1. Volume 117, Issue 3, Pages 584-594 (September 1999)
Development of interstitial cells of Cajal and pacemaking in mice lacking enteric nerves 
Sean M. Ward, Tamas Ördög, Julia R. Bayguinov, Burton Horowitz, Ann Epperson, Liya Shen, Heiner Westphal, Kenton M. Sanders 
Gastroenterology 
Volume 117, Issue 3, Pages (September 1999)
DOI: /S (99)
Copyright © 1999 American Gastroenterological Association Terms and Conditions

2. Fig. 1
GDNF−/− mice lack enteric neurons. (A) Myenteric ganglia (mg) from the gastric antrum of a D0+/+ animal labeled with PGP 9.5 immunoreactivity. (B) Complete lack of PGP 9.5 immunoreactivity in the myenteric plexus regions of the antrum of a GDNF−/− animal. (C) PGP 9.5–positive nerve fibers (arrows) running between the circular and longitudinal muscle layers of the stomach in a GDNF−/− animal. The close association of the nerve fibers with a blood vessel (Bv) suggests that these fibers are extrinsic in origin. (D) GFAP-like immunoreactivity within the myenteric ganglia (mg) and interganglionic connectives in the jejunum of a +/+ animal. Fiber tracts within the circular muscle (cm) also contained GFAP immunoreactivity (arrowheads). (E) Lack of GFAP immunoreactivity in the bowel wall of a GDNF−/− animal. (F) Occasional GFAP-like immunoreactivity was observed in fiber tracts between the circular and longitudinal muscles in GDNF−/− animals (arrows). (G) NADPH-diaphorase staining of cell bodies and processes within myenteric ganglia (arrows) and the circular muscle of the jejunum of a +/+ animal. (H) NADPH-diaphorase staining was completely absent distal to the most rostral regions of the stomach in GDNF−/− animals. (I) NADPH-diaphorase labeled some neurons and processes within the esophagus of GDNF−/− animals (arrows).
Gastroenterology  , DOI: ( /S (99) )
Copyright © 1999 American Gastroenterological Association Terms and Conditions

3. Fig. 2
Kit-like immunoreactivity in the stomach, small bowel, and colon. Kit immunoreactivity in the (A) fundus, (B) antrum, (C) jejunum, and (D) colon labeled ICC of different classes. The same populations of cells were found in the (E) fundus, (F) antrum, (G) small bowel, and (H) colon of GDNF−/− animals (see text for descriptions of Kit-positive cells). Bar in H applies to all panels.
Gastroenterology  , DOI: ( /S (99) )
Copyright © 1999 American Gastroenterological Association Terms and Conditions

4. Fig. 3
Slow wave activity in the gastric antrum of D0 mice. (A) At birth the mouse antrum produces rhythmic electrical slow waves. (A and B) Variability in slow waves in newborn wild-type mice. (C) Spontaneous electrical slow wave activity was also present in the antral muscles of GDNF−/− mice. Average electrical parameters are tabulated in Table 1.
Gastroenterology  , DOI: ( /S (99) )
Copyright © 1999 American Gastroenterological Association Terms and Conditions

5. Fig. 4
Slow wave activity in the jejunum of D0 mice. (A) At birth the mouse jejunum produces regular rhythmic electrical slow waves. (B) Spontaneous electrical slow wave activity was also present in the jejunums of GDNF−/− mice. Average electrical parameters are tabulated in Table 1.
Gastroenterology  , DOI: ( /S (99) )
Copyright © 1999 American Gastroenterological Association Terms and Conditions

6. Fig. 5
Slow wave electrical activity develops normally in cultured GDNF−/− tissues. (A) A representative electrical recording of slow wave activity from a newborn GDNF−/− animal. Slow wave activity was similar to that recorded from wild-type animals (Ward et al.8). (B and C) Antral slow waves from GDNF−/− animals developed more regular activity when maintained in an organ culture system for up to 9 days.
Gastroenterology  , DOI: ( /S (99) )
Copyright © 1999 American Gastroenterological Association Terms and Conditions

7. Fig. 6
Electrical slow waves from the jejunum of GDNF−/− animals were maintained in organ culture for up to 5 days. (A) Typical slow wave activity recorded from the jejunum at birth. Slow wave activity from jejunal tissues of GDNF−/− mice developed robust slow waves with larger amplitudes over 5 days in culture: (B) 1-day, (C) 4-day, and (D) 5-day culture. The development of slow wave activity in the jejunum of GDNF−/− mice was similar to that recorded in wild-type animals.8
Gastroenterology  , DOI: ( /S (99) )
Copyright © 1999 American Gastroenterological Association Terms and Conditions

8. Fig. 7
Quantitative RT-PCR determining transcriptional levels of SCF in small intestine for wild-type and GDNF−/− mice. Competitive PCR products were resolved on 2% ethidium bromide agarose gels. (A) A representative gel of quantitative RT-PCR for SCF in murine small intestine for wild-type and GDNF−/−. Tenfold serial dilutions of mimic DNA (shown as attomoles of mimic DNA added to PCR reaction) were included in the PCR reactions, whereas target cDNA (SCF) concentration remained constant. (B) The actual concentrations of target cDNA were calculated and expressed relative to β-actin cDNA concentration. Results are expressed as means ± SEM; no significant difference was detected in transcriptional expression of SCF between wild-type and GDNF−/− mice.
Gastroenterology  , DOI: ( /S (99) )
Copyright © 1999 American Gastroenterological Association Terms and Conditions

9. Fig. 8
Qualitative RT-PCR on dispersed and isolated smooth muscle cells from the small intestine of wild-type and GDNF−/− mice. RT-PCR amplification products electrophoresed on a 2% agarose gel. An SCF-specific 327–base pair (bp) amplification product was generated from mRNA derived from small intestinal smooth muscle cells of both wild-type and GDNF−/− mouse. The RT(−) lane was included as a negative control, indicating that the amplification products were not from DNA contamination in the mRNA preparation. Molecular weight (MW) standards are a 1000-bp ladder, and the bands shown are 200–400 bp.
Gastroenterology  , DOI: ( /S (99) )
Copyright © 1999 American Gastroenterological Association Terms and Conditions

10. Fig. 9
Small intestinal tissues that lacked enteric nerves were responsive to inhibitory and excitatory agonists. (A) Effect of exogenous application of the nitric oxide donor sodium nitroprusside (SNP, 10−6 mol/L). SNP induced a membrane hyperpolarization and a reduction in the frequency of slow waves that was reversible on washout. (B) Slow waves under control conditions, in the presence of SNP and after washout, at a faster sweep speed. (C) Acetylcholine (ACh, 10–6 mol/L) caused a depolarization in membrane potential and increased slow wave frequency. Recordings were made in the presence of nifedipine (10–6 mol/L).
Gastroenterology  , DOI: ( /S (99) )
Copyright © 1999 American Gastroenterological Association Terms and Conditions