1. Nitric Oxide–Sensitive Guanylyl Cyclase Is Dispensable for Nitrergic Signaling and Gut Motility in Mouse Intestinal Smooth Muscle 
Dieter Groneberg, Peter König, Doris Koesling, Andreas Friebe 
Gastroenterology 
Volume 140, Issue 5, Pages (May 2011)
DOI: /j.gastro
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2. Figure 1
Lack of NO-induced relaxation in fundus from GCKO mice. (A) Western blots for fundus and jejunum strips of WT and GCKO mice probed with antibodies specific for the α1 and the β1 subunit of NO-GC (NO-GC-α1 and NO-GC-β1). (B) Traces of organ bath experiments showing DEA-NO–dependent relaxation in fundus strips from WT and GCKO mice after precontraction with 0.1 μmol/L CCh. (C) Quantitative analysis of experiments performed in panel B. (D) Traces of EFS of fundus strips from WT and GCKO mice after precontraction with 0.1 μmol/L U (E) Quantitative analysis of experiments performed in panel D. Data shown are mean ± standard error of the mean of n = 5–6 per genotype.
Gastroenterology  , DOI: ( /j.gastro )
Copyright © 2011 AGA Institute Terms and Conditions

3. Figure 2
Lack of NO-induced sphincter relaxation in GCKO mice. Representative (A) original recordings and (B) quantitative analysis of DEA-NO–induced relaxation of LES from WT and GCKO mice precontracted with CCh (1 μmol/L). Data shown are mean ± standard error of the mean of n = 4–7 per genotype. Representative (C) original recordings and (D) quantitative analysis of DEA-NO–induced relaxation of pyloric sphincter from WT and GCKO mice precontracted with CCh (1 μmol/L). Data shown are mean ± standard error of the mean of n = 5 per genotype.
Gastroenterology  , DOI: ( /j.gastro )
Copyright © 2011 AGA Institute Terms and Conditions

4. Figure 3
Effect of NO-GC deletion in duodenum and jejunum. (A) Spontaneous rhythmic contractions (representative original recordings) of duodenum from WT and GCKO mice. (B) Quantitative analysis of rhythmic contractions of duodenum and jejunum of WT and GCKO mice. Data shown are mean ± standard error of the mean of n = 3–6 per genotype. Representative (C) original recordings and (D) quantitative analysis of DEA-NO–induced relaxation of precontracted duodenum from WT and GCKO. Data shown are mean ± standard error of the mean of n = 5 per genotype. Measurements were performed in the presence of nifedipine (1 μmol/L) to prevent rhythmic contractions.
Gastroenterology  , DOI: ( /j.gastro )
Copyright © 2011 AGA Institute Terms and Conditions

5. Figure 4
Time-dependent loss of NO-induced relaxation of fundus from SM-GCKO mice. Deletion of NO-GC in SM-GCKO mice induced with tamoxifen, and fundus was analyzed in organ bath experiments 5, 10, 20, 32, and more than 50 days after the last tamoxifen injection (indicated as: d5, d10, d20, d32, and >d50; pre indicates control animals not receiving tamoxifen). (A) Deletion of NO-GC was regarded as complete 50 days after the last tamoxifen injection.27 Data shown are mean ± standard error of the mean of n = 6–7 per genotype. (B) Comparison of tamoxifen-treated and untreated WT mice showed that tamoxifen does not influence NO-dependent relaxation. Data shown are mean ± standard error of the mean of n = 7–8 per genotype. EFS of fundus from WT and SM-GCKO mice precontracted with U46619 (0.1 μmol/L). Shown are (C) representative recordings and (D) quantitative analysis. EFS-induced relaxation was sensitive to the specific NO-GC blocker ODQ (10 μmol/L). Data shown are mean ± standard error of the mean of n = 6 per group.
Gastroenterology  , DOI: ( /j.gastro )
Copyright © 2011 AGA Institute Terms and Conditions

6. Figure 5
Relaxation of LES from SM-GCKO mice and downstream PKG signaling in LES and fundus. LES from (A) SM-GCKO mice and (B) WT were precontracted with CCh (1 μmol/L) and subsequently relaxed with DEA-NO. Pre, control animals that did not receive tamoxifen; >d50, tamoxifen-injected animals analyzed 50 days or more after tamoxifen treatment. (A) ODQ (10 μmol/L) was administered when indicated to block NO-GC–dependent signaling. (C) Fundus and (D) LES from WT and SM-GCKO mice were treated with the direct PKG activator 8-Br-cGMP to test downstream signaling. Data shown are mean ± standard error of the mean of n = 5–6 per genotype.
Gastroenterology  , DOI: ( /j.gastro )
Copyright © 2011 AGA Institute Terms and Conditions

7. Figure 6
Immunohistochemical analysis of NO-GC expression. Expression of the β1 subunit of NO-GC (NO-GC-β1) was visualized in the fundus of (A) WT, (B) GCKO, and (C) SM-GCKO animals (bars, 50 μm). (A) Circles and asterisks indicate strong immunoreactivity in the LMM and in vascular smooth muscle cells of the mucosal layer (M), respectively. Arrowheads show the weak staining in the circular and longitudinal muscle layers, and arrows indicate the very strong NO-GC expression in cells dispersed throughout the muscularis externa (ME). To show the absence of NO-GC in smooth muscle cells, a section of the fundus from SM-GCKO mice was double-stained with an antibody specific for (D) NO-GC-β1 and (E) α-smooth muscle actin (α-SMA). (D and E) Arrows and arrowheads indicate the loss of NO-GC expression in smooth muscle of the lamina muscularis mucosae and vascular smooth muscle, respectively. (F) Merge of panels D and E; yellow signals do not indicate colocalization of NO-GC and α-smooth muscle actin but result from overexposure of the α-smooth muscle actin signal of the muscularis externa that was necessary to show adequate staining of vascular smooth muscle.
Gastroenterology  , DOI: ( /j.gastro )
Copyright © 2011 AGA Institute Terms and Conditions

8. Figure 7
Whole-gut transit time of WT, heterozygous, and SM-GCKO mice. Each symbol represents the transit time of one animal tested. The bars represent mean values. n = 8–16 per genotype.
Gastroenterology  , DOI: ( /j.gastro )
Copyright © 2011 AGA Institute Terms and Conditions