Fast recovery of disrupted tip links in hair bundles
Hearing and balance rely on the capacity of mechanically sensitive hair bundles to transduce vibrations into electrical signals that are forwarded to the brain. Hair bundles possess tip links that interconnect the mechanosensitive stereocilia and convey force to the transduction channels. A dimer of dimers, each of these links comprises two molecules of protocadherin 15 (PCDH15) joined to two of cadherin 23 (CDH23). The “handshake” that conjoins the four molecules can be disrupted in vivo by intense stimulation and in vitro by exposure to Ca2+ chelators. Using hair bundles from the rat's cochlea and the bullfrog’s sacculus, we observed that extensive recovery of mechanoelectrical transductio and hair-bundle stiffness can occur within seconds after Ca2+ chelation, especially if hair bundles are deflected towards their short edges. Investigating the phenomenon in a two-compartment ionic environment that mimics natural conditions, we combined iontophoretic application of a Ca2+ chelator to selectively disrupt the tip links of individual frog hair bundles with displacement clamping to control hair-bundle motion and measure forces. Our observations suggest that, after the normal Ca2+ concentration has been restored, mechanical stimulation facilitates the reconstitution of functional tip links.
Left: In a displacement-clamp experiment, a feedback system imposes a ramp displacement on a hair bundle (first trace), moving it first in the positive direction, then more extensively in the negative direction. At three times during this paradigm, a 500 ms epoch of ±25 nm, 40 Hz sinusoidal stimulation is superimposed on the displacement-command signal. An iontophoretic pulse (third trace and blue band) releases EDTA to break tip links. The force (second trace) necessary to clamp the bundle at the outset (Initial) diminishes after exposure EDTA (Exposure) but recovers almost completely by the experiment's end (Recovered). Right: Data from seven hair cells reveal a significant decrease in hair-bundle stiffness after iontophoretic pulses; the stiffness then recovers following negative hair-bundle displacements.
