Decay and recovery of TRPM3 activity in inside-out membrane patches. (A) Time course of TRPM3 currents at −120 and +120 mV in cell-attached mode and, after patch-excision, in cell-free inside-out patch from a HEK-M3 cell. TRPM3 was activated by inclusion of 100 µM PregS in the extracellular (pipette) solution. The vertical dotted line in this and subsequent panels indicates the time point of patch excision. Membrane patches were always excised into intracellular solution. (B) I-V traces at different time points as indicated in A. (C) Representative time course showing the effect of diC8 PI(4,5)P₂ on TRPM3 currents measured in inside-out configuration. (D) I-V traces at different time points as indicated in C. (E) Representative time course showing the effect of 2 mM ATP on TRPM3 currents measured in inside-out configuration. (F) I-V traces at different time points as indicated in E. (G) Representative time course showing the effect of 2 mM ATP on TRPM3 current upon excision. (H) Representative time course comparing the effects of diC8 and brain derived natural PI(4,5)P₂ applied to the cytoplasmic side of an inside-out excised membrane patch. (I) Maximal current recovery evoked by PI(4,5)P₂ and ATP. Currents were normalized to the peak current upon excision. Numbers of individual membrane patches are indicated in parentheses.