-frequency assessment. They found flat responses out to 10 kHz at room

-frequency assessment. They found flat responses out to 10 kHz at room temperature. Importantly, the direct effects of temperature on OHC displacement currents and NLC have been evaluated and shown to substantially affect NLC Vh (indicative of transition-rate effects) of both OHC and prestintransfected cells when the bath temperature is altered (30,54,55). Shifts of 20 mV/ C were found. Additionally, temperature jumps using an infrared laser on prestin-transfected cells (56) induced Cm changes attributable to NLC as well as linear Cm, as originally described by Shapiro et al. (57). NLC Vh shifted with rates up to 14 V/s over the course of a 5 ms infrared pulse. Thus, it is clear that temperature will influence the frequency dependence of OHC NLC. Consequently, after correcting for temperature, Gale and Ashmore (12) arrived at a 25 kHz cutoff for NLC, still far below the eM cutoff observed at room temperature (11). We suggest that these incompatible measures arise from technical issues. Considering the characteristics of our recently espoused meno presto model (24,28), we further suggest that sufficiently long stimulations of the OHC will drive substantial numbers of prestins into the chloride-bound, voltage-enabled state where they may rapidly respond to voltage perturbations with cutoff frequencies possibly unencumbered by the chloride-binding step. Thus, the performance of the OHC may modulate between two frequency regimes, high and low; the latter likely related to a slow transport function of the A-836339 cancer protein (40,41,43). Recently, Homma et al. (58) have measured the frequency dependence of OHC NLC using our dual-sine approach, but only with discrete dual-sine frequencies and without presentation of linear capacitance data. Interestingly, they found that NLC in control mouse OHCs was frequency independent with high intracellular iodide solutions, but frequency dependent with high intracellular chloride conditions. The latter results are similar to our results under high-chloride conditions. Thus, we concur that anions are influential in controlling prestin kinetics, and we now must consider the effects of chloride substitutes on prestin kinetics. Here, we used gluconate–previously confirmed to be similar to aspartate substitution (18)–to lower chloride to near2558 Biophysical Journal 110, 2551?561, June 7,Chloride Controls Prestin Kineticsphysiological levels (16). Whether any of the effects of iodide were due to chloride reductions remains to be investigated. Interestingly, Albert et al. (33) also presented data showing low-pass NLC activity in rat prestin using singlesine measurements (see their Fig. 3 E), which they attributed to their recording equipment. Yet they note very fast clamp speeds, and furthermore, they do not claim any untoward influences on the low-pass nature of zebra fish prestin in that same study. How can low-pass prestin sensor charge movement that directly drives eM underlie cochlear amplification? Cochlear amplification provides a boost to auditory sensitivity ranging from 100- to 1000-fold. It is thought to be maximal at high acoustic frequencies, in the tens of kilohertz range. There is ample evidence that prestin-driven OHC electromechanical activity underlies cochlear amplification, yet how can a voltage-dependent process that relies on a low-pass voltage sensor to drive mechanical activity work? We have previously Tirabrutinib clinical trials estimated that mechanical responses at high acoustic frequencies would be markedly smaller th.-frequency assessment. They found flat responses out to 10 kHz at room temperature. Importantly, the direct effects of temperature on OHC displacement currents and NLC have been evaluated and shown to substantially affect NLC Vh (indicative of transition-rate effects) of both OHC and prestintransfected cells when the bath temperature is altered (30,54,55). Shifts of 20 mV/ C were found. Additionally, temperature jumps using an infrared laser on prestin-transfected cells (56) induced Cm changes attributable to NLC as well as linear Cm, as originally described by Shapiro et al. (57). NLC Vh shifted with rates up to 14 V/s over the course of a 5 ms infrared pulse. Thus, it is clear that temperature will influence the frequency dependence of OHC NLC. Consequently, after correcting for temperature, Gale and Ashmore (12) arrived at a 25 kHz cutoff for NLC, still far below the eM cutoff observed at room temperature (11). We suggest that these incompatible measures arise from technical issues. Considering the characteristics of our recently espoused meno presto model (24,28), we further suggest that sufficiently long stimulations of the OHC will drive substantial numbers of prestins into the chloride-bound, voltage-enabled state where they may rapidly respond to voltage perturbations with cutoff frequencies possibly unencumbered by the chloride-binding step. Thus, the performance of the OHC may modulate between two frequency regimes, high and low; the latter likely related to a slow transport function of the protein (40,41,43). Recently, Homma et al. (58) have measured the frequency dependence of OHC NLC using our dual-sine approach, but only with discrete dual-sine frequencies and without presentation of linear capacitance data. Interestingly, they found that NLC in control mouse OHCs was frequency independent with high intracellular iodide solutions, but frequency dependent with high intracellular chloride conditions. The latter results are similar to our results under high-chloride conditions. Thus, we concur that anions are influential in controlling prestin kinetics, and we now must consider the effects of chloride substitutes on prestin kinetics. Here, we used gluconate–previously confirmed to be similar to aspartate substitution (18)–to lower chloride to near2558 Biophysical Journal 110, 2551?561, June 7,Chloride Controls Prestin Kineticsphysiological levels (16). Whether any of the effects of iodide were due to chloride reductions remains to be investigated. Interestingly, Albert et al. (33) also presented data showing low-pass NLC activity in rat prestin using singlesine measurements (see their Fig. 3 E), which they attributed to their recording equipment. Yet they note very fast clamp speeds, and furthermore, they do not claim any untoward influences on the low-pass nature of zebra fish prestin in that same study. How can low-pass prestin sensor charge movement that directly drives eM underlie cochlear amplification? Cochlear amplification provides a boost to auditory sensitivity ranging from 100- to 1000-fold. It is thought to be maximal at high acoustic frequencies, in the tens of kilohertz range. There is ample evidence that prestin-driven OHC electromechanical activity underlies cochlear amplification, yet how can a voltage-dependent process that relies on a low-pass voltage sensor to drive mechanical activity work? We have previously estimated that mechanical responses at high acoustic frequencies would be markedly smaller th.