Fig. 3.

Bicarbonate does not activate other membrane GCs or sGCs. (A) Bicarbonate did not stimulate the cyclase activity of cells expressing GC-A, GC-C, GC-E, or GC-F. It appeared to inhibit the basal cyclase activity of GC-E and GC-F. GC-C was assayed on T84 cells, whereas other GCs were transiently expressed in HEK-293T cells. Numbers 0, 25, and 50 in leftmost panel indicate NaHCO₃ concentrations and apply to all panels. *, P < 0.05; ***, P < 0.001; t test. (B–D) Ca²⁺ imaging reviewed that CO₂ responses of GC-D⁺ neurons were independent on the activity of sGCs. (B) GFP labeling of GC-D⁺ neurons within an intact epithelial preparation from a GCD-ITG mouse (Left) and loading of calcium-sensitive Rhod-2/AM dye into GC-D⁺ neurons (Right). (C) Application of a selective sGC blocker, ODQ (50 μM), had no effect on CO₂-evoked Ca²⁺ signals. Arrows indicated the start of perfusion of CO₂ solution. (Scalle Bars, 5% ΔF/F and 50 s.) (D) Group data showing that the CO₂-evoked responses before and after ODQ application were not statistically different (P = 0.08; n = 27 cells; paired t test).