Download Lara Slough

Histology of the Digestive System of Scyliorhinus retifer,
the Chain Catshark
By Lara Slough
Advisor: Dr. John Morrissey
Abstract
I prepared a histological description of Scyliorhinus retifer, the chain catshark, to
show that the digestive tissues resemble those of other vertebrates. After taking ten tissue
samples, I prepared them for observation. I observed my slides using light microscopy
and took photomicrographs. My results show similarities between the digestive tissues of
Scyliorhinus retifer and those of other vertebrates. These results indicate similar function
of the digestive systems as well as a common ancestor.
Introduction
During the summer of 2008, I conducted a research project about the digestive
system of Scyliorhinus retifer, the chain catshark. This small, deep-dwelling
elasmobranch inhabits the waters of the outer continental shelf and slope from Nova
Scotia to Nicaragua. Elasmobranchs, which include sharks and rays, are in a sub-class of
the class Chondrichthyes that have yet to be studied extensively. The chain catshark
reaches approximately 45cm in length as an adult. Seeking an optimal temperature of 1112°C, it is found at average depths of 70-550m. Little is known about its biology and
habits. (Compagno, 1984) I wondered if the scarcity of food in the deep sea might have
led to adaptations in the digestive system of this shark. This was certainly a possibility.
However, because all vertebrates with rare exception have similarly functioning digestive
systems, I expected it to function in basically the same way as in other vertebrates. I
studied histology to confirm or deny similarities. There have only been a couple of
publications regarding the basic range and distribution of this species, and there is some
information regarding its reproduction, but no studies about its digestive system and
histology have been published to date. (J. Morrissey, pers.com) Histology is the study of
tissues, groups of cells with similar structure that function together as a unit. If the
tissues appear similar, the function is also similar. (Gartner, 2001) My hypothesis was
that the digestive tissues of Scyliorhinus retifer would appear similar to the digestive
tissues of other vertebrates.
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I compared my results to other vertebrates to prove my hypothesis. Because
human beings have already been compared histologically to numerous vertebrates, I used
the human digestive system as a standard for comparison. (Derrickson, 2006) I also
compared my results to primitive fishes, such as sturgeons and elasmobranchs. Humans
and sharks have similar digestive systems, the basic function of which is to break down
food for absorption. The systems appear somewhat similar upon macroscopic
observation of the organs. The major organs involved include the mouth, esophagus,
stomach, intestine, and rectum. There are some differences although the function is the
same. The stomach and intestine differ slightly in humans and sharks. The stomach
serves to break down larger pieces of food into smaller pieces of food in both humans and
sharks. The intestine of the shark is adapted to the stream-lined body shape of the shark.
The spiral valve results in a compact design as well as providing maximum surface area
for nutrient absorption. (Holmgren, 1999) In the human, the intestine is long and coiled;
the small and large intestine have a combined length of nearly 9 meters. This coiled
design also provides maximum surface area for nutrient absorption, but is more
appropriately shaped for the human organism (Derrikson, 2006).
Food follows a specific path through the digestive tract. In the human, the food
enters the oral cavity, travels through the esophagus, and into the cardiac stomach. This
is the first section of the human stomach, and is heavily muscled to break larger pieces of
food into smaller pieces of food. The muscles of the stomach create a churning motion
that breaks up the food. Then the food travels through the second section of the stomach,
the fundus. Here, the smaller pieces of food are broken down into even smaller pieces of
food. The third section of the stomach is called the pyloric stomach, in which the food is
broken down into the smallest particles possible in the stomach. The pyloric stomach
opens into the intestine via the pyloric sphincter, the muscular gateway between the
pyloric stomach and the intestine. The food travels through the long coils of the
intestines, where the nutrients are absorbed, and the remaining waste is excreted through
the rectum and out the anal opening. (Derrickson, 2006)
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The path food follows in the shark is similar to the path food follows in the
human. In the shark, the food enters the oral cavity, travels through the esophagus and
into the cardiac stomach. Sharks have a two-section stomach (Figure 1a), whereas
humans have a three-section stomach. In the cardiac stomach, as in humans, larger
pieces of food are broken down into smaller pieces of food. The food enters the second
part of the stomach, the pyloric stomach, where smaller pieces of food are broken down
into even smaller pieces of food. After the pyloric stomach has sufficiently broken down
the food, it passes through the pyloric sphincter. The sphincter contracts to keep
insufficiently processed food from passing through and relaxes to allow sufficiently
processed food to pass through into the intestine. As the food travels through the
intestine, nutrients are absorbed. Because sharks have a spiral intestine, nutrients are
absorbed while traveling through an encased spiral valve (Figure 1b) rather than long
coils as in the human. The leftover waste travels through the rectum and out the anal
opening. (Holmgren, 1999)
Figure 1a. The Shark Digestive System.
(Holmgren, 1999)
Figure 1b. Inside the Spiral Intestine.
(Holmgren, 1999)
I studied histology because histology accounts for apparent differences such as
the human intestine and the shark intestine, which could be misleading. Histology, which
is the study of tissues, provides a description of an organism on a fundamental level.
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Similarities at this level are most important because they prove similarities in function.
There is a hierarchy to the structure of an organism. Individual cells make up tissues,
tissues make up organs, organs make up organ systems, and organ systems make up the
organism. The type of tissue indicates the function of the tissue, the function of the
organ, the function of the organ system, and the function of the organism. There are four
types of tissue: epithelial, connective, muscle, and nervous. Epithelial tissue protects,
secretes, absorbs, lines hollow organs, and also forms glands. Connective tissue provides
a supportive framework for the body. Muscle tissue allows the body to move. Nervous
tissue controls all functions of the body. The tissues of the human intestine and the shark
intestine are histologically similar. Therefore, they are functionally similar as well. I set
out to prove that the chain catshark was histologically similar to other vertebrates.
The focal tissues of my project, epithelial and muscular, are most prevalent within
the digestive system, although all four tissue types are present. There are several
different types of epithelial tissue, each descriptively named based on the number of cell
layers and cell shape. There are three types of muscle. Smooth muscle is involuntarily
controlled and is found in the digestive tract. Skeletal muscle is voluntarily controlled
and is found, for example, in the biceps. Cardiac muscle is involuntarily controlled and is
found in the heart. (Carneiro, 2005)
Methods
To compare the digestive system tissues of the chain catshark to those of other
vertebrates, I completed several preparatory steps. After the digestive system had been
removed from the dissection specimen, I took tissue samples from certain areas along the
digestive tract, collecting ten samples in all (Figure 2).
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1. Oral Cavity
Dorsal
3. Esophagus
8. Spiral
Intestine A
9. Spiral
Intestine B
10. Rectum
7. Pyloric
Sphincter
6. Pyloric
Stomach
2. Oral Cavity
Ventral
4. Cardiac
Stomach A
5. Cardiac
Stomach B
Figure 2. Tissue Sample Locations 1-10 (in numerical order following food path).
After collecting the tissue samples, I prepared them for observation. I dehydrated
the tissue samples by soaking them in ethanol, which is water soluble, and Safe-clear, a
tissue clearing agent. This process removes water from the sample, resulting in the space
normally taken up by water, about 80% of the tissue (Derrickson, 2006), being filled with
something more conducive to sectioning. I then embedded the tissue samples in paraffin
wax, which creates a hardened sample that is ideal for sectioning. I left the samples in
the refrigerator overnight to make sure they were thoroughly hardened.
To ensure the samples were hard enough to section, yet soft enough to avoid
splintering, I removed the samples from the refrigerator 30 minutes before sectioning.
Using a microtome set to 10 micrometers, I sectioned each sample. I placed the ribbon of
wax with embedded tissue into a water bath and let it float to flatten out. The water bath
was 45°C, to soften yet not melt the wax. After the wax had softened, I submerged a
microscope slide in the water and caught the ribbon on the microscope slide as I pulled it
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up and out of the water. I placed the slides on a slide warmer, also 45°C, to dry. I made
at least three slides from each tissue sample.
After the slides were dry, I began the staining process. Staining requires soaking
the slides in a series of chemicals that ultimately darken the tissue sample and stain it
purple, making the tissues visible. I loaded the slides into a tray and submerged them in
the appropriate sequence of chemicals for the appropriate amounts of time. I had to break
down the paraffin and re-hydrate the tissues so that the dye could sufficiently permeate
the tissues to make them visible. This required reversing the procedure I had followed
earlier in order to dehydrate the tissue samples. I soaked the slides in Safe-clear and
ethanol, the dyes hematoxylin and eosin, and associated chemicals. After the staining
process was complete, I let the slides dry, applied cover-slips, and let them dry a final
time. They were finally ready to observe under the microscope.
I observed my slides using light microscopy, noting muscle and epithelial layers.
I took numerous photomicrographs, taking care to capture a good image of each of the
tissue samples at different magnifications, finding that 4x, 10x, and 40x magnifications
gave the best results. These photomicrographs provided me with a histological
description of the chain catshark which I compared to humans as well as primitive fishes.
I compared tissues and layers present, noting similarities.
Results
The photomicrographs I took of the digestive tissues of Scyliorhinus retifer
appear similar to photomicrographs taken of the digestive tissues of other vertebrates.
White spaces inside the organs, epithelial linings, muscle layers, and white spaces outside
the organs are all present. Figure 3a-f shows my results from the cardiac stomach, Figure
4a-f shows my results from the pyloric stomach and the pyloric sphincter, and Figure 5a-f
shows my results from the spiral intestine. My results indicate that the basic histology of
the gut wall of the chain catshark is similar to that of other vertebrates.
Following the path of food, I examined my slides and took photomicrographs,
focusing on the stomach and the intestine. I took photomicrographs from two areas of the
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cardiac stomach to observe any structural changes traveling father down the digestive
tract. My photomicrographs of the cardiac stomach proximal to the esophagus (Figure
3a, 3c, 3e, 3f) show the layered structure of the stomach wall. There is white space
indicating the inside of the organ, mucosa (a lining composed of simple columnar
epithelium), sub-mucosa (loose connective tissue that supports the mucosa), dual muscle
layers (interior: circular muscle, CM, exterior: longitudinal muscle, LM), and white
space indicating the outside of the organ (Figure 3a). Dense surface cells for secretion
are visible upon closer examination (Figure 3c). There are dense dual muscle layers.
The layers formed are thick and distinct (Figure 3e, 3f). My photomicrographs of the
cardiac stomach distal to the esophagus (Figure 3b, 3d) show the slight differences in
structure in this area of the stomach. There is white space indicating the inside of the
organ, mucosa, submucosa, circular muscle, longitudinal muscle, and white space
indicating the outside of the organ (Figure 3b). Notice that while the same layers are
present as in the proximal stomach, yet they are neither as dense nor clearly defined
(Figure 3d).
Mucosa
White
Space:
Inside
Organ
CM
&
LM
CM
&
LM
White
Space:
Inside
Organ
Mucosa
Surface
Cells for
Secretion
White
Space:
Inside
Organ
Mucosa
Submucosa
White Space:
Outside
Organ
Submucosa
3a. Cardiac Stomach A.
3b. Cardiac Stomach B.
3c. Cardiac Stomach A.
4x magnification
4x magnification
10x magnification
Submucosa
Mucosa
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Dual
Muscle
Layers
CM
&
LM
Mucosa
Close-up of
Circular
Muscle
Submucosa
Close-up of
Longitudinal
Muscle
White Space:
Outside Organ
3d. Cardiac Stomach B.
10x magnification
3e. Cardiac Stomach A.
10x magnification
3f. Cardiac Stomach A.
40x magnification
Figure 3a-f. Photomicrographs of the Cardiac Stomach of Scyliorhinus retifer.
The pyloric stomach has white space indicating the inside of the organ, mucosa,
submucosa, circular muscle, longitudinal muscle, and white space indicating the outside
of the organ (Figure 4a). The mucosa is simple columnar epithelium, and the submucosa
is loose connective tissue, as in the cardiac stomach. My photomicrographs clearly show
these layers at higher magnifications (Figure 4b, 4c). The layers appear different than the
layers in the cardiac stomach (Figure 3a, 3b). There is a clear distinction between the
basal membrane of the epithelial layer and the inner layer of submucosa (Figure 4c). The
white space indicating the inside of the organ, the circular shape of the pyloric sphincter,
and the white space indicating the outside of the organ are visible in the cross-section
(Figure 4d). The smooth muscle that makes up this sphincter is shown at higher
magnifications as well, showing the density of the fibers (Figure 4e, 4f).
Submucosa
Close-up of
Basal Membrane
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Mucosa
White Space:
Inside Organ
Mucosa
CM & LM
Submucosa
Close-up of
Submucosa
CM
&
LM
White Space:
Outside Organ
4a. Pyloric Stomach.
4b. Pyloric Stomach.
4x magnification
10x magnification
4c. Pyloric Stomach.
40x magnification
White Space:
Inside Organ
Smooth
Muscle
Tissue
Muscular
Ring
Muscular
Ring
Close-up of
Muscular
Ring
White
Space:
Outside
Organ
4d. Pyloric Sphincter.
4x magnification
4e. Pyloric Sphincter.
10x magnification
4f. Pyloric Sphincter.
40x magnification
Figure 4a-f. Photomicrographs of the Pyloric Stomach and the Pyloric Sphincter of
Scyliorhinus retifer.
I compared the structure of the intestine at different places along the digestive
tract to see if the structure changed as I had done with the cardiac stomach. I took
photomicrographs of two samples from the spiral intestine, one proximal to the pyloric
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sphincter (Figure 5a, 5b) and one distal to the pyloric sphincter (Figure 5c, 5d, 5e, 5f).
There is white space indicating the inside of the organ, an epithelial layer lining the
interior of the organ with pockets for absorption, a layer of muscles lining the exterior of
the organ, and white space indicating the outside of the organ (Figure 5a, 5c). The
structure of the intestinal wall is similar to the stomach wall, mucosa, submucosa, circular
muscle, and longitudinal muscle. The spiral valve is visible in cross-section (Figure 5c).
The photomicrographs clearly show the contrast between the larger pockets for
absorption proximal to the pyloric sphincter (Figure 5a), and the smaller pockets for
absorption distal to the pyloric sphincter (Figure 5b). The epithelial lining and outer
muscle layers are not easily distinguishable in the proximal sample (Figure 5a). The
epithelial lining is more easily distinguished from the outer muscle layer in the distal
sample (Figure 5d, 5e). The distal sample also shows surface cells for absorption (Figure
5d, 5f).
White
Space:
Outside
Organ
CM
&
LM
CloseUp of
White
Space:
Inside
Organ
Closeup of
Inner
Lining
CM &
LM
Smaller
Pockets for
Absorption
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Larger
pockets for
absorption
Spiral
Valve
White
Space:
Inside
Organ
White
Space:
Inside
Organ
5a. Spiral Intestine A.
4x magnification
White
Space:
Inside
Organ
CM
&
LM
White
Space:
Outside
Organ
5b. Spiral Intestine A.
40x magnification
White
Space:
Outside
Organ
Closeup of
CM &
LM
5c. Spiral Intestine B.
4x magnification
White
Space:
Inside
Organ
Surface
Absorptive
Cell
White
Space:
Inside
Organ
Close-up of
Surface
Absorptive
Cell
5d. Spiral Intestine B.
10x magnification
5e. Spiral Intestine B.
40x magnification
5f. Spiral Intestine B.
40x magnification
Figure 5a-f. Photomicrographs of the Spiral Intestine of Scyliorhinus retifer.
Discussion
The photomicrographs I took showed muscle layers and epithelial layers matching
the general structure found in other vertebrates in all major digestive organs, though I
focused on the stomach and intestine. These two organs are responsible for the majority
of the digestive process, and so similarities in these organs are most important.
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Mechanical digestion occurs in the stomach, and absorption occurs in the intestine. The
gut wall of Scyliorhinus retifer has the same basic structure as human: epithelial layers
lining the inside and muscle layers lining the outside. The epithelial layers in the
stomach contain cells that secrete acids, enzymes, and mucus. The acids and enzymes
aid in digestion and the mucus protects the underlying tissue from damage. The mucosa
of the gut wall is simple columnar epithelium, which is a single layer of column-shaped
epithelial cells. It is supported by the submucosa, which is loose connective tissue, and
dual muscle layers beneath. The dual muscle layers, the circular muscle and the
longitudinal muscle, contract perpendicular to each other to create the churning motion
that moves food along the digestive tract and breaks up larger pieces into smaller pieces.
Gut wall layers are consistent throughout the digestive tract, differing slightly depending
on location and function. The mucosa of the stomach is specialized for secretion, while
the mucosa of the intestine is specialized for absorption. The layers also vary in density
and thickness. (Cormack, 1993) In my photomicrographs, the differences in layers are
visible along the digestive tract as the organ function changes, depending on the stage of
digestion. I observed the organs in order along the digestive tract: the cardiac stomach A
(Figure 3a) and B (Figure 3b), the pyloric stomach (Figure 4a), the pyloric sphincter
(Figure 4d), and the spiral intestine A (Figure 5a) and B (Figure 5b). The distinction
between the layers is most apparent in the cardiac stomach (Figure 3e, 3f).
The histological changes that occur along the digestive tract to compensate for the
smaller particles of food are apparent in my results. The first part of the intestine that the
food passes through, just after the pyloric sphincter, has larger areas for absorption which
is consistent with the larger pieces of food that would be found there. By the time the
food reaches the latter part of the intestine, it has been broken down into much smaller
particles, and the structure reflects this change in size. (Derrickson, 2006) In the case of
the spiral intestine, there are much smaller and more numerous pockets for absorption
farther down the intestine. (Holmgren, 1999) This progression is clearly visible in my
photomicrographs. The contrast between the proximal (Figure 5a) and the distal (Figure
5c) intestine is striking.
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Conclusion
I observed macroscopic and microscopic similarities between the digestive organs
of the chain catshark and the digestive organs of other vertebrates. To prove my
hypothesis, I compared my results to photomicrographs taken of tissue samples taken
from humans and primitive fishes. Of course, I took many more photomicrographs than I
was able to include in my figures for this paper. I considered all of these when making a
histological comparison between Scyliorhinus retifer and another vertebrate. When
comparing my results to the human, I found gut wall structure was similar. (Cormack,
1993) When comparing my results to the shovelnose sturgeon, Scaphirhynchus
platorynchus, I found similarities in the dorsal and ventral oral cavity, including taste
buds and cartilage. I also found similarities in the spiral intestine and other organs.
(Weisel, 1979) The similar structures of the esophagus and spiral intestine were
especially striking in a shark, the narrownose smooth-hound, Mustelus schmitti. (Codon,
1988) I noticed similarities to a ray as well, the thornback ray, Raja clavata, particularly
in the structure of the gut wall and the rectum. (Holmgren, 1999) Similarities in the oral
cavity, intestine, and other organs were apparent in freshwater fishes as well, the
longnose sucker, Catostomus catostomus, and the squawfish, Ptychocheilus oregonense.
(Weisel, 1962) All of these comparisons led me to conclude that these results indicate
similar tissues, similar tissue function, and therefore similar digestive organs and
digestive systems. Similarities at this basic structural level between seemingly vastly
different organisms also indicate a common ancestor and evolution. Basic structural
components retained from an ancestor and then built upon by evolution may not be
obvious, but can be observed. The shark intestine seems entirely different from the
human intestine, but histological comparison shows the same types of cells present in
both organs, showing that the shark intestine and the human intestine are the same type of
organ made up of the same types of cells, evolved into different shapes more suited to the
particular organism. The results support my hypothesis: the digestive tissues of
Scyliorhinus retifer appear similar to the digestive tissues of other vertebrates.
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References
“The Biology of Sharks and Rays”. Website.
http://www.elasmo-research.org/education/white_shark/digestion.htm
Carneiro, Jose. Junqueira, Luiz Carlos. Basic Histology Text and Atlas: 11th Edition.
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Gartner, Leslie P. Hiatt, James L. Color Textbook of Histology: Second Edition. W.B.
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Morrissey, John. Faculty Sponsor. Homepage.
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Weisel, George F.. “Histology of the Feeding and Digestive Organs of the Shovelnose
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