SPD model fitting to experimentally observed TRPC6/7 currents and AVP-evoked TRPC6/7-like currents in A7r5 cells. (A) Fitting of the receptor-operated TRPC6 current simultaneously measured with PI(4,5)P₂ by FRET with the SPD model (top; light blue trace). The experimental data were obtained from the low strength of receptor stimulation carried by CCh application (100 µM) to endogenous muscarinic receptor in HEK293 cells. Back-calculated FR of PI(4,5)P₂ concentrations at the respective points (bottom; solid) was normalized against the time zero FR (Norm.FR; middle; light blue), and that was overlaid on the experimental Norm.FR (middle; circles). The bottom panel displays the fitting resultant changes in PI(4,5)P₂ (solid), DAG (dashed), and PA (thin dashed). The similarity of Norm.FR was assessed by SD as described in Materials and methods. (B) Fitting of experimental TRPC6 current data from M₁R-overexpressing cells. The initial parameters are identical to those in A. (C) Fitting of experimental TRPC6/7-like currents recorded from A7r5 aorta–derived smooth muscle cells. The currents were evoked by 1 µM AVP. The inset in the top panel shows A7r5 cells expressing the PI(4,5)P₂ sensor. (D) Fitting of TRPC7 currents. Local PI(4,5)P₂ dynamics were detected using a FRET donor linked to the TRPC7 channel at the end of its C-terminal domain (top; inset). Transient increments in FRET were detected during the plateau or biphasic response (middle; arrow).