Mechano-electrical transduction: new insights into old ideas

Abstract

The gating-spring theory of hair cell mechanotransduction channel activation was first postulated over twenty years ago. The basic tenets of this hypothesis have been reaffirmed in hair cells from both auditory and vestibular systems and across species. In fact, the basic findings have been reproduced in every hair cell type tested. A great deal of information regarding the structural, mechanical, molecular and biophysical properties of the sensory hair bundle and the mechanotransducer channel has accumulated over the past twenty years. The goal of this review is to investigate new data, using the gating spring hypothesis as the framework for discussion. Mechanisms of channel gating are presented in reference to the need for a molecular gating spring or for tethering to the intra- or extracellular compartments. Dynamics of the sensory hair bundle and the presence of motor proteins are discussed in reference to passive contributions of the hair bundle to gating compliance. And finally, the molecular identity of the channel is discussed in reference to known intrinsic properties of the native transducer channel.

Fig. 1

Hair bundles are dynamic structures that come in various sizes and shapes. (a–c) are scanning electron micrographs of cultured rat inner (a), mature rat outer (b) and mature mouse vestibular (c) hair bundles from rat. (d)isa transmission electron micrograph (TEM) where long arrows point to side links, short arrows to the rootlets (rt). CP is cuticular plate. The schematic of (e) shows the typical staircase pattern of hair bundles with the directionally sensitive placement of the tip-link. (f) TEM of the stereociliary pair shows the tenting on the apical surface of the stereocilia as well as the tip-links. Arrows indicate insertional plaques. The inset is a surface plot rendering of the freeze-etching view of the helical structure of the tip-link (from Kachar et al., 2000), made using the Image J image-processing program. (g) Schematic representation of the hair bundle depicting the active turnover process, termed treadmilling. (h) Schematic depicting the location of myosins, in particular the myosin XVa proposed to be critical in setting stereocilia height. Other myosins include myosin VI, VIIa and Ic.

Fig. 2

The mechanism of MET channel activation is unknown. These schematic representations illustrate that a variety of possible mechanisms exist. In each panel the arrow indicates the direction of the applied force. (A) shows the traditionally accepted view that the tip-link is directly associated with the MET channel and exerts force perpendicular to the membrane onto the channel. (B) demonstrates that the tip-link does not need to be associated with the channel but could simply exert force onto the lipid membrane. (C) similarly demonstrates that, dependent on geometry, the force exerted onto the lipid may be parallel to the lipid just as well as perpendicular. (D) further demonstrates that accessory structures like myosins may be linked to the membrane and not the channel. This configuration can produce adaptation. Neither existing data nor existing theoretical arguments can distinguish between these possibilities.

Fig. 3

Voltage-clamp recordings of mechanoelectric transduction currents and hair bundle movements that underlie the gating-spring theory. (A) Upper panel shows mechanical stimulus paradigm and lower panel, MET currents obtained from a hair cell in response to flexible fiber stimulation. (B) Measured hair bundle motion in response to force stimulation. Note the notch at the onset and the slower increase in movement with larger stimuli. (C) Plotting the applied force against displacement at the timepoint of peak current response results in a nonlinear plot that underlies the gating-spring hypothesis. The region of minimal slope corresponds to the steepest portion of the MET activation plot (D) and indicates a reduction in hair bundle stiffness at these positions. (D) Differentiating the force plot results in a stiffness plot (blue trace) that clearly shows a minimum in stiffness that correlates well with the steepest portion of the activation plot. Note that the stiffness axis start is nonzero. A single Boltzmann function was used to fit the activation plot (see Eq 3).

Fig. 4

Mechanical and electrical properties of a two-state gating spring model of the MET channel. (A) Free energies as a function of hair bundle position of the two states. A_o is the free energy of the open state (dashed line) and A_c that of the closed state (dash-dotted line). The channels average free energy A_ch (continuous line;) A_ch = p_oA_o + p_cA_c). The energy specifically related to gating is indicated as A_g (dotted line). Note that A_ch also equals A_ch = A_o + A_g (Eq. 4). The closed state is engaged at, X ≥ X_c; whereas the open state is activated at X ≥ X_o. (B) Forces acting on a single transduction channel, as resulting from the free energies shown in (A) F_ch = dA_ch/dX is the sum of F_o = dA_o/dX and F_g = dA_g/dX (C) Stiffness resulting from forces depicted in (B). S_ch = dF_ch/dX is the sum of S_o = dF_o/dX and S_g = dF_g/dX. The gating compliance is the reduction of S_ch with respect to k_gs. (D) Open probability as obtained from the free energy difference of A_c and A_o (Eqs. 3 and 4). Parameters: ε_o = 3.5 kT; ε_o = 0 kT; ; X_o = −30 nm; X_c = −5 nm, so that D = X_o − X_c = 25 nm; k_gs = 10 μN/m.

Fig. 5

The native MET channel is a nonspecific cation channel that shows little rectification in high external calcium solutions. (A) Plot of the MET current against voltage. The reversal potential of +7 mV demonstrates the nonselective nature of the channel. Little rectification is observed. The data were fit with the single-site binding equation that allows for the estimate of the location of a binding site (presumably a calcium binding site) within the pore (Woodhull, 1973; Kros et al., 1992; Farris et al., 2004). A location approximately 0.5 of the distance into the electric field was obtained. (B) Examples of single-channel recording responses from four stimulus cycles from a high (red) and low (blue) frequency cell. Larger currents are obtained from high-frequency cells than from low-frequency cells. (C) Schematic representation of the MET channel pore illustrating the estimated molecular dimensions of the channel. Curare is shown acting as an open-channel blocker in the pore of the channel and represents the prominent mechanisms of block (Farris et al., 2004).