Objectives:

The first week, concentrate on the esophagus and stomach.
The second week, on the small and large intestine.

After this laboratory, students should be able to identify different organs of the alimentary canal from histological sections and explain the meaning of the terms listed at the end of the lab exercise. You will be expected to not only identify the major organs, i.e. esophagus, stomach, small intestine, colon, rectum and anus, but to identify the particular region of the organs, i.e. duodenum, jejunum and ileum of the small intestine; cardiac, fundic and pyloric regions of the stomach; upper, middle and lower regions of the esophagus etc. These different structural variations are summarized at the beginning of the slide descriptions of the particular organs. The histology of the alimentary canal is summarized at the end of the lab manual.
 
 

 
1. 4 layers of GI tract;

4. Muscularis: inner circular layer, outer longitudinal layer.
Use this to determine orientation of section:

77a Esophagus, upper part 

Compare this slide with the previous one. It is pale stained but reveals the difference in content of the muscularis, skeletal muscle is present here. Compare the skeletal muscle fibers with the smooth muscle fibers of the muscularis mucosa. The muscularis mucosa is difficult to fine but it lies immediately underneath the epithelium.
 
 
 

 77b Esophagus, upper, human
This slide is unusual because it has some submucosal mucous glands and their ducts, which are lined with a stratified squamous epithelium. These glands are located throughout the esophagus, but it is rare to get a slide that shows them. The muscularis consists of entirely skeletal muscle, which can be compared in appearance with the smooth muscle of the muscularis mucosae.
 
 

 
79a Esophagus & stomach
There are two groups of slides here. The older ones show both cardiac stomach and fundic stomach so that you can compare the glands. The newer slides have a piece of skeletal muscle attached. Notice how the muscularis mucosa thins out as it enters the stomach. Near the junction, the glands are typical cardiac glands with mucous secreting cells at the baase and duct cells along the neck. Further along the epithelium, it changes to fundic glands and you can see several different cell types.
 
 

79b Esophagus & stomach, human
The preservation here is not too good, but you can get a feeling for the dramatic change that occurs at the gastroesophageal junction. The cardiac stomach epithelium is a bit confusing for several reasons. First, there are eosinophilic cells throughout the glands but especially at the base where mucous cells are supposed to be. These cells look alot like pareital cells from the adjacent fundic glands. In addition some of the glands look a lot like fundic glands. The mucous surface cells, where preserved are typical. The cardiac type of glands are found very near the junction.
 
 

79c Esophagus-stomach junction, human
This slide illustrates the tripartite structure of the cardiac glands of the stomach. In a few places the glands are cut in longitudinal orientation. Note that the pits extend only about 1/3 of mucosa. About halfway through the remaining mucosa the glands change in appearance. This 1/3 - 1/3 - 1/3 division is characteristic of cardiac glands. Note also that in each region of the glands, there seems to be a homogeneous cell type. In other places of the section, the gastric mucosa is cut in oblique section. Again there seem to be 3 regions of the glands. The esophagus has an interesting feature, namely the presence of an esophageal cardiac gland. Note the abrupt change from stratified squamous to simple cuboidal. At the other end of the GI tract at the rectal-anal junction, this transition from simple columnar to stratified squamous is accomplished through a short bridging stretch of pseudostratified columnar epithelium.
 
 

 
A. Cardiac stomach

80a Stomach, cardiac region, human
80b Cardiac stomach
This slide of cardiac stomach has the appearance of three regions of cells in the mucosa, i.e. mucous surface, duct and mucous cells in the glands. Note that the mucous cells at the base of the gland have a flattened basal nucleus while the mucous surface cells have a more columnar shape. This division into three regions is characteristic of cardiac stomach. The pyloric stomach, on the other hand has the appearance of 2 regions, the pits and the glands; the middle region is much less apparent. In some regions the serosa is intact.
 
 

B. Fundic stomach

1. 4 layers of GI tract; mucosa, submucosa, muscularis externa, serosa
2. Mucosa contains 3 layers, simple columnar epithelium, lamina propria and muscularis mucosa of smooth muscle
3. Mucosa arranged as wide bore pits leading to long, narrow bore relativley straight tubular glands. Glands are long relative to the pit
4. Epithelium: mucous surface cells line the surface and the pits. Glands contain three major cell types (diagnostic for this region of stomach):

• parietal cells (fried-egg shaped, producing HCl),
• chief cells (granular cytoplasm, producing pepsinogen) and
• mucous neck cells (pale staining granules, producing mucus).

82c Stomach, Fundus, H&E
The cellular preservation in this slide is quite poor. But even so, you should be able to identify pareital and chief cells in the mucosal glands. The one redeeming feature of this tissue is the presence of a serosa, which remains largely intact with the mesothelial cells somewhat enlarged and therefore more visible.
 
 

82a Fundic stomach

This is the best stomach slide. There are 4 types of cells in the glands. There is only one layer in the muscularis and the serosa is missing. One very interesting feature of this slide is the fact that the muscularis mucosa shows three layers. In the small intestine the mm shows 2 layers, just line the muscularis. But in the stomach where the muscularis has three layers, the muscularis mucosa also has three layers, at least in the example, which comes from an unknown species.
 
 

 82d Stomach, Fundus, MA
This slide reveals the connective tissue distribution pretty well but it will be difficult to detect some of the mucosal cell types. The section is thick so there is a lot of superposition. Mucous neck cells and chief cels are pretty indistinguishable. Pareital cells are the large blue cells and if you look carefully you can see that they have the characteristic fried egg appearance. Te serosa is visible as the red outline but the mesothlial cells are not very squamous, perhaps a post mortem change or an en face view.
 
 

 82b Stomach, Fundic, Human
This is a good slide of fundic stomach. The preservation is relatively good except that some of the mucosal cells are a bit shrunken. You should be able to visualize 4 cell types in the fundic glands. There is also a bit of the mucous lining of the surface intact. Auerbach’s plexus is a bit difficult to see. Meissner’s plexus is quite difficult to find. Because of the positioning of Auerbach’s plexus between muscularis layers, you don’t need to see neurons. To clearly identify Meissner’s plexus from a peripheral nerve, you need to observe a ganglion cell. If you think you have found one, share the experience.
 

82e Stomach, fundic region, human

There are only 2 copies of this slide but it is a good one to look at because it shows the pits in both cross and longitudinal section. It is possible to distinguish the 4 cell types of the pits and glands although the mucous neck cells will have to be identified by process of elimination. The parietal cells are fried-egg shaped, the chief cells have obvious granules and the ones that are left are the mucous neck cells, with pale staining cytoplasm.

C. Pyloric stomach

84b Stomach, pyloric region, human
The preservation here is not wonderful, but it is good enough for you to get the essence of pyloric stomach structure. Note that throughout the glands, there is only one obvious cell type. The pits are very deep which is a characteristic of pyloric stomach. The glands also have a relativley wide lumen and the mucous secreting cells (nearly all of the cells are mucous) are relativley large, much like the mucous surface cells.
 

85 Pyloric stomach, Monkey, H&E, plastic

Contraction of the muscularis has caused this tissue to curl inside out. The mucosa is on the outside and the serosa is on the inside. Mostly this is pyloric stomach but there is a small piece of fundic stomach attached which gives you the opportunity to appreciate the gradual conversion from fundic glands to pyloric glands. Unfortunately the cellular preservation in the glands is not too good. Look for areas of pyloric glands that are cut in longitudinal section and observe how deep the pits penetrate into the mucosa. In the fundic glands, the pits are much shallower. There are numerous examples of Auerbach’s plexi. The mesothelium is not easily visualized.
 

D. Self Test

55a Stomach, human H&E
This slide is unlabeled with regard to the stomach location. Can you determine what region of the stomach it came from? There are several different slides in this group so each one gives a different view. Concentrate on the glands in the mucosa. The muscularis is in poor condition and some of the surface epithelium is missing. However there is enough material left that you can identify the region.
 
 

88c Stomach-duodenum junction, l.s.
This slide brings out an important difference between stomach and small intestine. In the stomach the surface epithelium consists of a single kind of cell the mucous surface cell. In the small intestine, there are two kinds of cell, absorptive enterocytes and goblet cells. The presence of villi are difficult to establish because of the apparent presence of food in the lumen. In the stomach, the mucosa consists of pyloric glands. These can be identified because the pits are deep and occupy about 1/2 of the mucosa, the other half is occupied by the gland. This slide also has a piece of pancreas attached.
 
 
 

 
A. Duodenum

87b Duodenum, human
This is an H&E stained slide. The general preservation is poor. The surface epithelium has exfoliated, but the Bruener’s glands stand out very well as does the muscularis. Auerbach’s plexus is easily visible as the pale staining regions between the inner and outer muscle layers. A palely stained serosa can be seen with difficulty.
 

B. Jejeunum

90c Jejunum, Masson, human 

90d Jejunum, c.s.
This is a good cross section of jejunum, but it is not human. Because it is a cross section, the plicae circulares are not prominent, but traces can be seen by the periodic thickening of the submucosa. Meissner’s plexi are numerous and can be identified by the large, pale staining nuclei with prominent nucleoli. Paneth cells are not prominent. Mitotic figures are very numerous in the crypts.
 

90a Jejunum, human, c.s.
There are 2 groups of these slides. This section has nice tall plicae, tall villi, short Brunner’s glands. There are many cross and longitudinal sections through villi. The serosa is intact, but the hematoxylin staining is so faint that the mesothelium nuclei are difficult to see.
 
 

90e Small intestine human, H&E
This slide has tall villi but the apical epithelium has been lost in most places. It is difficult to see a brush border on the enterocytes. However, you can see goblet cells clearly so identification of the enterocytes is easy by process of elimination.
 

Jejunum, rabbit 1.5µm, BF-TB
This is a 1.5µm thick plastic section stained with basic fuscin and toluidine blue. The orientation of the section is approximately transverse but many of the villi are sectioned obliquely. As a result of the oblique cut of the section, the simple characteristic of the epithelium is obscurred and it looks stratified. However, you will find regions where the true type of the epithelium can be discerned. Goblet cells stain a reddish brown color with this stain. You can see how the goblet cells disrupt the brush border in places. The brush border is very distinct. The section is so thin that you will think that you can resolve the microvilli. Instead you are actually seeing clumps of the microvilli. Try finding a classical goblet cell and enterocyte and then sketch them together below. At the base of the crypts of Leiberkühn, you will find Paneth cells. You can identify them by the pale staining granules in their cytoplasm. In the villi, you will find both capillaries and lacteals. This animal had just eaten prior to being sacrificed so the lacteals are swollen. Lacteals can be differentiated from capillaries by the presence of what structure or feature? In the lamina propria you can find plasma cells occasionally. There are areas through the columnar ejpithelium where the cells can be seen outlined by a dark line (in most areas they are outined by a white line). What is this structure giving rist to the dark line. In the submucosa are Meissner’s plexi. In this slide the submucosa contains, in addition to collagen fibers, arteries, veins and Meissner’s plexus. Find the Meissner’s plexus by eliminating the other possibilities. Auerbach’s plexus can be found in the muscularis. In the outer muscle layer that is sectioned transversely, you can see the serrated edges of the smooth muscle cells. The serosa is present in most places.
 
 

C. Ileum

1. 4 layers of GI tract; mucosa, submucosa, muscularis externa, serosa

2. Mucosa contains 3 layers, simple columnar epithelium, lamina propria and muscularis mucosa of smooth muscle

3. Mucosa extensively folded into villi with narrow bore crypts at base, the crypts of Lieberkühn. Villi may reveal pronounced lymphatics, the lacteals

4. 3 major epithelial cell types: enterocytes (brush border), goblet cells, Paneth cells at base of crypts (intensely basophilic cytoplasm)

Presence of mucosal folds, the plicae circularis with villi extending off of them. Submucosa of dense irregular connective tissue with no glands. Villi are short compared with jejunum.

Submucosa of dense irregular connective tissue with no glands

7. Presence of Peyer's patches (five or six aggregeted lymph folliculi together) in submucosa. Lymphocites can invade the muscularis mucosae and the propria as well. Diagnostic if present. However, Peyer's patches are located only in the distal ~6 inches of the organ at the antimesenterial portion . Most of the ileum lacks them.

8. Muscularis externa contains 2 clearly defined layers of smooth muscle. Inner is circular, outer is longitudinal (use to determine section orientation).
 

92b Ileum human, sec.
The orientation of this section is roughly longitudinal so you can see plicae readily. In addition the villi are cut in cross and longitudinal section thereby showing clearly that the mucosal foldings are villi and not pits. There are only a few lymph nodules, not the dense arrangement that constitutes Peyer’s patches.
 

92c Ileum, H&E
The epithelium in this slide is nearly completely destroyed. However, a row of lymph nodules can be seen which would be suggestive of Peyer’s patches. That is about all that there is to look at in this slide.
 
 

92e Ileum, human, H&E
This slide, which is a longitudinal section shows truly dramatic examples of plicae circulares. The epithelium is nicely preserved even though it has detached from the underlying lamina propria. You should be able to see clearly the brush border and distinguish the goblet cells from the enterocytes. At the base of the crypts, notice the cells with red granules. These are the Paneth cells. What do these granules contain? The Meissner’s and Auerbach’s plexi are quite numerous.
 

92 Ileum Peyer’s patches, monkey
This side was looked at back in the defense lab. The lymph nodules are distributed rather differently than Peyer’s patches, but the number of patches suggests that this is indeed ileum.
 
 

Small intestine composite

There are two groups of this slide. Use this slide as a test to see if you can identify the three regions, which are not separately labeled.

For 94a the duodenum has relatively sparse submucosal glands, but they are in sufficient numbers to enable you to identify the duodenum. For the ileum, the Peyer’s patches are located on only one side, which is classically the distribution. This is the only slide that really shows this feature correctly. In the ileum, notice that there are relatively more goblet cells than are present in the other regions. The preservation of the surface epithelium is relatively poor.

For 94b, the epithelium preservation is much better. For the duodenum example, there is a strange structuring of the submucosa in the duodenum example, that may represent Brunner’s glands, but you would never know it. The jejunum has a piece of glandular tissue attached to it, which is the pancreas (topic for next lab). The ileum example in the new group has only a single lymph nodule, which would hardly classify the tissue as ileum. The presence of Peyer's patches is diagnostic for ileum, but the absence of Peyer's patches is not diagnostic for jejunum. Without Peyer’s patches, the identification of ileum from jejunum is very difficult and you must rely on such things as height of plicae and villi as well as the rather higher proportion of goblet cells in the ileum
 

  - Before beginning this section it is worth noting that slides of colon rarely preserve the surface epithelium although they usually do a good job with the crypts. Do not be surprised if the surface epithelium seems to be missing, because most of the time it is. Slide 98a has much of the surface epithelium in tact and so is worth examining, if only for that reason.

1. Ileum-colon junction

96b Colon, Mucicarmine, human

96a Colon, human, H&E
This is a fairly typical H&E preparation. Note that the glands have many goblet cells and that they have constant diameter. There is some suggestion that the outer part is bound by a serosa, but the mesothelium is not obvious. Some slides show a gathering of the outer muscle layer into tenae coli.
 
 
 

97b Colon, human, sec
Another fairly typical colon prep. This one does not reveal any hint of tenia coli in the outer muscularis externa.
 
 

97a Colon, c.s.
There are 2 groups of slides here but both are equally good. A clear mesothelium in visible in the older group, but is also present in the newer group though more difficult to see. Neither slide shows tenia coli in the external muscle layer. Note that the glands contain cells that are not goblet cells. In some regions these cells will reveal a brush border.
 

1. Rectum

a. Stratified squamous epithelium that eventually becomes keratinized (cutaneous zone)

b. Layering becomes that of epidermis

c. Hair follicles with sebaceous glands

d. Eccrine sweat glands

e. May contain submucosal mucous glands - anal glands

f. May contain external anal sphincter identifiable because it has skeletal muscle

g.May contain apocrine scent glands (of Montgomery) , the circumanal glands

98b Rectum, human, H&E
The typical structure of the rectum is a mucosa of straight, "test-tube" shaped glands with many goblet cells. The surface of the mucosa is very well preserved. Though this slide is stained with H&E, note that under the surface epithelium is a pale stained layer that probably is the collagen table (RR&K, p. 469). There are some very well preserved Auerbach’s plexi in the muscularis. Note that the outer muscle layer is relatively uniform in thickness.
 

99a Rectum & anus, Masson

The stain makes this one look kind of peculiar, but it has all the typical structures of this part of the GI tract. Of particular note is the transition from columnar to stratified squamous. Hair follicles in the stratified squamous epithelium identify this tissue as having a piece of anus attached. The stratifed squamous epithelium is keratinized over only part of its surface. Another notable feature is the large amount of skeletal muscle, the external anal sphincter. The glands in the rectum portion are similar to those in the rest of the colon except that nearly all the cells are mucous secreting. Note that as the anal canal is approached that the muscularis mucosae gradually disappears.
 
 
 
 

99c Rectum-anus junction, human
This slide shows the typical test-tube shaped glands of the rectum with predominant goblet cells. Also present, and quite a large stretch is a region of stratified columnar epithelium between the rectum and the stratified squamous epithelium of the anus. Note the presence of hair follicles of the anus as well as apocrine scent glands. The large external anal sphincter is easily identified.
 
 

100 Anal canal, monkey
This is a nice longitudinal section that gives a good idea of the transition from columnar to stratified squamous. There is a large piece of skeletal muscle present, the external anal sphincter. Everything else about this slide is pretty typical.
 
 

99b Rectum & anus, Masson

You will have trouble navigating this slide. The structure of the rectal portion is strange looking because there are few longitudinal cuts through the glands. Superficially, the glands appear to have a wide bore leading to narrow bore glands but the presence of skin features in the anal portion will eliminate the possibility of stomach. In the anal portion are some mucous secreting glands that are clearly different from those in the rectum. These are the anal glands. The adventitia is unremarkable. The simple columnar epithelium is quite irregular in folding because there are some rather large circular profiles indicating some large luminal openings.
 
 

 

General  tunica mucosa  epithelium mucosae  lamina propria mucosae  muscularis mucosae tela submucosa  Meissner’s plexus tunica muscularis   peristalsis  Auerbach’s plexus=myenteric plexus tunica serosa  mesothelium  mesentery tunica adventitia  GALT  Esophagus  mucosa glands  esophageal cardiac glands  pyrosis  gastroesophageal junction Stomach  chyme  Rugae  gastric pits regions  cardia/cardiac  fundus/fundic  pylorus/pyloric glands  fundic glands  cardiac glands  pyloric glands epithelial cells  mucous surface  mucous neck  chief  parietal/oxyntic  enteroendocrine  APUD  GEP secretions  pepsinogen/pepsin  hydrocholric acid (HCl)  intrinsic factor  gastrin hormones  gastrin  cholecystokinin  secretin  gastric inhibitory peptide (GIP)

Small Intestine  plicae circulares (of Kerkring)  villi  crypts of Lieberkühn  lacteal vessels  microvilli/striated border  glycocalyx regions  duodenum  jejunum  ileum epithelial cells  Paneth cells  enterocytes  goblet cells  enteroendocrine cells  M cells (microfold cells) Brunner’s glands  Peyer’s patches  Large Intestine  collagen table  pericryptal fibrobast sheath  teniae coli Rectum  anorectal junction Anus  Columnar zone  Hemorrhoid zone  Cutaneous zone  anal columns  anal sinuses  anal glands  circumanal glands  internal anal sphincter  external anal sphincter