Skip to main content
. 2016 Mar 16;43(8):1062–1074. doi: 10.1111/ejn.13209

Figure 4.

Figure 4

The 9‐AC block is chloride‐sensitive and 9‐AC can access from both sides of the plasma membrane. (A) Dose dependency of the extracellularly applied 9‐AC on the maximum amount of motor charge moved, Q max, measured with extracellular 9‐AC concentrations of 0.05, 0.5, 5, 50, 500 and 5000 μm. The effect on Q max is presented as relative reduction. Circles labelled ‘CsCl’, the control condition with intracellular solution containing 135 mm CsCl. Triangles labelled ‘Cs2 SO 4’, intracellular chloride concentration reduced by substituting 115 mm chloride with sulfate in the patch pipette. Squares labelled ‘redCl’, both intracellular and extracellular chloride concentrations reduced by substituting 130 mm CsCl with gluconate in both the patch‐pipette and bath solutions. This latter condition significantly shifted the dose dependency to the left. Lines indicate logistic‐function fits (Eqn (3)) with estimates of the IC 50 and the slope parameter, respectively, of 928 ± 66, 1012 ± 192, 79 ± 14 μm and 0.82 ± 0.04, 0.56 ± 0.06, 0.71 ± 0.08 for the three conditions. (B) The relative reduction in Q max in response to extracellular application of 500 μm 9‐AC, using the indicated intracellular solutions, patch configuration, drugs and cell types. Labelling on the abscissae has the following meaning. ‘CsCl’ (n = 11 for OHCs, n = 11 for HEK 293 cells), ‘Cs2 SO 4’ (n = 5), and ‘redCl’ (n = 4), as in panel A. ‘9‐AC i’, 500 μm 9‐AC in the patch solution (n = 5 for OHCs, n = 5 for HEK 293 cells). ‘InOut’, inside‐out excised‐patch configuration (n = 3); all other measurements were made in the whole‐cell configuration. Data for the bar labelled ‘OHC/CsCl’ are the same as those for the bar labelled ‘Control’ in Fig. 2D. Notice for OHCs, only intracellular and extracellular chloride substitution with gluconate (redCl; n = 4) increased the relative reduction in Q max compared with CsCl control. That is, the blocking efficacy of extracellular 9‐AC was increased by lowering the OHC [Cl−] gradient by reducing the intra‐ and extracellular [Cl−]. Notice for HEK 293 cells, the addition of 9‐AC to the cytosol (9‐AC i; n = 5) decreased the blocking efficacy of extracellularly applied 9‐AC.