and the mouse (283).
Figure 12. Figure 12. Outer hair cell contractility mediated by prestin. (A) Schematic of outer hair cell with prestin molecules in lateral wall. Force applied to hair bundle open MET channels, causing depolarization and cell contraction due to change in conformation of prestin. (B) Transmission electron micrograph of rat outer hair cell immunolabeled for prestin shows gold particles in the lateral wall; abbreviations: st, stereociliary bundle; cp cuticular plate; cy, cytoplasm; jc junctional complex. (C) Contractions of outer hair cell evoked by voltage steps from −120 mV to +50 mV; length change measured with dual photodiode; (D) plots of length change in outer hair cell recorded with chloride‐based and sulfate‐based intracellular solutions. With chloride, the prestin was half‐activated at −50 mV, but sulfate shifted the activation relationship ∼150 mV positive. B taken, with permission, from (176); C and D taken, with permission, from (141).
Figure 13. Figure 13. Deformation of organ of Corti during stimulation. (A) Excitatory (rarefaction) sound stimulus causes upward deflection of basilar membrane and organ of Corti. On conventional view, the entire organ moves upward without changing shape and causes abneural displacement of hair bundles; brown background denotes resting position and black outline new stimulated position. (B) Electrical stimulation elicits contraction of outer hair cells and compression of the organ of Corti, with the reticular lamina being pulled down and basilar membrane pulled up. During normal stimulation it is envisage that both processes in A and B will occur sequentially but the exact timing is still uncertain.
Figure 14. Figure 14. Synaptic potentials and action potentials in an auditory afferent. (A) Microelectrode recordings from an auditory nerve terminal in the turtle cochlea showing the spontaneous synaptic potentials and action potentials in the absence of a sound stimulus (top) and the response evoked by a tone at 265 Hz, 54 dB SPL (bottom). (B) Peristimulus histograms showing phase locking of action potentials to a 265 Hz tone (top) and a 520 Hz tone (bottom) from cell in (A); modified, with permission, from reference (53).
Figure 15. Figure 15. The synapse between the inner hair cell and cochlear afferent fiber. (A) Inner hair cell makes synaptic contacts with multiple (10‐20) afferent fibers on its basolateral aspect, each synapse having one presynaptic ribbon (blue) and release site onto one afferent. Fibers synapsing on the pillar side are thought to have low thresholds and high resting spontaneous firing; fibers synapsing on the modiolar side have high threshold and low spontaneous discharge. The ribbons are smaller and the postsynaptic glutamate receptor densities (blue strip) are larger for the low threshold fibers. (B) Enlargement of the (blue) ribbon surrounded by halo of (yellow) synaptic vesicles. The ribbon is composed of ribeye and piccolo proteins and anchored to the membrane of the release site by bassoon. Vesicles are exocytosed by Ca2+ influx through Cav1.3 Ca2+ channels on presynaptic membrane and glutamate neurotransmitter binds to GluA2/3 receptors on the postsynaptic membrane. (C) High power view of synaptic vesicle, glutamate transporter Vglut3, and Ca2+ sensor otoferlin with six #c2 domains. (D) Conventional view of the life cycle of the synaptic vesicle, from docking at the release site, interaction of vesicular and target SNARE proteins and priming for release, fusion and reuptake. Several of these processes are thought to be Ca2+‐sensitive and possibly mediated by otoferlin.


Figure 1. Schematic of the sound transmission pathway from the eardrum to the cochlea. Sound stimuli impinge on the tympanum (t), or eardrum, at the end of the ear canal and the vibrations (denoted by red arrows) are transmitted through the three bones of the middle ear: malleus (m), incus (i), and stapes (s). The footplate of the stapes behaves like a piston in the oval window and initiates pressure waves in the cochlear fluids so setting in vibration the basilar membrane. The pressure is relieved at the round window (rw). The cochlea, here depicted as straight, is in situ coiled like a snail's shell and embedded in the petrous temporal bone. It is subdivided into three compartments containing perilymph or endolymph fluid, the two outer compartment being connected by the helicotrema. The total length of the cochlea is 35 mm (humans), 26 mm (cat), 18 mm (guinea pig), and 6 mm (mice).


Figure 2. Cross section though the cochlear duct showing the cellular structure. The scala media is delimited by Reissner's membrane, the spiral ligament, and the basilar membrane, which is surmounted by the organ of Corti. The width of the basilar membrane ranges from approximately 100 to 500 μm in humans. The scala media is filled with a K+‐based endolymph, here colored pink. The organ of Corti contains the sensory hair cells embedded in assorted supporting cells of distinct shape. The hair‐cell stereociliary bundles are covered in an acellular tectorial sheet and the cells are innervated by the cochlear branch of the VIIIth cranial nerve. Inner hair cells are contacted by afferents (orange) whereas outer hair cells are innervated mainly by efferent fibers (yellow). The stria vascularis is an epithelial strip on the lateral wall that is specialized for secreting endolymph.


Figure 3. Schematic of the stria vascularis. The stria comprises two cellular layers separated by an intrastrial space. Marginal cells face the endolymph and intermediate/basal cells, interconnected by gap junctions (blue pairs of lines), are exposed to fibrocytes of the strial ligament and perilymph; adjacent cells in each layer are linked by tight junctions (purple). (Note that the orientation is reversed with regard to that shown in Fig. 2.) Flow of K+ ions is facilitated by the inwardly rectifying KCNJ10 K+ channel on intermediate cells and the KCNQ1/KCNE1 K+ channel on the endolymphatic aspect of the marginal cells. Ionic balance is maintained by Na/K ATPase, Na‐2Cl‐K and Cl transporters. The voltages given (+90, +100, and +10 mV) refer to the static potentials of the extracellular spaces with respect to the scala tympani. The endolymphatic potential of +90 mV is attributable to a Nernst K+ equilibrium potential of ∼100 mV across the highly K+ selective apical membrane of intermediate cells. The intrastrial space has low K+ due to uptake of the ion by the Na‐2Cl‐K cotransporter and the Na/K ATPase and K+ is then secreted into endolymph across the K+‐selective membrane of marginal cell.


Figure 4. Stereociliary bundles and the transduction apparatus. Scanning electron micrographs of stereociliary bundles of (A) an outer hair cell and (B) an inner hair cell, showing the staircase in heights of the rows. (C) Transmission electron micrograph of an outer hair cell showing a tip link connecting two stereocilia; the insertion sites of the tip link (TL) are heavily electron dense suggesting dense protein densities. (D) Schematic of the molecular structure of the tip link apparatus deduced from various mutations. USH‐1 and USH‐2 denote different Usher type 1 and type 2 mutations. The association between the N‐termini of protocadherin‐15 and cadherin‐23 is Ca2+ dependent. Two MET channels (red) are situated at the lower end of the tip link and are present as complexes with TMIE, LHFPL5, TMC1, and possibly other proteins. Modified, with permission, from (80).


Figure 5. Mechanoelectrical transducer (MET) currents in outer hair cells. (A) Schematic of the stimulating and recording techniques. OHCs are patch clamped and the stereociliary bundle is deflected either by a glass probe attached to a piezoelectric device or by a fluid jet. Displacement of the bundle are calibrated by projection of image onto a photodiode array (55,243). (B) MET currents for family of step displacements, X, of a hair bundle, displaying rapid rise to peak and then adaptive decline to a steady level. (C) Plot of peak MET current against bundle displacement with an operating range of ∼0.25 μm. (D) Expanded scale of MET current onset showing that it develops as quickly as the displacement step (shown above) but then adapts with a time constant, τA, of 100 μs. (E) MET currents in OHCs from the apex and base of the cochlea for sinusoidal modulation of hair bundle position (top). Bundle motion was calibrated by projecting its image on to a pair of photodiodes, the noisy grey trace denoting the photocurrent. (F) MET current increases from apex to base of cochlea; current amplitude was 50% larger in the reduced Ca2+ of the endolymph solution‐bathing bundle. All currents measured at a holding potential of −84 mV. Modified, with permission, from (80,146).


Figure 6. Single MET channels in mouse hair cells. (A) Apical outer hair cell: four representative single channel records for 150 nm hair bundle displacement steps; middle, ensemble average of 10 responses; bottom amplitude histograms giving mean single‐channel current of 6.2 pA. (B) Basal outer hair cell: four representative single channel records for 150 nm hair bundle stimuli; middle, ensemble average of 10 responses; bottom, amplitude histograms giving mean single‐channel current of 12 pA. (C) Single‐channel current and conductance (mean ± 1 SD) as a function of position in the cochlea, expressed as relative distance from the apical end. Total length of cochlea is 6 mm. All measurements made at room temperature and −84 mV holding potential. Modified, with permission, from (23).


Figure 7. Adaptation assayed with two‐pulse experiment. (A) MET currents for two series of brief bundle displacements, the first are control steps and the second are test steps, which are preceded by a long adapting step. Note the current decay during the adapting step. (B) Current‐displacement relationships for first (control) pulse and for second (test) pulse after adapting step. The current I is scaled to its maximum value, Imax. Note the positive shift, ΔX0.5, in the current‐displacement relationship. (C) Schematic of experiment where the amplitude of the adapting step was varied. (D) Plot of shift in current‐displacement relation, ΔX0.5, as a function of the size of the adapting step. The slope is typically 0.5–0.6. All currents measured in outer hair cells at a holding potential of −84 mV. Results, with permission, from reference (18).


Figure 8. Tonotopic variations in membrane properties of rodent outer hair cells. (A) Principal membrane currents determining potential of outer hair cell. MET current, IMT, carried mainly by K+ ions, flows in through MET channels down a potential gradient determined by the positive endolymphatic potential (EP, 90 mV) and the resting potential (VR, ∼ −50 mV); the K+ current exits mainly via GK,n channels in lateral wall, down a K+ concentration gradient into the perilymph. (B) MET conductance, GMT, increases with the characteristic frequency at the location of the hair cell. (C) Voltage‐dependent K+ conductance, GK,n, increases with hair‐cell characteristic frequency. (D) Membrane capacitance decreases with hair‐cell characteristic frequency, signifying a progressive decrease in the size, mainly the length, of the outer hair cell. Combining results in B, C, and D, implies a significant reduction in the membrane time constant determined by C/(GMET + GKn). Results are combined measurements from gerbils (filled circles) and rats (filled squares) and were taken, with permission, from (129). (E) OHC length (and hence membrane area and electrical capacitance) decreases with increase in characteristic frequency in different mammals: (a) chinchilla, human; (b) guinea pig; (c) chinchilla, gerbil; (d) guinea pig, chinchilla; (e) gerbil, rat; (f) chinchilla, mouse, rat; (g) guinea pig, rat, human; (h) rat, bat; (i) mouse; (j) bat. Data, with permission, from [(60); rat, bat, guinea pig, and gerbil], [(25); chinchilla], [(239,235); human], and from author's laboratory (rat, mouse, and gerbil).


Figure 9. Filtering of receptor potentials by inner hair cell. (A) Changes in IHC membrane potential elicited by current pulses of magnitudes given next to each trace in isolated guinea pig inner hair cell. Note the voltage inactivation for larger responses. (B) Schematic of organ of Corti showing the IHC and innervation by multiple afferents. The medial and lateral sides of the IHC are often referred to as “modiolar” and “pillar,” the orientation of which is shown beneath the schematic. (C) Receptor potentials in an inner hair cell of an anesthetized guinea pig for tones of different frequencies, given in Hz alongside the traces. At low frequencies, the response is purely sinusoidal, reflecting the sound stimulus. At frequencies above 1000 Hz, the periodic (AC) component is filtered by the membrane time constant leaving a sustained depolarizing (DC) component. (D) Synchronization index, indicating phase‐locking in auditory nerve discharge, as a function of the frequency of the sound stimulus in auditory nerve fibers of cats (crosses) and guinea pigs (filled and open squares). An index of 1.0 denotes perfect synchronization of the spikes to a specific phase on every cycle of the tone, whereas an index of 0 denotes no relationship between the spike firing and the sound cycle. Records in (A) modified, with permission, from (153) and (C) and (D), with permission, from (222). See also Figure 14 for examples of phase locking.


Figure 10. Tonotopic organization of the turtle auditory papilla. Left, medial view with the hair‐cell papilla on the right‐hand side of the basilar membrane; scale bar = 100 μm. Right, examples of electrical resonance in hair cells at different positions along the epithelium. Resonant frequency, given beside traces, increases from apex to base. Each record is the voltage response to a small depolarizing current step, the timing of which is shown at top; cells had resting potentials in the range −44 to −51 mV. Figure taken, with permission, from (246).


Figure 11. Mechanical and electrical tuning curves in the mammalian cochlea. (A) Solid curves are frequency‐threshold tuning curves for two auditory nerve fibers in the chinchilla cochlea, with characteristic frequencies of 0.4 and 9.5 kHz. Superimposed on each nerve‐fiber tuning curve at similar locations are the basilar membrane vibrations: iso‐displacement response (dotted curves, 1‐nm left and 2.7 nm right) and isovelocity response (dashed curves, 2.5 μm/s left, and 164 μm/s right). The results indicate almost all of the frequency tuning is present in the basilar membrane vibrations, with isovelocity responses giving better fits to the nerve fiber frequency‐threshold curve; from (254). (B) Schematic of auditory nerve fiber tuning curves for the cat cochlea based on results in references (165,125). Similar sets of tuning curves are also available for other mammals including the Mongolian gerbil (215) and the mouse (283).


Figure 12. Outer hair cell contractility mediated by prestin. (A) Schematic of outer hair cell with prestin molecules in lateral wall. Force applied to hair bundle open MET channels, causing depolarization and cell contraction due to change in conformation of prestin. (B) Transmission electron micrograph of rat outer hair cell immunolabeled for prestin shows gold particles in the lateral wall; abbreviations: st, stereociliary bundle; cp cuticular plate; cy, cytoplasm; jc junctional complex. (C) Contractions of outer hair cell evoked by voltage steps from −120 mV to +50 mV; length change measured with dual photodiode; (D) plots of length change in outer hair cell recorded with chloride‐based and sulfate‐based intracellular solutions. With chloride, the prestin was half‐activated at −50 mV, but sulfate shifted the activation relationship ∼150 mV positive. B taken, with permission, from (176); C and D taken, with permission, from (141).


Figure 13. Deformation of organ of Corti during stimulation. (A) Excitatory (rarefaction) sound stimulus causes upward deflection of basilar membrane and organ of Corti. On conventional view, the entire organ moves upward without changing shape and causes abneural displacement of hair bundles; brown background denotes resting position and black outline new stimulated position. (B) Electrical stimulation elicits contraction of outer hair cells and compression of the organ of Corti, with the reticular lamina being pulled down and basilar membrane pulled up. During normal stimulation it is envisage that both processes in A and B will occur sequentially but the exact timing is still uncertain.


Figure 14. Synaptic potentials and action potentials in an auditory afferent. (A) Microelectrode recordings from an auditory nerve terminal in the turtle cochlea showing the spontaneous synaptic potentials and action potentials in the absence of a sound stimulus (top) and the response evoked by a tone at 265 Hz, 54 dB SPL (bottom). (B) Peristimulus histograms showing phase locking of action potentials to a 265 Hz tone (top) and a 520 Hz tone (bottom) from cell in (A); modified, with permission, from reference (53).


Figure 15. The synapse between the inner hair cell and cochlear afferent fiber. (A) Inner hair cell makes synaptic contacts with multiple (10‐20) afferent fibers on its basolateral aspect, each synapse having one presynaptic ribbon (blue) and release site onto one afferent. Fibers synapsing on the pillar side are thought to have low thresholds and high resting spontaneous firing; fibers synapsing on the modiolar side have high threshold and low spontaneous discharge. The ribbons are smaller and the postsynaptic glutamate receptor densities (blue strip) are larger for the low threshold fibers. (B) Enlargement of the (blue) ribbon surrounded by halo of (yellow) synaptic vesicles. The ribbon is composed of ribeye and piccolo proteins and anchored to the membrane of the release site by bassoon. Vesicles are exocytosed by Ca2+ influx through Cav1.3 Ca2+ channels on presynaptic membrane and glutamate neurotransmitter binds to GluA2/3 receptors on the postsynaptic membrane. (C) High power view of synaptic vesicle, glutamate transporter Vglut3, and Ca2+ sensor otoferlin with six #c2 domains. (D) Conventional view of the life cycle of the synaptic vesicle, from docking at the release site, interaction of vesicular and target SNARE proteins and priming for release, fusion and reuptake. Several of these processes are thought to be Ca2+‐sensitive and possibly mediated by otoferlin.
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Teaching Material

R. Fettiplace. Hair Cell Transduction, Tuning, and Synaptic Transmission in the Mammalian Cochlea. Compr Physiol 7:2017, 1197-1227. doi:10.1002/cphy.c160049

Didactic Synopsis

Major Teaching Points:

  • Sound is detected by hair cells, the mechanoreceptors of the cochlea, the spiral cavity of the inner ear. Hair cells are excited by submicrometer vibrations of their stereociliary (hair) bundles, which are converted into changes in membrane potential graded with sound intensity.
  • Hair bundles are bathed in an extracellular fluid, high in potassium and low in calcium and at a positive 100 mV potential, which optimizes the transduction process.
  • There are two types of hair cell with disparate functions: outer hair cells contain prestin, a piezoelectric motor protein that generates force to amplify the mechanical stimulus, inner hair cells communicate with auditory nerve fibers via a ribbon synapse at the cochlear output.
  • The frequency of a sound is analyzed by both passive and active tuning mechanisms, with subsets of hair cells along the cochlear being tuned to different frequencies.
  • Transduction can be disrupted by mutations in single genes leading to permanent deafness; many of these genes are linked to the hair bundle structure.

Didactic Legends

The figures—in a freely downloadable PowerPoint format—can be found on the Images tab along with the formal legends published in the article. The following legends to the same figures are written to be useful for teaching.

Figure 1. This figure shows the pathway for conducting sound waves from the eardrum via three small bones of the middle ear to the cochlea. The stapes acts like a piston, generating pressure waves in the cochlear fluid, which result in small displacements of the basilar membrane synchronized to the sound stimulus. A pure tone (sinusoidal) sound stimulus will produce a traveling wave on the basilar membrane that peaks at a position characteristic of the stimulus frequency. The properties of the cochlea vary from one end to the other, which causes the frequency components in the sound to be separated along it length like an acoustic prism. Thus, low frequencies are most effective at the distal apex near the helicotrema, whereas high frequencies elicit vibrations, which are confined to the proximal base. This is referred to as a tonotopic organization. The cochlea is divided into three compartments, with the central compartment containing endolymph fluid having an ionic composition which, with its high K+ concentration, differs from that in most extracellular spaces.

Figure 2. This figure depicts a slice through the cochlear tube of Figure 1 showing the location of the sensory hair cells (in green), which detect the sound stimulus. The hair cells are embedded in an array of specially named supporting cells in the organ of Corti. There are two types of hair cell: inner hair cells and outer hair cells. Inner hair cells produce the major output of the cochlea, and synapse on afferent neurons of the VIIIth cranial nerve, which generate action potentials. Outer hair cells contract and amplify the sound-evoked motion. Both types of hair cell are covered in a gelatinous structure termed the tectorial membrane, and they detect vibrations of this membrane relative to the basilar membrane. Bundles of modified microvilli, referred to as stereocilia, project from the tops of each hair cell, and are the site of mechanically sensitive ion channels where transduction occurs. The stereociliary bundles are immersed in a fluid of special ionic composition known as endolymph that fills the scala media (indicated in pink).

Figure 3. This shows a schematic of the stria vascularis, an epithelial strip on the side wall of the cochlea, which secretes the K+-based endolymph and generates ~100 mV endocochlear potential (EP). Both features facilitate hair cell transduction. These properties are achieved by two layers of cells, each tightly interconnected, with membranes containing K+ ion channels (KCNQ1, KCNJ10) and transporters (NKCC), that participate in the secretion of endolymph. Ion channels are proteins with a central pore that are embedded in the lipid bilayer of the plasma membrane, through which ions traverse. The voltages given (+90, +100, and +10 mV) refer to the static potentials of the extracellular spaces with respect to the scala tympani.

Figure 4. Details of the ultrastructure and protein composition of the stereociliary bundles and the transduction apparatus. Scanning electron micrographs of stereociliary bundles of an outer hair cell and an inner hair cell illustrate the staircase in heights of the rows. Deflection towards the taller row of stereocilia is excitatory. The rows are interconnected by various extracellular filaments, the most important of which is the tip link connecting the tip of one stereocilium with the side wall of its taller neighbor. A number of proteins form the tip link and the transduction apparatus, which includes two mechanoelectrical transducer (MET) ion channels situated at the lower end of the tip link. Much of this information has been gleaned from study of genetic mutations, especially those that occur in the deafness-blindness Usher syndrome.

Figure 5. This figure illustrates the properties of the electrical currents evoked in outer hair cells in response to displacement of their stereociliary bundles. The electrical currents are measured by patch clamping hair cells during a series displacement steps to the bundles. Mechanoelectrical transducer (MET) currents show a rapid rise to a peak followed by a subsequent decline in a process known as adaptation. As in other sensory receptors, adaptation optimizes transducer sensitivity around the resting state. The peak MET current encodes bundle displacements over a fraction of a micron. The maximum amplitude of the MET current depends on the location of the hair cell in the cochlea, and increases from the low-frequency apex to the high-frequency base. The systematic change in the amplitude of the macroscopic current along the cochlea is largely accounted for by a changes in the single-channel conductance. Its role may be to enhance the sensitivity in high frequency cells.

Figure 6. As shown in Figure 4, the ion channels underlying the macroscopic transducer current in hair cells are localized to the tips of the shorter stereocilia where they are activated by tension in tip links. Channels can be isolated by severing almost all of the tip links with exposure to submicromolar Ca2+. The figure shows examples of single MET ion channel events. The currents look square, with the channel being either closed (C) or open (O). The amplitude of the current events is larger in outer hair cells at the base than at the apex. The gradient in channel conductance largely accounts for the systematic change in macroscopic current shown and it implies that the molecular composition of the MET channel varies along the cochlea.

Figure 7. This figure shows one method of quantifying the adaptation of the MET current. This is done by presenting two sets of brief displacement stimuli, the second of which is preceded by a long adapting step. Adaptation is manifested by a shift in the sigmoidal current-displacement relationship along the displacement axis. Varying the amplitude of the adapting step shows that the positive shift, DX0.5, in the current-displacement relationship increases as a function of the adapting amplitude.

Figure 8. Multiple properties of the basilar membrane and the hair cells change along the cochlea to enable the distribution of frequency components in a sound stimulus. The figure shows how both structural and ionic properties of the outer hair cells vary systematically along the cochlea, each location being identified with a characteristic sound frequency. Factors increasing with characteristic frequency include the magnitude of the MET conductance, the number of K+ channels that determine the resting membrane potential, and the electrical capacitance. The electrical capacitance is attributable to the plasma membrane, and its magnitude reflects the surface area of the outer hair cell. Systematic variation is produced mainly by a change in outer hair cell length which varies in different mammals between 8 and 80 microns.

Figure 9. Inner hair cells are the main output of the cochlea and are innervated by multiple auditory afferent fibers. A sinusoidal sound stimulus produces a receptor potential in an inner hair cell, and action potentials are first generated in the auditory nerve fibers. This figure summarizes the change in the IHC receptor potential and in the firing of action potentials for different sound frequencies. At low frequencies, the IHC response is purely sinusoidal, reflecting the sound stimulus. At frequencies above 1000 Hz, the periodic (AC) component of the receptor potential is filtered by the membrane time constant leaving a sustained depolarizing (DC) component. The periodic component is also manifest in the phase locking of auditory nerve action potentials synchronized to the cycles of the sound at low frequencies but phase locking is lost at high frequencies.

Figure 10. In all vertebrate cochleas, a hair cell’s characteristic frequency changes systematically along the organ. The figure illustrates this tonotopic organization in the turtle cochlea, where the membrane potential of each hair cell resonates when the cell is stimulated electrically. In most nonmammals, frequency tuning is accomplished by an electrical resonance achieved by the interplay of K+ and Ca2+ channels in the hair cell membrane. These channels tune the receptor potential, with different resonant frequencies being generated by altering the density and kinetics of the K+ channels. Although the frequency range (60 to 400 Hz) is much lower than in mammals, the orientation of the frequency map is the same.

Figure 11. This figure shows frequency tuning curves in the mammalian cochlea. For each plot, the sound level required to elicit a threshold response is an approximately V-shaped function of the sound frequency, with the tip of the “V” being referred to as the characteristic frequency (CF). The sound threshold is expressed in decibels (dB), which is a logarithmic measure, 20 dBs being a ten-fold pressure change. Comparison of the frequency-threshold tuning curves for two auditory nerve fibers with the frequency tuning in the basilar membrane vibrations indicates that almost all of the tuning is present in the basilar membrane motion. The figure also shows multiple frequency tuning curves for auditory nerve fibers in the cat cochlea, illustrating how the shape of the tuning curves changes across the frequency spectrum.

Figure 12. There has been uncertainty about the mechanisms involved in producing the narrow frequency tuning curves in Figure 11. It is well established that the passive mechanical properties of the basilar membrane generate some frequency tuning but another metabolically sensitive (active) mechanism is needed to augment the selectivity and produce narrow tuning curves. The active mechanism is mediated by a piezoelectric motor protein known as prestin in the outer hair cell membrane, which is activated by changes in membrane potential. This figure shows some features of prestin. Localization of prestin to the OHC lateral membrane can be demonstrated by antibody labeling and electron microscopy. Depolarization of the OHC causes the outer hair cell to contract, with the cell length showing a sigmoidal dependence on membrane potential around a resting potential of about -50 mV. Correct operation of the prestin mechanism requires the presence of chloride ions.

Figure 13. This figure shows that the organ of Corti is deformed during a sound stimulus. A reduction in sound pressure at the eardrum (a rarefaction) causes an upward displacement of the basilar membrane and organ of Corti. On the conventional view, the entire organ moves upward without changing its shape, and results in lateral displacement of hair bundles and hair cell excitation. The brown background indicates the organ of Corti at rest, and the black outline the new position when stimulated. This view of a rigid organ of Corti cannot be completely accurate because, upon stimulation, outer hair cells contract and cause the organ of Corti to compress, bringing the reticular lamina and basilar membrane closer together. The latter effect can be studied in the absence of a sound stimulus by electrically stimulating the organ of Corti to elicit outer hair cell contraction. During normal a stimulus, both processes in A and B will occur sequentially but their relative timing is still not precisely known.

Figure 14. This figure shows microelectrode recordings of the excitatory synaptic potentials and action potentials in an auditory afferent terminal. These recordings indicate that there are spontaneous synaptic potentials and action potentials in the absence of a sound stimulus. However, in response to a tone, the action potentials become synchronized, or phase locked, to the cycles of tones at 265 and 520 Hz. Not all synaptic potentials evoke action potentials.

Figure 15. This figure shows the schematic structure of the synapse between the inner hair cell and cochlear afferent fiber derived from transmission electron micrographs and antibody labeling against specific protein components. An inner hair cell makes synaptic contacts with multiple (10-20) afferent fibers, which have different sensitivities. This glutamatergic synapse is unusual in a number of respects. It is characterized by an electron dense presynaptic structure known as a ribbon, which is surrounded by a halo of synaptic vesicles containing the neurotransmitter. Fusion of the synaptic vesicles with the plasma membrane is triggered by an increase in cytoplasmic Ca2+. Unlike most synapses, Ca2+ influx occurs via L-type Ca2+ channels, whereas at other synapses in the central nervous system N-type and P-type Ca2+ channels are more common. Furthermore, exocytosis is not regulated by the Ca2+ binding protein synaptotagmin, which is absent. Instead, several of the steps in synaptic vesicle docking, fusion and reuptake are thought mediated by the Ca2+-binding protein otoferlin. The glutamate neurotransmitter released into the synaptic cleft binds to AMPA receptors on the postsynaptic membrane.


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Article Title: Hair Cell Transduction, Tuning, and Synaptic Transmission in the Mammalian Cochlea
 

How to Cite

Robert Fettiplace. Hair Cell Transduction, Tuning, and Synaptic Transmission in the Mammalian Cochlea. Compr Physiol 2017, 7: 1197-1227. doi: 10.1002/cphy.c160049