Claims 48
From the paper A novel theoretical framework reveals more than one voltage-sensing pathway in the lateral · claims file · checked 2026-11-06: ready
- cohort
The data set holds the paper's 89 OHCs, 39 from adult males (17 albino, 22 pigmented), 21 from prepubertal males (15 albino, 6 pigmented), 28 from prepubertal females (10 albino, 18 pigmented) and 1 pigmented cell from an adult female.
Electrophysiological data were obtained from 89 OHCs, namely 39 (17 albino and 22 pigmented) from adult males, 21 (15 albino and 6 pigmented) from prepubertal males, 28 (10 albino and 18 pigmented) from prepubertal females, and 1 pigmented from an adult female
Materials and methods, Animals and OHC isolation, paragraph 4How it will be judged
Counted from the data set's per-cell records of colony, sex and maturity. Reproduced when all eight counts of the quote match exactly; partially reproduced when the total is 89 and no group count is off by more than 2; not reproduced otherwise. The counts of cells with 10 mM chloride (3, Methods, Whole-cell patch-clamp, paragraph 1), with XE991, with Rs < 10 MOhm (81 of 89) and the recording times (47 to 182 min) are reported as well.
- figs1-example-cell
The example cell of Fig. S1 (original number 99, juvenile pigmented female, 60 uM XE991) has CL 24.45 pF, CLn 25.94 pF, CLW 18.56 pF, V0.5 -0.022 V, QTm 3.02 pC and alphaB 34.98 V-1.
Cell (original number 99) was obtained from a juvenile normal (pigmented) female guinea pig, and the experiment was conducted in the presence of 60 μM XEE91. Voltages were corrected for the voltage drop across the pipette where the measured parameters are C L , 24.45 pF; C L n , 25.94 pF; C L W , 18.56 pF; V 0 . 5 , −0.022 V; Q T m , 3.02 pC; and α B , 34.98 V −1
Figure S1 legendHow it will be judged
The cell is found by its original number 99 (and colony, sex, maturity and XE991 as the legend gives them). Values are recomputed from its capacitance record by the shared method (opening comment) and also read from any stored per-cell parameters; both are shown. Reproduced when all six recomputed values are within tolerance; partially reproduced when four or five are; not reproduced otherwise. The legend's "XEE91" is read as XE991.
- nlw-capacitance
The capacitance of the non-lateral-wall region is about 4.82 pF, close to the 4.86 pF reported before for adult males, and it does not differ between phenotypes or between adult and juvenile animals.
The capacitance of the nonlateral wall region was found to be ∼4.82 pF and is similar to that previously reported (4.86 pF) when only cells isolated from adult male guinea pigs were considered ( Corbitt et al., 2012 ). We found no differences when cells were isolated from animals of either phenotype or when cells were isolated from adult or juvenile animals with this dataset
Results, Experimental magnitude of CLW, QTm, alphaB and V0.5, paragraph 1How it will be judged
Method from Expressions used to determine the experimental parameters, paragraph 2 (Eqs. 8b and 8c, whose rendering is missing). QTm of each cell is plotted against CL - CSB and the root exp(-gamma/beta) of the fitted relation is found, with beta and gamma fitted constants; the prose and Results paragraph 1 ("QTM increases with the logarithm of lateral wall capacitance") fix the relation as QTm = gamma + beta ln(CL - CSB), so the root is the value of CL - CSB where QTm is zero. CNLW is then CSB plus that root. CSB, the stereocilia bundle capacitance, is "calculated" by a method the paper takes from Corbitt et al. (2012) and does not give; open. Readings (n-a) CSB as stored per cell in the data set, if it is; (n-b) CNLW and CLW as stored per cell; (n-c) CSB = 0, so CNLW is the root alone. The form of Eq. 8c (CNLW = CSB + root) is rebuilt from the prose. Which cells enter the fit is not given; readings all 89 cells, or the 86 with 144 mM chloride (the population of Fig. 1). Reproduced when under one reading CNLW is within tolerance and separate fits for pigmented against albino, and adult against juvenile, give roots whose 95% confidence intervals overlap (the paper names no test for "no differences"; overlap of intervals is the reading used here, and the size of each difference is shown).
- clw-by-turn
The lateral membrane capacitance ranges from 5.0 (+/- 2.4) pF for turn 1 cells to 17.9 (+/- 2.74) pF for turn 4 cells.
The capacitance of the lateral membrane ranged from 5.0 (± 2.4) pF for cells that originate from turn 1 to 17.9 (± 2.74) pF for cells from turn 4
Results, Experimental magnitude of CLW, QTm, alphaB and V0.5, paragraph 1How it will be judged
Tolerances are the spread the paper states. Mean CLW (shared method) over the cells of turn 1 and of turn 4. The cells are not named; readings (c-a) all cells of the turn; (c-b) the 144 mM chloride cells (the population of Fig. 1); (c-c) the 70 cells of Fig. 2 (no XE991). The standard deviations are reported and compared with 2.4 and 2.74 (within 10%).
- fig1-charge-not-linear
The charge does not grow linearly with the lateral wall capacitance as identical and independent sensors would give (Eq. 6b fit, adjusted R2 0.516); the square of the charge rises much more steeply with CLW^2 for basal cells (CLW < 10 pF) than for apical cells.
In contrast, Q T M increases with the logarithm of lateral wall capacitance ( Corbitt et al., 2012 ), or the square of the charge increases much more steeply with C L W 2 for sensors found on basal originating cells ( C L W < 10 pF) than for sensors found on the apical originating cells ( Fig. 1 )
Figure 1; Results, Experimental magnitude of CLW, QTm, alphaB and V0.5, paragraph 1How it will be judged
Cells, the 86 with 144 mM chloride in the pipette (Fig. 1 legend), with and without XE991. Eq. 6b as rebuilt (QTm^2 proportional to CLW^2) is fitted by least squares. The paper does not say whether the fit has an intercept; readings (f-a) through the origin, as Eq. 6b has none; (f-b) with an intercept; and whether the R2 is taken on the scale of QTm^2 against CLW^2 (as the Results describe Fig. 1) or of QTm against CLW, both shown. The adjusted R2 is reported under each. Reproduced when under one reading the adjusted R2 is within tolerance and the slope of QTm^2 on CLW^2 fitted to cells with CLW < 10 pF is larger than the slope fitted to cells with CLW >= 10 pF (same intercept reading), with non-overlapping 95% confidence intervals of the two slopes (the paper gives no test for "much more steeply"; this is the reading used, and the ratio of the slopes is shown).
- fig1-two-state-bounds
The charge predicted by Eq. 6b for a membrane holding only one of the two sensor states (lambda 3.95 kB T with C_Leq 0.584 aF, and lambda 2.16 kB T with C_Leq 1.07 aF) bounds the measured charges, cells between the two limits holding mixtures of the two states.
We calculate with Eq. 6b the predicted charge if only one of the states exists within the lateral membrane and compare these calculations with the experimental data to show that these two states cover the data range ( Fig. 1 , dashed red lines)
Discussion, Kinetic scheme, paragraph 4; Figure 1 legendHow it will be judged
With Eq. 6b as rebuilt, the two lines are QTm = CLW sqrt(2 lambda / C_Leq), that is dVmin_exp = 0.2356 V and 0.1286 V (lambda in joules at 297.15 K). The 86 cells of Fig. 1. The paper gives no measure of "cover the data range"; readings (b-a) at least 90% of cells have dVmin_exp between the two values widened by 10% of each; (b-b) the cells of CLW < 10 pF lie near the upper line and those of CLW >= 10 pF near the lower line, judged by the median dVmin_exp of each group falling within 10% of 0.2356 V and 0.1286 V respectively (the legend places the two lines in the left and right panels). Reproduced when either reading holds; the fraction inside the band is reported.
- alpha-pigmented-vs-albino
The maximum voltage sensitivity alpha(V0.5) is significantly greater for OHCs from pigmented animals (8.62 +/- 0.95 V-1, 28 cells) than from albinos (7.65 +/- 0.88 V-1, 41 cells).
We found the maximum voltage sensitivity α V 0 . 5 for OHCs originating from pigmented animals (8.62 ± 0.95 V −1 ) to be significantly greater (P < 0.0001; number of observations, 28) than α ( V 0.5 ) observed for cells isolated from albinos (7.65 ± 0.88 V −1 ; number of observations, 41)
Results, Experimental magnitude of CLW, QTm, alphaB and V0.5, paragraph 2How it will be judged
Tolerances are the stated standard deviations. alpha(V0.5) per cell by the shared method. The 28 pigmented cells match the pigmented cells without XE991 in Table 1 (12 + 16); the 41 albino cells match Table 1 (11 + 30). The test is not named; Table 1 uses one-way ANOVA, so readings (t-a) one-way ANOVA on colony (equivalent to a two-sided pooled t-test); (t-b) Welch's t-test. Reproduced when the pigmented mean exceeds the albino mean with p < 0.0001 under one reading and both means are within tolerance, and also within 10% of the reported value (shown); partially reproduced when the direction holds with p < 0.05 only.
- alpha-gradient-pigmented
For pigmented animals alpha(V0.5) falls from base (9.32 +/- 0.69 V-1) to apex (8.08 +/- 0.76 V-1), a significant difference, while albinos show no gradient (basal 7.76 +/- 0.98, apical 7.59 +/- 0.83 V-1).
we found that the maximum voltage sensitivity decreases across the cochlea from the base (9.32 ± 0.69 V −1 ) to apex (8.08 ± 0.76 V −1 ) for OHCs isolated from pigmented animals, with no gradient (basal, 7.76 ± 0.98 V −1 ; apical, 7.59 ± 0.83 V −1 ) observable for OHCs isolated from albinos ( Fig. S2 ). ... The positional differences between basal and apical regions for cells from pigmented animals are statistically significant
Results, Experimental magnitude of CLW, QTm, alphaB and V0.5, paragraph 2; Figure S2How it will be judged
Tolerances are the stated standard deviations. The pigmented values times four are Table 1's rows (b) (37.3 and 32.3 V-1); the albino values times four (31.0 and 30.4 V-1) are close to but not equal to rows (c) (31.3 and 30.5 V-1), so the albino cells may differ slightly from Table 1's. The cells are taken as those rows' (pigmented without XE991, albino) and the test is Table 1's one-way ANOVA; basal and apical by readings (r-a) and (r-b) of the opening comment. Reproduced when the four means are within tolerance under one region reading, the pigmented difference has p < 0.05 and the albino difference has p >= 0.05 (the absence judged by the same ANOVA, as Table 1 does).
- table1-pigmented-xe991
With XE991 in the bath, cells from pigmented animals have higher alphaB at the base (36.4 V-1, z 0.93, 4 cells) than at the apex (33.8 V-1, z 0.86, 11 cells), P = 0.0227.
How it will be judged
Tolerances are the standard deviations Table 1 prints. alphaB = 4 alpha(V0.5) and z = alphaB kB T / e0 at 297.15 K (opening comment). Cells, pigmented with XE991 and 144 mM chloride. One-way ANOVA of alphaB on region (Table 1 footnote). Reproduced when under one region reading the counts are 4 and 11, the means are within tolerance and p < 0.05; partially reproduced when the direction holds and p < 0.05 with other counts.
- table1-pigmented-no-xe991
Without XE991, cells from pigmented animals have higher alphaB at the base (37.3 V-1, z 0.96, 12 cells) than at the apex (32.3 V-1, z 0.83, 16 cells), P < 0.0001.
How it will be judged
As table1-pigmented-xe991, for pigmented cells without XE991. Reproduced when under one region reading the means are within tolerance and p < 0.0001; partially reproduced when the direction holds with p < 0.05.
- table1-albino-no-gradient
Cells from albinos show no difference of alphaB between base (31.3 V-1, z 0.80, 11 cells) and apex (30.5 V-1, z 0.78, 30 cells), P = 0.5294.
How it will be judged
A claim of absence, judged on the paper's test, a one-way ANOVA of alphaB on region for the albino cells, at p < 0.05. Reproduced when the means are within tolerance and the test gives p >= 0.05 under the region reading that gives counts closest to 11 and 30 (r-a unless r-b matches better); not reproduced when it finds the difference. The difference is reported either way.
- table1-xe991-no-effect-alpha
Adding XE991 to the bath has no significant effect on alphaB (P = 0.7551), so pigmented cells with and without XE991 are pooled, giving basal alphaB 37.1 +/- 2.5 V-1 (z 0.95, 16 cells) and apical 32.9 +/- 2.7 V-1 (z 0.84, 27 cells).
Therefore, we grouped the results (XE991 and no XE991) from cells obtained from pigmented animals and calculated the mean α B and z for cells originating from basal and apical regions of 37.1 ± 2.5 V −1 ( z , 0.95 ± 0.06; number, 16) and 32.9 ± 2.7 V −1 ( z , 0.84 ± 0.07; number, 27)
Table 1 footnote; Results, Experimental magnitude of CLW, QTm, alphaB and V0.5, paragraph 2How it will be judged
The absence is judged on the paper's test, an "ANOVA effect test" of XE991 (Table 1 footnote). Its model is not given; readings (x-a) a two-way ANOVA of alphaB on region and XE991 over the pigmented cells (the footnote's rows a and b); (x-b) a two-way ANOVA on region and XE991 over all three groups of Table 1, with colony as a third factor; (x-c) a one-way ANOVA of alphaB on XE991 over the pigmented cells. Reproduced when under one reading the XE991 effect has p >= 0.05 and the pooled means are within tolerance with counts 16 and 27; not reproduced when the XE991 effect has p < 0.05 under the reading closer to the paper's counts. The effect size is shown. The pooled counts (16, 27) are those of Table 1 (4 + 12, 11 + 16).
- figs2-alphab-vs-clw
alphaB decreases monotonically with lateral membrane capacitance for cells from pigmented animals, with no relationship for cells from albinos.
When α B is plotted as a function of lateral membrane capacitance, it decreases monotonically for cells from pigmented animals, with no relationship observed for the population of cells isolated from albinos ( Fig. S2 )
Results, Experimental magnitude of CLW, QTm, alphaB and V0.5, paragraph 2; Figure S2, right panelsHow it will be judged
The paper gives no test. Readings (s-a) Spearman correlation of alphaB with CLW; (s-b) least-squares linear regression of alphaB on CLW. Cells, as Fig. S2, the pigmented and albino cells (the legend does not say whether XE991 cells are included; with and without are both shown). Reproduced when, under one reading, the pigmented relation is negative with p < 0.05 and the albino relation has p >= 0.05. This rests on a test the paper does not make, and is marked as resting on an assumption.
- splice-isoform-x1
Only isoform X1 of guinea pig prestin is found in the cochleae of pigmented and albino animals, so splice variants cannot explain the voltage-sensitivity gradient.
In all experiments, only isoform X1 was detected upon sequencing ( Fig. S3 ). We conclude that splice variants cannot explain the voltage sensitivity gradient
Results, Experimental magnitude of CLW, QTm, alphaB and V0.5, paragraph 3; Figure S3How it will be judged
Rests on PCR and sequencing of cochlear cDNA (Methods, Sequencing the prestin gene), which are not electrophysiology recordings; the data set, by its description, holds only the voltage-clamp data. Reproduced only if the sequencing products are found.
- v05-by-turn
V0.5 is not constant but ranges from -0.083 +/- 0.030 V (turn 1) to -0.050 +/- 0.018 V (turn 4) for albino cells, and from -0.102 +/- 0.019 V to -0.052 +/- 0.020 V for pigmented cells.
V 0.5 is not constant but ranges from −0.083 ± 0.030 V for turn 1 to −0.050 ± 0.018 V for turn 4 cells extracted from albinos, and from −0.102 ± 0.019 V (turn 1) to −0.052 ± 0.020 V (turn 4) for cells extracted from a normal pigmented phenotype
Results, Experimental magnitude of CLW, QTm, alphaB and V0.5, paragraph 4How it will be judged
Tolerances are the stated standard deviations. V0.5 corrected for the pipette drop (shared method). Cells, readings (v-a) the 70 cells of Fig. 2 (144 mM chloride, no XE991); (v-b) all cells of the turn and colony. Reproduced when under one reading all four means are within tolerance and turn 1 is more negative than turn 4 in both colonies; the means are also compared within 10% of each reported value and shown.
- fig2b-v05-vs-clw
V0.5 increases monotonically with CLW, becoming more positive as CLW increases; the linear relation is better for pigmented cells (adjusted R2 about 0.6) than for albino cells (about 0.2).
V 0.5 increases monotonically with C L W , becoming more positive as C L W increases ( Fig. 2 b ). ... However, the linear relationship is better with cells from pigmented phenotype (adjusted R 2 of ∼0.6) compared with cells from albinos (adjusted R 2 of ∼0.2)
Figure 2b; Results, Experimental magnitude of CLW, QTm, alphaB and V0.5, paragraph 4How it will be judged
Cells, the 70 of Fig. 2 (144 mM chloride, no XE991; legend). Least-squares linear fit of corrected V0.5 on CLW per colony. The paper gives no test of the slopes; the p < 0.05 on each slope is a choice made here. Reproduced when both slopes are positive with p < 0.05 and both adjusted R2 are within tolerance; partially reproduced when the slopes are positive and the pigmented R2 exceeds the albino R2 but one is outside tolerance. The paper's "about" values carry the default 10% tolerance.
- fig2b-no-phenotype-difference
The V0.5 gradient with CLW does not differ statistically between the two phenotypes.
There is no statistical difference between the gradient observed for each phenotype
Results, Experimental magnitude of CLW, QTm, alphaB and V0.5, paragraph 4How it will be judged
A claim of absence. The paper does not name its test. Readings (g-a) the CLW by colony interaction in a linear model of V0.5 on CLW, colony and their interaction; (g-b) a t-test of the difference of the two fitted slopes. Reproduced when the test gives p >= 0.05 under (g-a) (the standard test of equal slopes), not reproduced when p < 0.05; (g-b) shown as a variant, with the two slopes and their difference.
- v05-gradient-length
In guinea pig OHCs the V0.5 gradient along the cochlea is 0.7 um/mV in terms of cell length.
The gradient is more than that measured in cells from guinea pigs, at 0.7 μm/mV
Results, Experimental magnitude of CLW, QTm, alphaB and V0.5, paragraph 4How it will be judged
Cell length from the bright-field image (Methods, Whole-cell patch-clamp, paragraph 2). The fit is not given; readings (l-a) slope of length regressed on V0.5 in mV; (l-b) the inverse of the slope of V0.5 regressed on length; each on (l-1) the 70 cells of Fig. 2 or (l-2) all cells with a length. Reproduced when under one reading the gradient is within tolerance.
- xe991-voltage-clamp
XE991 improves the voltage clamp at least twofold, the fractional pipette drop staying below 13% (against e.g. 26% at voltages above 0.08 V without), while conductance at V0.5 with XE991 reaches 335 pS.
When XE991 was added to the extracellular solution, it caused at least a twofold improvement in the voltage clamp, with the fractional drop consistently <13%. In addition, the conductance measured at V 0.5 in the presence of XE991 was as high as 335 pS
Results, Experimental magnitude of CLW, QTm, alphaB and V0.5, paragraph 5How it will be judged
Fractional drop is Rs / (Rs + Rm) (the complement of the correction factor in Methods, Whole-cell patch-clamp, paragraph 2). Readings for the voltage at which it is taken, (d-a) at V0.5; (d-b) at positive voltages (0.09 to 0.15 V, where Rm and Rs are measured). Reproduced when under one reading every XE991 cell has a drop below 13% (widened by 10%), the median drop without XE991 is at least twice the median with XE991 (the paper names no measure of "twofold improvement"; the medians are a choice made here, the means shown as a variant), and the largest conductance at V0.5 among XE991 cells is within tolerance.
- table2-xe991-v05-shift
XE991 shifts V0.5 by about 20 mV in the depolarizing direction (ANOVA effect test P = 0.0035 in pigmented cells) without affecting the other electrical parameters.
We also report a surprising 20mV depolarizing shift of V 0.5 upon addition of XE991 ( Table 2 ); this channel blocker did not affect the other electrical parameters
Results, Experimental magnitude of CLW, QTm, alphaB and V0.5, paragraph 5; Table 2 footnoteHow it will be judged
The tolerance is not the default 10%; the text's "20mV" is a rounding of Table 2, whose own means give shifts of 0.029 V (basal), 0.019 V (apical) and 0.028 V (count-weighted pooled means), so 0.010 V is used to admit the paper's own numbers. Pigmented cells of Table 2 rows (a) and (b). Model of the effect test open, as in table1-xe991-no-effect-alpha; readings (x-a) and (x-b). The shift is read as (h-a) the difference of region-matched means averaged over basal and apical (Table 2 gives 0.029 and 0.019 V), or (h-b) the XE991 coefficient of the two-way model. Reproduced when under one reading the XE991 effect has p < 0.05 and the shift is within tolerance; partially reproduced when p < 0.05 with the shift outside tolerance. "Did not affect the other parameters" is judged on the same model for QTm, CLW and alphaB at p >= 0.05 and reported.
- table2-pigmented-xe991
With XE991, V0.5 of pigmented cells is more negative at the base (-0.062 V, 4 cells) than at the apex (-0.035 V, 11 cells), P = 0.0399.
How it will be judged
Tolerances are the standard deviations Table 2 prints. One-way ANOVA of corrected V0.5 on region (Table 2 footnote, "The P value of the ANOVA is shown for each group"). Reproduced when under one region reading the means are within tolerance and p < 0.05.
- table2-pigmented-no-xe991
Without XE991, V0.5 of pigmented cells is more negative at the base (-0.091 V, 12 cells) than at the apex (-0.054 V, 16 cells), P = 0.0003.
How it will be judged
As table2-pigmented-xe991. Reproduced when under one region reading the means are within tolerance and p < 0.001 (the reported p is 0.0003); partially reproduced when p < 0.05.
- table2-albino
In albino cells V0.5 at the base (-0.071 V, 11 cells) and apex (-0.057 V, 30 cells) do not differ significantly (P = 0.0697).
How it will be judged
Judged on the paper's test, a one-way ANOVA on region at p < 0.05. Reproduced when the means are within tolerance and p >= 0.05 under the region reading closest to the counts 11 and 30; the p-value and difference are reported (the reported p, 0.0697, is near the threshold, so both region readings are shown).
- fig3a-eq3a-fit
Binned by lateral membrane capacitance (5 pF), V0.5 falls as the charge density dVmin_exp rises, as Eq. 3a predicts; the one-parameter fit gives Vd 0.033 V (0.034 V in the legend, adjusted R2 0.916), and an unconstrained linear fit has slope -0.401 (adjusted R2 0.95).
We determined the pseudo-minimum at the depolarizing voltage, V d to be 0.033 V and determined an unconstrained linear fit of −0.401 (fit not shown; adjusted R 2 = 0.95)
Figure 3a; Results, Comparison of the experimental results with Eq. 3a, paragraph 1How it will be judged
Method from Results, Comparison with Eq. 3a, paragraph 1, and the Figure 3a legend. Cells without XE991, both colonies pooled; the chloride is not stated, and the 144 mM cells are used (the 10 mM cells are shown apart in Fig. 4). Cells grouped by CLW in 5 pF bins; per bin the mean dVmin_exp (QTm / CLW) and mean corrected V0.5. Fit of V0.5 = Vd - dVmin_exp / 2 (Eq. 3a as rebuilt) with Vd the only free parameter, weighted by the number of cells per bin (legend); an unconstrained linear fit, (u-a) weighted the same way, (u-b) unweighted (the legend states the weights for the Eq. 3a fits only). Open, (e-a) bin edges at multiples of 5 pF from 0; (e-b) bins starting at the smallest CLW; and whether the adjusted R2 is computed (w-a) with or (w-b) without the weights. Vd is 0.033 V in the text and 0.034 V in the legend; both fall within the tolerance of 0.034. Reproduced when under one reading all four values are within tolerance; partially reproduced when Vd and the slope are but an R2 is not.
- fig3a-xe991-fit
Cells with XE991 follow Eq. 3a with Vd 0.05 V (0.0499 V, adjusted R2 0.793), and an unconstrained linear fit of slope -0.37 (adjusted R2 0.87).
Also shown on the plot are the data determined in the presence of XE991, where V d is at 0.05 V with an unconstrained linear fit of −0.37 (fit not shown; adjusted R 2 = 0.87)
Figure 3a legend; Results, Comparison of the experimental results with Eq. 3a, paragraph 1How it will be judged
As fig3a-eq3a-fit, for the cells with XE991 (the legend says 60 uM; the Methods give 30 to 60 uM; readings (k-a) all XE991 cells, (k-b) 60 uM only, if the concentration is recorded). Reproduced when under one reading all four values are within tolerance; partially reproduced when Vd and the slope are.
- fig3a-high-density-deviation
Below a charge density of 0.25 V the data lie close to the Eq. 3a line, while above it V0.5 is larger than expected and more variable.
When Δ V m i n e x p < 0.25 V, the experimental data lies close to the central tendency of the theoretical line, but when Δ V m i n e x p > 0.25 V, V 0.5 is larger than expected and exhibits greater variance
Results, Comparison of the experimental results with Eq. 3a, paragraph 1; Figure S4How it will be judged
Cells of fig3a-eq3a-fit, with the fitted Vd. Per cell the residual V0.5 - (Vd - dVmin_exp / 2). No test is given; reading used, the cells with dVmin_exp > 0.25 V have a mean residual above zero (one-sample t-test, p < 0.05) and a larger residual variance than cells below 0.25 V (F-test, p < 0.05), and the mean residual below 0.25 V is smaller in magnitude than above. The sentence is about the binned data of Fig. 3a and Fig. S4; the same three conditions are also computed on the 0.01 V bins of Fig. S4 (bin means, and spread of bin means) as a reading. Reproduced when all three hold under one reading; partially reproduced when the mean residual holds but not the variance. Rests on tests the paper does not name.
- figs4-fine-bins
With cells binned by charge density in 0.01 V bins (dVmin_exp < 0.4 V fitted), Eq. 3a gives Vd 0.030 V (adjusted R2 0.389) without XE991 and Vd 0.054 V (adjusted R2 0.737) with XE991.
Data were averaged based on the value of Δ V min exp with a bin width of 0.01 V. The bars are SEM. The dashed blue line was fit to Eq. 3a , where Δ V min exp < 0.4 V was included in the fit. V d is 0.030 V; adjusted R 2 = 0.389. The dashed red line was fit to Eq. 3a with V d of 0.054 V; adjusted R 2 = 0.737
Figure S4 legendHow it will be judged
Cells as fig3a-eq3a-fit and fig3a-xe991-fit. Bins of dVmin_exp 0.01 V wide (edges at multiples of 0.01 V, a standard choice); per bin means; fit of Eq. 3a with Vd free over bins below 0.4 V. Whether the fit is weighted by count is not stated for Fig. S4; readings (w-1) weighted by count as in Fig. 3a, (w-2) unweighted. Whether the 0.4 V limit also applies to the XE991 fit is not stated; readings with and without. Reproduced when under one reading all four values are within tolerance; partially reproduced when both Vd are.
- tables3-by-region
Similar trends of V0.5 with charge density are found when the charge density is computed per cell and averaged by cochlear region.
Similar trends are found when Δ V m i n e x p is calculated per cell and aggregated based on the positional polarity of the cell in the cochlea ( Table S3 )
Results, Comparison of the experimental results with Eq. 3a, paragraph 1; Table S3How it will be judged
Table S3's values are not in the supplied text, so no number can be compared. Per cell dVmin_exp; mean dVmin_exp and mean V0.5 per region (basal, apical by readings r-a and r-b) and group (pigmented with XE991, pigmented without, albino, as the Table S3 caption lists). Reproduced when in every group the region with the higher mean dVmin_exp has the more negative mean V0.5. Table S3 is to be supplied to compare values.
- fig3a-other-rodents
Values reported for young albino rats and for wild type mice from five studies lie on the guinea pig relation of V0.5 with charge density, four of the five mouse values within the range of high-frequency guinea pig cells.
Four of the five values lie within the data observed for the high-frequency guinea pig cells, with one (red square) exhibiting a value that is more hyperpolarized for the reported Δ V m i n e x p . This shows that V 0.5 is related to Δ V m i n e x p , as predicted by Eq. 3a for OHCs from guinea pigs, rats, and mice
Figure 3a; Results, Comparison of the experimental results with Eq. 3a, paragraph 2How it will be judged
Outside inputs, the values of V0.5, QTm and cell length and radius reported by Mahendrasingam et al. (2010), Cheatham et al. (2005), Homma et al. (2013), Yamashita et al. (2012) and Zhu et al. (2013, 2015), with CLW = 2 pi r (l - r) x 0.01 pF/um2 and the radii and lengths of the Fig. 3a legend; for the rats the mean of that CLW and the Eq. 8a value (8.9 and 3.3 pF; 9.56 and 3.55 pF). Reproduced when four of the five mouse points have dVmin_exp and V0.5 within the range spanned by the guinea pig bins of CLW < 10 pF, and the rat points lie within 0.02 V of the guinea pig Eq. 3a line (a tolerance chosen here, as the paper gives none).
- fig3c-engineered-mice
In engineered mice, neo/neo and neo/prestin KO OHCs have 35% and 57% lower charge density with little change of V0.5, 499 KI OHCs a tenfold lower charge density with a large positive V0.5, and C1 KI OHCs a wild-type charge density with a negative V0.5.
In these cases, the Δ V m i n e x p is 57% (neo/prestin KO) and 35% (neo-neo mouse) lower, while the V 0.5 is slightly depolarized or about the same for both cases
Figure 3c; Results, Comparison of the experimental results from engineered mice with Eq. 3a, paragraph 2How it will be judged
Rests wholly on values from other papers (Zhu et al. 2013, 2015; Yamashita et al. 2012; Homma et al. 2013; Gao et al. 2007; Cheatham et al. 2005), with the radii and lengths of the Fig. 3c legend; the data set holds none of it. Reproduced when the percentages computed from those values are within 10% of 57% and 35%.
- capacitance-asymmetry
Linear capacitance at hyperpolarized voltages exceeds that at positive voltages by 2.76 +/- 1.32 pF (62 cells); excluding differences above 5 pF, the difference is 2.87 pF for apical and 2.53 pF for basal cells, a 13% and 21% increase in CLW.
For our data, the membrane capacitance observed in the hyperpolarized region is 2.76 ± 1.32 pF (number of observations, 62) more than the capacitance measured at positive voltages ( Fig. S5 ). ... When Δ C > 5 pF, we discounted the measurements and report a Δ C of 2.87 (±1.27) and 2.53 (±1.47) pF for cells from apical and basal regions. This amounts to a 13% (apical) and 21% (basal) increase in the linear capacitance of the lateral membrane
Results, Asymmetry of C versus V function, paragraph 1; Figure S5How it will be judged
Tolerances for the differences are the stated standard deviations, for the percentages 10%. dC = CLn - CL per cell, for cells where CLn could be estimated (the capacitance reached its hyperpolarized plateau); how CLn is taken is not given beyond that, open (stored value, or by the CL method applied at negative voltages). The percentage is read as (m-a) the mean of dC / CLW per cell; (m-b) the ratio of the mean dC to the mean CLW. Reproduced when under one reading all values are within tolerance and the count is 62 (+/- 3) cells.
- figs5-specific-capacitance
The specific capacitance of the lateral membrane is 0.0095 pF/um2 at positive voltages (intercept 2.14 pF, adjusted R2 0.79, 83 cells) and 0.01106 pF/um2 at negative voltages (fit with the same intercept, adjusted R2 0.807, 59 cells).
The solid line is fitted to data where capacitance was determined at positive voltages with slope 0.0095 pF/μm 2 and intercept 2.14 pF; adjusted R 2 = 0.79 (83 cells). The dashed line is fit-constrained to the same intercept with slope 0.011062 pF/μm 2 ; adjusted R 2 = 0.807
Figure S5 legend; Results, Asymmetry of C versus V function, paragraph 1How it will be judged
Linear least-squares fits of CL (83 cells) and CLn (59 cells) on cell surface area, the second with the intercept fixed at the first's. Tolerance of the negative slope is the stated spread (0.01106 +/- 0.0016 pF/um2, Results). The surface area is from the bright-field image (Corbitt et al. 2012); its formula is not given here; readings (a-a) a stored area; (a-b) 2 pi r (l - r) from length and radius as used for mice in Fig. 3a; (a-c) the total cylinder surface 2 pi r l. Reproduced when under one reading all five values are within tolerance; partially reproduced when both slopes are.
- fig3b-negative-clw-fit
With CLW taken at negative potentials (44 cells, no XE991), Eq. 3a fits with an intercept of 0.02 V (adjusted R2 0.91), while an unconstrained linear fit has slope -0.664 and intercept 0.046 V (adjusted R2 0.97).
The dashed blue line is the fit to Eq. 3a with an intercept at 0.02 V (adjusted R 2 , 0.91). The dashed black line is the linear fit of data with a slope of −0.664 and an intercept at 0.046 V (adjusted R 2 , 0.97)
Figure 3b; Results, Asymmetry of C versus V function, paragraph 1How it will be judged
CLWn = CLW + dC per cell (Results, Asymmetry, paragraph 1), for the 44 cells without XE991 that have CLn (legend). dVmin^n = QTm / CLWn. "We partitioned the data making use of CLWn" is read as binning by CLWn in 5 pF bins as Fig. 3a (bin and weighting readings of fig3a-eq3a-fit), and whether cells with dC > 5 pF are kept is open (readings kept, dropped). Reproduced when under one reading all five values are within tolerance; partially reproduced when the Vd and slope are.
- mean-slope-half
The mean of the three unconstrained slopes for guinea pig cells (-0.4, -0.37, -0.66) is about -0.48, close to the -1/2 Eq. 3a predicts for more than one hyperpolarized state.
Therefore, we conclude upon fitting the guinea pig data unconstrained to a linear function ( Fig. 3 ) that the mean value for all unconstrained slopes is approximately −0.48 (i.e., mean of slopes −0.4, −0.37, and −0.66) which is approximately −0.5
Results, Asymmetry of C versus V function, paragraph 1How it will be judged
The mean of the three slopes found in the claims it depends on. Reproduced when within tolerance.
- fig4a-vh-rises-vd-constant
The hyperpolarized pseudo-minimum Vh increases monotonically by threefold across cells while the depolarized pseudo-minimum Vd stays about constant, evidence for more than one hyperpolarized state.
We calculated with Eq. 3 V h and V d for prestin sensors on the lateral membrane of rodents and found that V h increases monotonically by threefold while V d remains approximately constant ( Fig. 4 a )
Figure 4a; Results, More than one configurational state of the prestin sensor when hyperpolarized, paragraph 1How it will be judged
Per bin of Fig. 3a (with and without XE991, legend of Fig. 4a), Vh = V0.5 - dVmin_exp / 2 and Vd = V0.5 + dVmin_exp / 2 (Eq. 3 as rebuilt). "Increases monotonically" is judged by a Spearman correlation of Vh with V0.5 (or with CLW) of +1 allowing at most one inversion between neighbouring bins; "threefold" by the ratio of largest to smallest magnitude of Vh. The paper fixes no measure of "approximately constant"; readings (q-a) the range of Vd across bins is less than one third of the range of Vh; (q-b) a weighted linear fit of Vd on V0.5 has a slope not different from zero at p < 0.05. Fig. 4a plots the bins with and without XE991 together, and the text does not say over which set the threefold is taken; readings (o-a) the bins without XE991; (o-b) all bins pooled. Reproduced when the Vh conditions hold and Vd is constant under one reading.
- fig4d-cleq-two-limits
The capacitance of the membrane where one sensor resides, C_Leq = n e0 / dVmin_exp, has two limits, 0.584 (+/- 0.065) aF at mean V0.5 below -0.06 V, rising almost exactly twofold to 1.07 (+/- 0.038) aF as V0.5 rises to -0.03 V.
Likewise, C L , e q is calculated with n e o / Δ V m i n e x p and found to exhibit two limits. The first is 0.584 (±0.065) aF, which is apparent at a mean V 0.5 less than −0.06 V and increases monotonically to 1.07 (±0.038) aF as V 0.5 increases to −0.03 V ( Fig. 4 d ). This change in the capacitance is almost exactly twofold
Figure 4d; Results, Determine q, Amean, C_Leq and V0.5, paragraph 2; Table 3How it will be judged
Tolerances are the stated spreads. With delta = 1, n = z = alphaB kB T / e0 (Fig. 4d legend, "hence z = n"). Per group of cells, C_Leq = z e0 / dVmin_exp from the group means. The groups are "the data obtained with linear capacitance measured at positive potentials (Fig. 3 a and Fig. S4)", read as (j-a) the 5 pF CLW bins of Fig. 3a; (j-b) the 0.01 V bins of Fig. S4; (j-c) per cell; each pooling cells with and without XE991 (Table 3 and Fig. 4f pool them). The split of groups between the limits is taken from Table 3 and Discussion, Kinetic scheme, paragraph 2, which give the V0.5 range of each pathway, -0.1 to -0.056 V for the 0.584 aF limit and -0.043 to -0.013 V for the 1.07 aF limit; the lower limit is the mean over groups with V0.5 at or below -0.056 V, the upper over groups at or above -0.043 V, and groups between are left out and shown (the text's "less than -0.06 V" is shown as a variant split). Fig. 4d plots the rat and mouse values of Fig. 3a with the guinea pig values, and Table 3 is for all wild type, so the paper's limits may include them; the data set gives the guinea pig groups only, and this is noted. Reproduced when under one grouping both values are within tolerance and C_Leq rises with V0.5 (Spearman, p < 0.05, a test chosen here).
- fig4c-sensor-area
The limiting membrane area of one wild type sensor is 51 to 121 nm2 (from dVmin_exp and a specific capacitance of 0.0095 pF/um2), with |V0.5| falling as a power of the area, |V0.5| = 38.5 Amean^-1.15 in the Fig. 4c legend (exponent -1.5 in the text).
We found for our experimental data that the limiting membrane area for the wild type voltage sensor was 51 to 121 nm 2 , which we calculated from the experimental values of Δ V m i n e x p and a specific capacitance of 0.0095 pF/μm 2 ( Fig. S5 )
Figure 4c; Results, Determine q, Amean, C_Leq and V0.5, paragraph 1How it will be judged
Amean = C_Leq / 0.0095 pF/um2 per group, with C_Leq of fig4d-cleq-two-limits (same groupings). Range over groups of guinea pig cells (the paper's range may include rats and mice, so the claim holds when the guinea pig range lies within 51 to 121 nm2 widened by 10%, and the end values are shown). Power law fitted to guinea pig groups without XE991 (open blue circles, legend), units of |V0.5| and Amean not given (readings mV with nm2, V with nm2). The exponent is -1.15 in the legend and -1.5 in the text; the claim holds if the fitted exponent is within 10% of either. The paper's own numbers favour -1.5 with V and nm2 (38.5 x 61.5^-1.5 = 0.080 V and 38.5 x 112.6^-1.5 = 0.032 V, against Table 3's -0.076 and -0.031 V, with Amean from 0.584 and 1.07 aF); -1.15 gives 0.34 and 0.17 V. Reproduced when the range and one exponent reading hold.
- fig4e-sensor-charge
The charge moved per sensor is similar for all sensors, 0.873 +/- 0.0577 e0 at the 0.584 aF limit and 0.872 +/- 0.083 e0 at the 1.07 aF limit, 0.859 +/- 0.066 e0 over all wild type sensors.
The magnitude of the charge is similar for all sensors at 0.873 ± 0.0577 (maximum 0.952; minimum 0.767; C L , e q 0.584 aF) and 0.872 ± 0.083 (maximum 0.95; minimum 0.756; C L , e q 1.07 aF) with a range for all wild type at 0.859 ± 0.066 (maximum 0.952; minimum 0.746)
Figure 4e; Results, Determine q, Amean, C_Leq and V0.5, paragraph 2; Table 3How it will be judged
Tolerances are the stated spreads. z per group, as in fig4d-cleq-two-limits and with its groupings and limit split. The "all wild type" value includes rat and mouse values from other papers; for the data set the guinea pig groups alone are used and compared, and the difference is noted. Reproduced when the first two values are within tolerance and do not differ (Welch's t-test p >= 0.05, a test chosen here as the paper names none).
- table3-pathway-energies
Pathway 1 (K1, K2) has C_Leq 0.584 aF, V0.5 -0.076 V and z 0.859, giving a reorganization energy of 2 x 3.97 kB T and barriers of 2 x 2.68 kB T (forward) and 2 x 0.125 kB T (reverse); pathway 2 (K3, K4) has C_Leq 1.07 aF, V0.5 -0.031 V, a reorganization energy of 2.16 kB T and barriers of 1.19 (forward) and 0.145 kB T (reverse).
The V 0.5 associated with pathway 1 ( K 1 and K 2 ) is −0.076 ± 0.014 (minimum −0.1; maximum −0.056) V. We calculate the activation energy barrier Δ G § at 2.68 κ B T /monomer ... Likewise, the V 0.5 associated with K 3 and K 4 is −0.031 V (± 0.007; minimum −0.043; maximum −0.013), where we calculate Δ G § of 1.2 κ B T /dimer (0.7 kcal/mol of dimers) and reorganization energy at 2.16 κ B T /dimer
Table 3; Discussion, Kinetic scheme, paragraph 2How it will be judged
V0.5 tolerances are the stated spreads, the rest 10%. V0.5 per pathway is the mean V0.5 of the groups at each C_Leq limit (fig4d-cleq-two-limits). lambda = q^2 / (2 C_Leq) (Eq. 6a as rebuilt) with q = z e0, z being the mean over all groups for both pathways, as Table 3 gives 0.859 for each; the forward (activation) barrier (lambda - q V0.5 / kB T)^2 / (4 lambda) and the reverse (lambda + q V0.5 / kB T)^2 / (4 lambda) (Eq. 7 as rebuilt), at 297.15 K; the rebuilt forms give Table 3's values from Table 3's inputs, which is checked first and shown. Reproduced when all six values computed from the data are within tolerance; partially reproduced when the V0.5 values are but a derived energy is not. The text's 3.95 and the table's 3.97 kB T both fall within tolerance. Rate ratios (pathway 2 forward 64 times faster than pathway 1, e^-1.2 / e^-5.36; Discussion, Physiological implications) and the chemical free energy of 6.7 kB T follow from these and are reported.
- alternative-limit
Under the alternative limit of constant C_Leq (1.098 aF), 1.7 charges (1.44 to 1.92) would move a fraction delta = 0.535 (0.432 to 0.63) of the field, which the authors judge less logical than delta = 1 with C_Leq changing.
finds 1.7 charges [maximum 1.92, minimum 1.44] moving a fraction of the distance, δ = 0.535 [maximum 0.63, minimum 0.432]
Results, Determine q, Amean, C_Leq and V0.5, paragraph 3How it will be judged
Per group (groupings of fig4d-cleq-two-limits), n = dVmin_exp x 1.098 aF / e0 and delta = z / n (Results, paragraph 3; the text prints "[n/z]", but z = n delta by Eq. 5, and 1.7 x 0.535 = 0.91 is a z, so z / n is the reading). Reproduced when the means are within tolerance under one grouping and the ranges lie within 1.44 to 1.92 and 0.432 to 0.63 widened by 10%.
- fig4f-all-rodents
Charge density against V0.5 for the sensors of all rodents, with heterozygote density corrected, fits Eq. 3a with Vd 0.037 V (adjusted R2 0.538), regardless of bath composition.
The dashed line is the fit of data from all rodents with and without XE991 in the bath to Eq. 3a with intercept, V d 0.037 V (adjusted R 2 , 0.538)
Figure 4f legend; Results, Determine q, Amean, C_Leq and V0.5, paragraph 3How it will be judged
Guinea pig groups from the data set (with and without XE991) plus the rat and mouse values of fig3a-other-rodents and the heterozygotes of Fig. 3c with density corrected as Results, paragraph 2 says (C_Leq times the ratio of heterozygote to wild type density), leaving out 499 KI, heterozygote 499, C1 KI and neo/prestin KO. Fit of Eq. 3a with Vd free; weighting open (by count, unweighted). Reproduced when both values are within tolerance under one reading.
- low-chloride-similar
With 10 mM intracellular chloride and glutamate, results are similar to those with high intracellular chloride.
When we performed experiments in 10 mM intracellular chloride with coion glutamate, results were similar to those observed at high concentrations of intracellular chloride (magenta circles in Fig. 4, d–f )
Discussion, Kinetic scheme, paragraph 2; Figure 4d to f (open magenta circles)How it will be judged
The three cells with 10 mM chloride (pigmented, XE991; Fig. 4 legend). Per cell z, dVmin_exp, C_Leq and V0.5. No test is given (three cells); reading used, each cell's C_Leq lies within the range of the 144 mM chloride groups (0.584 - 0.065 to 1.07 + 0.038 aF), its z within 0.746 to 0.952, and its V0.5 within 0.02 V of the Eq. 3a line fitted to the XE991 cells. Reproduced when all three cells meet all three; partially reproduced for two cells.
- molecular-model
The I-TASSER model of the guinea pig prestin transmembrane domain has radii of 3.4 and 3.15 nm and a maximum area of about 36.3 nm2, slightly larger than SLC26Dg (34.2 nm2), smaller than the area from electrophysiology; the STAS domain is strongly negative (-16.3 or -14 charges).
The maximum area of this domain is ∼36.3 nm 2 , slightly larger than that obtained for the fumarate bacterial transporter, SLC26Dg ( Geertsma et al., 2015 ), at 34.2 nm 2 ( Fig. 4 c )
Figure 4b; Results, Determine q, Amean, C_Leq and V0.5, paragraph 1; Discussion, Kinetic scheme, paragraph 3; Table S4How it will be judged
Rests on a structural model (I-TASSER, templates PDB 5DA0 and 3LLO; PROPKA 3.0 on PDB2PQR; OPM), not on the data set. Reproduced only if the model is rebuilt; the area is then compared with 10% tolerance. Table S4's charges are not in the supplied text.
- figs7-hearing-loss
Hearing loss at 32 kHz in engineered mice correlates with charge density measured ex vivo (best linear fit slope -0.0051 V/dB, intercept 0.2343 V).
The loss at 32 kHz relative to measured experimental charge density. The dashed line represents the best linear fit with a slope of −0.0051 V/dB and intercept 0.2343 V
Figure S7 legendHow it will be judged
Rests wholly on values reported in other papers (Dallos et al. 2008; Yamashita et al. 2012; Gao et al. 2007; Zhu et al. 2013); the data set holds none of them. Reproduced when a least-squares fit of charge density on loss gives slope and intercept within 10%.
- v05-temperature-corrected
After correction to physiological temperature, ex vivo V0.5 of basal cells is about -72 mV (range -47 to -105 mV), comparable with in vivo resting potentials, while turn 3 cells are centred at about -40 mV.
are comparable with our ex vivo measurements of V 0.5 at −72 mV (range −47 to −105 mV) after correcting to physiological temperature ( Meltzer and Santos-Sacchi, 2001 ; Fig. 4 d , bottom arrow)
Discussion, Caveats associated with the model and data, paragraph 3How it will be judged
The temperature correction is taken from Meltzer and Santos-Sacchi (2001), an outside input the paper does not state numerically, applied from 24 C to 37 C (the temperature of the in vivo arrows, Fig. 3c legend). "Basal" by readings r-a and r-b; the cells by readings v-a and v-b of v05-by-turn. The central tendency is read as (c-1) mean, (c-2) median. The turn 3 tolerance is the paper's stated spread, "-40 (± 0.022) mV", read as 0.022 V (a spread of 0.022 mV is implausible beside the other V0.5 spreads of 0.018 to 0.030 V). Reproduced when both values are within tolerance under one reading, and the basal range lies within -47 to -105 mV widened by 10%.
- recording-time
Recordings were made 47 to 182 min after sacrifice (mean 125 +/- 32 min pigmented, 112 +/- 36 min albino), and the electrical parameters do not correlate with recording time.
However, we did not find a correlation with the magnitude of the electrical parameters and time of the recording relative to the lifetime boundary of the animal ( Table S1 )
Materials and methods, Animals and OHC isolation, paragraph 4; Discussion, Caveats, paragraph 4; Table S1How it will be judged
Tolerances are the stated standard deviations. Table S1's contents are not in the supplied text, so its test is unknown. A claim of absence with no test given; reading used, Pearson correlation at p < 0.05 of each of QTm, CLW, V0.5, alphaB and dVmin_exp with recording time, per colony. Reproduced when the means are within tolerance and none of the ten correlations is significant (Bonferroni-corrected as a variant). Rests on a test the paper does not name.
- two-hyperpolarized-states
The relation of V0.5 to charge density, with slope near -1/2, Vd constant and Vh varying, shows that the prestin sensor has two hyperpolarized states and two charge-displacement pathways in the lateral membrane.
We show that a plot of V 0.5 as a function of Δ V m i n e x p has a negative slope, with an approximate value of one half for OHCs from rodents. This is evidence for more than one hyperpolarized state of the sensor
Theoretical model, the paragraph after Eq. 3; Discussion, Kinetic scheme, paragraph 1; Figure 5How it will be judged
The paper's interpretive conclusion. Reproduced when the claims it depends on are reproduced (the mean slope is within 10% of -0.48, Vd constant and Vh rising, and C_Leq shows two limits); partially reproduced when the slope and Vh conditions hold but C_Leq does not show two limits; not reproduced when the unconstrained slope is not negative or Vd is not constant.
Pipeline
- Paper
- waiting (not checked)
- Claims
- done
- Claims check
- done (ready)
- Attempt
- todo
- Questions
- none
- Verified
- none