**Simultaneous measurement of NLC and motility.** NLC (*A*) and OHC displacement (*B*) were measured simultaneously as described under “Experimental Procedures” and in supplemental Fig. S1. The data were analyzed using the two-state Boltzmann model-based equations ( and ) shown under “Results.” The *solid lines* are the fitting curves derived from the equations. The simultaneous NLC and motility measurements were performed on 36 different OHCs, and α values (*C*) and *V*_{pk} values (*D*) were compared against each other for testing their identity. The *error bars* represent the S.E. of the curve-fitting analyses. The *diagonal broken lines* indicate exact match (*y* = *x*). Deming linear regression analyses (*solid lines*) followed by *t* tests were performed for quantitatively testing identity (see “Experimental Procedures”). The *p* values for the *y* intercept and the slope were 0.83 and 0.62 (*C*) and 0.33 and 0.39 (*D*), none of which disagreed with the null hypothesis of identity, *i.e. a* = 0, *b* = 1. *E*, *Q*_{max} and *D*_{max} were estimated from the curve fitting analyses shown in *A* and *B. D*_{max} values are plotted against the corresponding *Q*_{max} (*n* = 36). The *error bars* represent S.E. of the curve-fitting analyses. Alternatively, the observed maximum OHC displacement (*d*) is plotted against the observed prestin-dependent charge movement (*q*) that is determined by the area between NLC curves and the *C*_{lin} (*E*, *inset*). The Deming linear regression analysis was performed with the *y* intercept fixed at zero (*y* = *ax*). *r*^{2} values for *Q*_{max}-*D*_{max} and *q-d* were 0.036 and 0.41, respectively. The NLC motility data (*n* = 36) were also analyzed by the three-state model, and the α_{1} and α_{2} values (*F*) and the *V*_{pk1} and *V*_{pk2} values (*G*) were compared against each other. Because the three-state fitting on NLC/motility data recorded in the absence of salicylate usually finds larger S.E., only data sets whose S.E. found in *V*_{pk1}/*V*_{pk2} were less than 100 mV are shown, with *bold symbols error bars*, whereas others are shown with *smaller pale symbols without error bars*. The *p* values determined by Deming linear regression analyses (*solid lines*) followed by *t* tests for the *y* intercept (=0) and the slope (=1) were less than 0.05 for α_{1}, α_{2}, *V*_{pk1}, and *V*_{pk2} comparisons, suggesting that α_{1}, α_{2}, *V*_{pk1}, and *V*_{pk2} values are different between NLC and motility. *H*, shown are computer simulations of *m*_{1}/*m*_{2}-dependent OHC displacements using . The parameters used were 0.0252 mV^{−1}, 0.0239 mV^{−1}, −85.7 mV, and −52.6 mV for α_{1}, α_{2}, *V*_{pk1}, and *V*_{pk2}, all of which were derived from the NLC measurements (*F* and *G*). The generated displacement data were analyzed by in which *m*_{1}/*m*_{2} was fix at unity, and the resultant fitting parameters were plotted against *m*_{1}/*m*_{2} for α_{1} and α_{2} (*I*) and *V*_{pk1} and *V*_{pk2} (*J*). The *broken lines* indicate the α and *V*_{pk} values used for generating the displacement data. *K* and *L*, NLC and OHC displacement were measured simultaneously as in *A* and *B* in the presence of 0.1∼1 mm salicylate. The NLC motility data (*n* = 15) were analyzed by the three-state model, and the α_{1} and α_{2} values (*K*) and the *V*_{pk1} and *V*_{pk2} values (*L*) were compared against each other. The *p* values determined by Deming linear regression analyses (*solid lines*) followed by *t* tests for the *y* intercept (=0) and the slope (=1) were all greater than 0.05 for α_{1}, α_{2}, *V*_{pk1}, and *V*_{pk2} comparisons, suggesting that the NLC and motility are coupled in terms of α_{1}–α_{2} and *V*_{pk1}–*V*_{pk2}.

## PubMed Commons