Figure 2.

Single channel properties of PIEZO1 V598M transiently expressed in HEK-293 cells. (A) Representative cell-attached patch from a cell transiently transfected 24 hours previously with cDNA encoding human PIEZO1 V598M. Channels opened predominantly during the pressure pulse and occasionally during the interpulse intervals (red boxes) with only residual intrinsic stress remaining in the patch. (B) Responses of the same cell-attached patch to 3 subsequent pulses of prolonged stimulation (50 seconds). The time-resolved inset for pulse 2 shows a subconductance level of 1 to 1.5 pA (red box). Holding potential in A and B was −60 mV. (C) Left, amplitude histogram with predominant amplitude at 2.5 pA, corresponding to trace of panel A; right, amplitude histogram with predominant amplitude of 3.06 pA, corresponding to traces of panel B. (D) Representative cell-attached patch at holding potential of −60 mV subjected to −50 mm Hg pressure step of 1 second duration, showing rapid activation of multiple channels with slow, partial inactivation to a lower steady-state activity level. Panels A–D are each representative of similar recordings from 3 cells. (E) Relationship between applied negative pressure (conditions as in panel D) and normalized magnitude of charge transferred during the entire pressure pulse, as fit with the Boltzmann equation for 3 cells. For all panels, bath (in mM) was 150 KCl, 1 CaCl₂, 1 MgCl₂, 10 HEPES, pH 7.4; pipette solution (in mM) was 150 KCl, 10 HEPES, pH 7.4. Additional single channel recording conditions were as described.^10.