Differential Cl- and HCO3- mediated anion secretion by different colonic cell types in response to tetromethylpyrazine

Figure 1 TMP-induced I_SC response in T84 and Caco-2 cell lines. The concentration-response curve for TMP-induced response in T84 and Caco-2 cells, and each datum was obtained from at least 4 individual experiments. A: Values are mean ± SE of maximal I_SC increase; B: Representative I_SC recordings in response to apical addition of TMP (1 mmol/L) in T84 cells; C: Basolateral application of TMP (5 mmol/L) in Caco-2 cells.

Figure 2 Cl^- dependence of TMP-induced I_SC increase in T84 cells. Representative I_SC recording with arrows indicating TMP (1mmol/L) added apically, which was blocked by apical adding DPC. Values are mean ± SE; ^cP < 0.001. A: Normal; B: Cl^-free; C: K-HS, comparison of TMP-induced total charge transferred in normal and Cl^--free K-HS.

Figure 3 HCO₃^- dependence of TMP-induced I_SC increase in Caco-2 cells. Representative I_SC recording with arrows indicating TMP (5 mmol/L) added basolaterally, which was abolished by apical application of glybenclamide. Values are mean ± SE; ^aP < 0.05, ^bP < 0.01 and ^cP < 0.001. A: Normal; B: Cl-free; C: K-HS, comparison of TMP-induced total charges transferred in normal and Cl^--free K-HS.

Figure 4 Effect of inhibitors of basolateral anion transporters on TMP-induced I_SC responses. Values are mean ± SE; ^cP < 0.001. A: Comparison of TMP (1 mmol/L)-induced I_SC responses in T84 cells in the absence and presence of basolateral addition of bumetanide (100 μmol/L); B: Comparison of TMP (5 mmol/L) -induced I_SC responses in Caco-2 cells in the absence and presence of basolateral addition of inhibitors indicated.

Figure 5 TMP-induced anion secretions in rat colonic mucosa. Representative I_SC recording values are mean ± SE; ^aP < 0.05, ^bP < 0.01, ^cP < 0.001. A: 1 mmol/L; B: 5 mmol/L of TMP added basolaterally in normal K-HS; C: Comparison of TMP (1 and 5 mmol/L)-induced I_SC obtained in normal and Cl^--free as well as both of Cl^- and HCO₃^- -free solutions; D: Comparison of TMP (1 and 5 mmol/L)-induced I_SC obtained in basolateral pretreatment of colonic mucosa with bumetanide (100 µmol/L) and DIDS (100 µmol/L).

Figure 6 RT-PCR analysis of mRNA expression of Na⁺- HCO₃^- cotransporter (NBC) in colonic cells. A: RT-PCR results with products as expected of NBC found in rat colon (1), T84 (2) and Caco-2 (3) with positive control (4) using pancreatic duct cells (CAPAN-1) and negative control (5) where cDNA was omitted. B: Semi-quantitative analysis of NBC expression in Caco-2 (1) ; C: T84 (2) cells with NBC to GAPDH (internal marker) ratio shown.