Block of the muscle nAChR by TMA⁺. (A) Macroscopic current response to a 2-s pulse of 50 mM TMA⁺ recorded at –80 mV from the mouse muscle adult-type nAChR in the outside-out configuration. Because TMA⁺ is both a pore blocker and a (desensitizing) agonist of the nAChR, TMA⁺ was applied only transiently rather than continuously (compare with Fig. 9, C and D). The solutions flowing through the two barrels of the perfusion tubing were (in mM) 142 KCl, 5.4 NaCl, 1.8 CaCl₂, 1.7 MgCl₂, and 10 HEPES/KOH, pH 7.4, with or without TMA⁺. In the schematic representation of the theta-tubing perfusion, the arrow indicates the application of TMA⁺. (B) Macroscopic current responses to a 1-ms pulse of 100-µM ACh or 50 mM TMA⁺ recorded at –80 mV in the outside-out configuration. The solutions flowing through the two barrels of the perfusion tubing were as described in A with or without ACh and with or without TMA⁺. The two traces are averages of 10 consecutive responses recorded from two separate representative patches and were normalized for displaying purposes in such a way that the current values attained upon ligand removal are the same; this emphasizes the block by TMA⁺ (see red asterisk). In the schematic representation of the theta-tubing perfusion, arrows indicate the application of ACh or TMA⁺. (C) Single-channel inward currents elicited by 0.5 mM or 50 mM TMA⁺ recorded at approximately –100 mV in the cell-attached configuration; openings are downward deflections. The pipette solution was (in mM) 142 KCl, 5.4 NaCl, 1.8 CaCl₂, 1.7 MgCl₂, 0.5 or 50 TMA⁺, and 10 HEPES/KOH, pH 7.4. The extent of block caused by TMA⁺ seems to be larger in the single-channel recordings than in the macroscopic recordings. Undoubtedly, this is due to the fast deactivation time constant of the nAChR, which leads to the underestimation of the true peak value of the macroscopic transient upon TMA⁺ removal.