Stimulation of cystic fibrosis transmembrane conductance regulator–dependent short- circuit currents across ΔF508 murine intestines  Wendy K. Steagall*, Mitchell L. Drumm*,‡  Gastroenterology  Volume 116, Issue 6, Pages 1379-1388 (June 1999) DOI: 10.1016/S0016-5085(99)70502-0 Copyright © 1999 American Gastroenterological Association Terms and Conditions

Fig. 1 Effect of PDE inhibitors on Isc in T84 cells. PDE inhibitor (100 μmol/L) or forskolin (10 μmol/L) was applied to cells in modified Ussing chambers and ΔIsc measured. I, 8-methoxymethyl-IBMX (class I PDE inhibitor); III, milrinone (class III PDE inhibitor); IV, Ro20-1724 (class IV PDE inhibitor); and V, dipyridamole (class V PDE inhibitor). Errors are standard errors of the means of the indicated number of experiments. aP = 0.04 class III PDE inhibitor vs. class I PDE inhibitor; bP = 0.003 class III PDE inhibitor vs. class IV PDE inhibitor (Fisher's PLSD). Gastroenterology 1999 116, 1379-1388DOI: (10.1016/S0016-5085(99)70502-0) Copyright © 1999 American Gastroenterological Association Terms and Conditions

Fig. 2 Effect of PDE inhibitors on Isc on sections from wild-type murine intestines. Appropriate sections of intestines were mounted in a modified Ussing chamber and exposed to either 100 μmol/L PDE inhibitor or 10 μmol/L forskolin. I, 8-methoxymethyl-IBMX (class I PDE inhibitor); III, milrinone (class III PDE inhibitor); IV (Ro20), Ro20-1724 (class IV PDE inhibitor); IV (rol), rolipram (class IV PDE inhibitor); and V, dipyridamole (class V PDE inhibitor). Errors are standard errors of the means of the indicated number of experiments. P values from Fisher's PLSD: a0.02, I vs. IV (rol); b0.0001, III vs. IV (rol); c0.04, III vs. V; d0.02, IBMX vs. III; e0.03, V vs. IV (rol); f0.01, IBMX vs. I; g0.02, IBMX vs. IV (rol); h0.04, IBMX vs. V; i0.03, I vs. IV (rol); j0.003, III vs. IV (rol); k0.04, IBMX vs. IV (Ro20); l0.0001, IBMX vs. IV (rol); m0.002, IBMX vs. V. Gastroenterology 1999 116, 1379-1388DOI: (10.1016/S0016-5085(99)70502-0) Copyright © 1999 American Gastroenterological Association Terms and Conditions

Fig. 3 Effect of PKA type-selective cAMP analogues on Isc on sections from wild-type murine intestines. Indicated sections of intestines were exposed to either type I–selective cAMP analogues (I) (8-AHA-cAMP plus 8-PIP-cAMP; 50 μmol/L each) or to type II–selective cAMP analogues (II) (N6-benzoyl-cAMP plus 8-CPT–cAMP; 50 μmol/L each). ΔIsc was then measured. Errors are standard errors of the means of the indicated number of experiments. Gastroenterology 1999 116, 1379-1388DOI: (10.1016/S0016-5085(99)70502-0) Copyright © 1999 American Gastroenterological Association Terms and Conditions

Fig. 4 Effect of PKA type-selective cAMP analogues on PKA activity. Sections of jejunum were incubated in buffer without stimulus (control), type I–selective cAMP analogues (8-AHA-cAMP plus 8-PIP-cAMP; 50 μmol/L each), or type II–selective cAMP analogues (N6-benzoyl-cAMP plus 8-CPT-cAMP; 50 μmol/L each), lysed, and fractionated into cytosolic (c) and particulate (p) fractions. PKA activity was measured and expressed as the percent of active PKA in that fraction compared with the whole cell. Errors are standard errors of the means of at least 3 experiments. *P < 0.05 vs. control; **P < 0.05 type I vs. type II. Gastroenterology 1999 116, 1379-1388DOI: (10.1016/S0016-5085(99)70502-0) Copyright © 1999 American Gastroenterological Association Terms and Conditions

Fig. 5 Effect of type-selective cAMP analogues and PDE inhibitors on Isc from murine jejunum. (A) Type II PKA-selective cAMP analogues (N6-benzoyl-cAMP plus 8-CPT-cAMP; 50 μmol/L each analogue) (II), PDE inhibitors I (8-methoxymethyl-IBMX; 100 μmol/L) and III (milrinone; 100 μmol/L) (I + III), or a mixture of all four compounds (M) were applied to either ΔF508 (F/F) or Y122X (Y/Y) homozygous or wild-type (WT) murine jejunum, and ΔIsc was measured. Errors are standard errors of the means of the indicated number of experiments. *P = 0.04 F/F vs. Y/Y. (B) DMSO (18 μL) was applied to F/F jejunum and ΔIsc measured. F/F jejunum was also mounted in chloride-free Ringer's solution, and ΔIsc was measured after the addition of cocktail. Errors are standard errors of the means of the indicated number of experiments. These mice were raised on solid food. **P = 0.047 mixture vs. DMSO; ***P = 0.0009 chloride-replete vs. chloride-free buffers. Gastroenterology 1999 116, 1379-1388DOI: (10.1016/S0016-5085(99)70502-0) Copyright © 1999 American Gastroenterological Association Terms and Conditions