INTRODUCTION TO DYNAMIC HISTOLOGY – ANAT 261
LABORATORY MANUAL
WRITTEN BY: CARLOS R. MORALES
INDEX
Electronic Slide Box …………………………………………………………….. Page 2
Lab 1 (Organization. Training. Skin and Epithelial tissue) ……………………… Page 3
Lab 2 (Skin and Annexes).……………………………………………………….
Page 4
Lab 3 (Cartilage and Bone).……………………………………………………… Page 5
Lab 4 (Bone Formation) ………………………………………………………..... Page 6
Lab 5 (Muscle Tissue) …………………………………………………………… Page 7
Lab 6 (Blood Vessels) …………………………………………………………… Page 8
Lab 7 (Respiratory System).……………………………………………………… Page 9
Lab 8 (Oral Cavity).………………………………………………………………. Page 10
Lab 9 (Digestive Tract).…………………………………………………………… Page 11
Lab 10 (Associated Glands).…………………………………………….………… Page 12
Lab 11 (Urinary System) …………………………………………………….…… Page 13
Lab 12 (Nervous System) ………………………………………………………… Page 15
1
Electronic Slide Box
1.
2.
3.
4.
5.
6.
7.
8.
Skin - Finger
Striated Skeletal Muscle (Diaphragm)
Cardiac Muscle
Lip - Skin with hair follicles
Spinal Cord
Brain
Cerebellum
Tongue
9.
10.
11.
12
13.
14.
15.
16.
17.
Esophagus, Trachea, Elastic Arteries, Large Veins
Urinary Bladder
Lung
Submandibular Gland
Pancreas
Liver
Pyloric Stomach, Duodenum
Kidney
Mature and Developing Teeth
18.
19.
20.
21.
22.
23.
24.
25.
26.
Sublingual Gland
Endochondral Bone Formation - Tendon
Endochondral Bone Formation
Compact Bone
Spinal Ganglion and Peripheral Nerve
Aorta
Rectum, Jejunum, Body of Stomach, Colon
Larynx (Elastic Cartilage)
Parotid Gland
27.
28.
29.
30.
Lung
Ileum
Ileum
Colon
2
ANATOMY AND CELL BIOLOGY
Introduction to Dynamic Histology
Laboratory 1
Objectives
1.
Organization: The teaching in the labs is organized as small group teaching. Six
or seven students are assigned to one station with a TA in charge of the group.
You will sit in the same station for the rest of the semester.
2.
Learn the use of the Electronic Slide Box: Most of the activities described in this
manual specify the use of electronic slides. The electronic slides have been listed
separately (see Electronic Slide Box Document)
Learn the use of digitized micrographs: These micrographs are found in Images.
Generally, they will be used by the TA to demonstrate the lab activities, to answer
specific questions, and to review the lab. This folder also contains useful drawings
and electron microscope images. At the end you will find a section named
“cytology”. Feel free to use these electronic images in your assignment.
A Laboratory Review describing the main concepts that you should learn in the
3.
4.
lab is also available. Be advised to read the text prior to the lab.
Assignments
1.
2.
3.
Become familiar with the laboratory systems: The use of the electronic slide box
will simulate light microscopy. At low power you will observe the image at a
0.4x. You can magnify the same field up to 40x which is equivalent to the highdry ocular of a light microscope. The small image at the corner has a red rectangle
that indicates the selected field. You can place it over the area you want to
observe. Additionally, the arrows at the bottom will allow you to move the picture
in different directions.
When using electronic slides, follow these steps of observation: A) Start with low
power (L.P) and end with high power (H.P.); B) What is a longitudinal, a cross
and an oblique section? C) Always identify your final magnification
Your TA may decide to show you first the Lab Images. In such a case use then the
electronic slide 1 for review. Identify the epidermis, dermis and hypodermis.
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ANATOMY AND CELL BIOLOGY
Introduction to Dynamic Histology
Laboratory 2
Objectives
1.
2.
3.
4.
To study the histology of the skin and its appendages (hair, sebaceous and sweat
glands).
To study the keratinized stratified squamous epithelium (epidermis).
To identify the phases of mitosis in the stratum germinativum (basal layer), i.e.,
interphase, prophase, metaphase, anaphase and telophase.
To identify the dermis and hypodermis.
Assignments
1.
2.
3.
4.
5.
6.
Use electronic slide 1. Make a drawing of the epidermis at L.P and H.P.
Use electronic slide 4. In the dermis, study the hair follicle and identify its layers
(C.T. sheath, glassy membrane, E.R.S., I.R.S., cuticle of hair shaft and cortex).
Use your Atlas of Histology and your drawings from the lecture. Note that the
I.R.S. is composed by the Henle’s layer, Huxley’s layer, and the cuticle of I.R.S.
Study the sebaceous gland at L.P. and H.P. Make a drawing.
Use slide 1. Study the sweat gland and its duct at L.P. and H.P.
Identify mitotic figures in the basal layer.
If you have time compare the epidermis to the dermis (C.T.) and hypodermis
(C.T.).
C.T.: connective tissue
4
ANATOMY AND CELL BIOLOGY
Introduction to Dynamic Histology
Laboratory 3
Objectives
1.
2.
3.
To study the classification and histology of the connective tissue (using the
dermis and hypodermis as study models).
Histology of cartilage
Introduction to the histology of the bone.
Assignments
1.
2.
3.
4.
5.
6.
Use electronic slide 1. Study the dermis and hypodermis of the skin. The dermis
is divided into a papillary layer (loose connective tissue) and a reticular layer
(dense irregular connective tissue). The hypodermis is composed of adipose tissue
(a variety of loose connective tissue). Locate sweat glands and their ducts in
these layers of the skin.
Use electronic slide 9. Locate the trachea. Examine the hyaline cartilage. Study
the perichondrium, the isogenous groups and the extracellular matrix. Identify
chondroblasts and chondrocytes. In slide 25 find elastic cartilage. Identify elastic
fibers.
Use electronic slide 19 and locate dense regular C.T. (tendon). Note that the
collagen fibers run in one direction.
Slide 19. Note that the articular surface is made of hyaline cartilage. Do you see
perichondrium in this location?
Use electronic slide 21. Bone. Cross section of shaft (long, compact lamellar
bone). Identify the following structures: periosteum, outer circunferential system
or lamellae, Haversian system, inner circumferential system or lamellae,
endosteum, marrow, osteoblasts, osteocytes, osteoclasts, Haversian canals
(vascular canals).
Make drawings.
5
ANATOMY AND CELL BIOLOGY
Introduction to Dynamic Histology
Laboratory 4
Objectives
1.
2.
To study the histology of the bone.
To study endochondral bone formation.
Assignments
1.
2.
3.
4.
5.
6.
6.
Use slide 21. Review bone structure. Note the following points: Bone tissue is
made up of cells and matrix (matrix consists of collagen and ground substance).
The collagen is impregnated with minerals in the form of hydroxyapatite crystals.
Cells are: a- Osteoblasts (seen during bone formation) line the bone surfaces and
are large when active and flat when inactive. b- Osteocytes (lie in bone). They
are seen in small holes called lacunae. Osteocytes have processes which run in
small canaliculi in the bone matrix. c- Osteoclasts (bone resorption). These are
large multinucleated cells which remove bone (matrix, minerals and cells).
In the rat there are no true Haversian systems. But the canals containing blood
vessels may be considered to be Haversian canals. Review the following:
periosteum, endosteum, inner and outer circumferential systems and interstitial
system.
Study the articular surface (Slide 19, hyaline cartilage without perichondrium).
Slides 19 and 20. The bone that is first formed is embryonic or woven bone. This
bone is laid down around the cartilage model of the bone. The cartilage model is
removed during endochondral bone formation leaving two discs of cartilage at
either end of the bone; these are the epiphyseal plates. They are responsible for
the growth in length of the bones.
Slides 19 and 20. Study the epiphyseal plate. Identify the different layers (resting
cartilage, zone of proliferation, hypertrophy, cell death and mixed spicules.
Make drawings
6
ANATOMY AND CELL BIOLOGY
Introduction to Dynamic Histology
Laboratory 5
Objectives
1.
2.
3.
To study the histology of striated (skeletal) muscle.
To study the histology of cardiac muscle.
To study smooth muscle (wall of intestine).
Assignments
1.
2.
3.
4.
Use slides 2 and 8. Make drawings of longitudinal, cross and oblique sections of
muscle fibers (striated skeletal muscle). In cross sections identify peripheral
nuclei and myofibrils. In longitudinal sections note long multinucleated fibers,
peripheral nuclei and cross striations of the fiber.
Compare striated skeletal muscle to cardiac muscle (slide 3). Cardiac muscle is
similar to striated skeletal muscle, but has central nuclei and the fibers (cells) are
joined together at intercalated discs.
Compare striated skeletal muscle to smooth muscle (slides 15 and 24). Note
fibers in cross sections presenting centrally located nuclei. In longitudinal section
the fibers are elongated, spindle shaped with central nuclei.
Make drawings at L.P. and H.P.
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ANATOMY AND CELL BIOLOGY
Introduction to Dynamic Histology
Laboratory 6
Objectives
1.
2.
To study the histology of blood vessels (elastic arteries, muscular arteries,
arterioles, large veins, medium size veins and venules).
To identify the different layers of these vessels.
Assignments
1.
2.
3.
4.
Find a muscular artery (media with four or more layers of smooth muscle cells)
and a medium size vein in slides 1, 4, 8 and 24. In each case, try to identify cross
and longitudinal sections of blood vessels. Examine the intima, media and
adventitia. In the case of the arteries find the internal elastic limiting membrane
(IELM).
Identify other vessels such as arterioles and venules in the same sections. How do
they differ from muscular arteries and medium size veins?
Study elastic and large size veins. Use slides 9 and 27. Sketch a portion of the
media of the elastic artery showing fenestrated elastic membranes and the smooth
muscle fibers between them. What is and where is located the vasa vasorum?
Make drawings of all these vessels at L.P. and H.P.
8
ANATOMY AND CELL BIOLOGY
Introduction to Dynamic Histology
Laboratory 7
Objectives
1.
To study the histological organization of the trachea
2.
To study the histology of the respiratory system
Assignments
1.
2.
3.
4.
Use slide 9. See that the lumen of the trachea is lined by a pseudostratified
columnar epithelium with cilia and goblet cells. Make a H.P. drawing. The thin
layer of connective tissue (loose) under the thickened basement membrane is the
lamina propria. This C.T. together with the epithelium is collectively called
mucosa. The connective tissue below the mucosa is the submucosa and contains
serous and mucous acini. Finally the submucosa merges with an open ring of
hyaline cartilage completely surrounded by perichondrium. Look from
chondrocytes (isogenic groups).
Find intrapulmonary bronchus (Slide 28). Study the folded mucosa, the
incomplete plates of cartilage and the presence of smooth muscle (Find slides in
the wooden box).
Slide 28. Find bronchioles (regular, terminal and respiratory), alveolar ducts,
alveolar sacs and alveoli. Identify macrophages and pneumocytes type II.
Make detailed drawings of these structures.
9
ANATOMY AND CELL BIOLOGY
Introduction to Dynamic Histology
Laboratory 8
Objectives
1.
To study the histology of the digestive system
a)
Teeth
b)
Tongue
c)
Esophagus
Assignments
1.
2.
3.
4.
5.
6.
7.
Teeth. Slide 17 (Cross section of monkey maxilla). Find erupted primary tooth
and unerupted permanent teeth. Examine the gingiva and the periodontal
ligament around the erupted tooth.
Study the gingiva. Find, if possible, the muco-gingival junction. Identify the
parakeratinized squamous epithelium.
Study the erupted tooth. Find the dentine and odontoblasts, the pulp, the
cementum and cementocytes. Find the Sharpey's fibers in the cementum. The
enamel (crown) is absent (removed during decalcification). What is different
about the cementum near the crown and at the tip of the root?
Developing crowns of permanent tooth. Find the enamel (white space and/or
bluish material), the ameloblasts (inner dental epithelium), stratum intermedium,
stellate reticulum, outer dental epithelium and Hertwig's epithelial root sheath.
What is the enamel organ?
Make drawings of all these structures. Also take a look at the inner aspect of the
lip (Slide 4).
Tongue (Slide 8). Find filiform, fungiform and circumvallate papillas. In the last
one, find the taste buds. Make drawings.
Esophagus (Slide 9). The mucosa consists of a non-keratinized stratified
squamous epithelium and a thin lamina propia (loose C.T.). There is a muscularis
mucosa. The submucosa is thick (dense C.T.) and contains mucous glands. The
tunica muscularis has two layers. The outermost layer is called adventitia. Make
drawings.
10
ANATOMY AND CELL BIOLOGY
Introduction to Dynamic Histology
Laboratory 9
Objectives
1.
Study of the histology and general organization of the gastro-intestinal tract
(esophagus, stomach and intestines).
Assignments
1.
Review esophagus.
2.
Reed on your atlas and lectures about the cardiac portion of the stomach (no
slides).
3.
4.
5.
6.
Body of the stomach. Slide 24. Find the mucosa and identify pits and glands.
The lamina propria is squeezed between the gastric glands. The gastric glands
extend down to the muscularis mucosa. Note the thin submucosa and the tunica
muscularis composed of three layers of smooth muscle. Sketch these layers under
L.P (10X). Identify the following cells: surface mucous cells, mucous neck cells,
parietal cells, zymogenic cells.
Pyloric portion of the stomach and duodenum. Slide 15. Follow the mucosa and
identify both regions. One follows the other. Note that in the pylorum, the glands
are lined by mucous cells. In the duodenum the mucosa forms villi and crypts.
The crypts are called crypts of Lieberkühn. In the epithelial lining (simple
columnar) identify enterocytes (columnar cells with brush border), goblet cells,
Paneth cells. Note the muscularis mucosa and beneath the submucosa containing
the Brünner's glands. Sketch these organs at L.P. and H.P.
Small intestine proper (Slides 24, 29 and 30). Identify the submucosa and
compare to the submucosa of the duodeneum. Slide 24: Note that this portion
corresponds to the jejunum; Why? Slides 29 and 30: Note that these images
correspond to the Ileum; Why? Make a drawing. Study the tunica muscularis.
What is the serosa? Make drawings.
Large intestine. (Slide 24, colon and rectum; Slide 31, colon). The mucosa
consists only of crypts of Lieberkühn. There are no villi. Sketch the mucosa.
Note the presence of many goblet cells and few striated border cells.
11
ANATOMY AND CELL BIOLOGY
Introduction to Dynamic Histology
Laboratory 10
Objectives
1.
To study the histology of the glands associated with the digestive tract (salivary
glands, pancreas and liver).
Assignment
1.
2.
3.
Salivary glands. Study the organization of compound glands in general. Note the
lobes subdivided into lobules. This subdivision is made by connective tissue as a
capsule with sub-partitions. Note the duct distribution as interlobar, interlobular,
and intralobular. Note that intralobular ducts are either intercalated or striated
ducts. a) Parotid Gland. Study in slide 26 the structure of the parotid gland. All
the acini are serous. Note that the intercalated ducts are short and that the striated
ducts are longer. b) Sublingual gland (slide 18). Most acini are mucous, usually
with serous demilunes. There are also a few serous acini. Study the duct system.
c) Submandibular gland: (slide 13). Most acini are serous, but there are some
patches of mucous acini.
Pancreas (slide 13). a) Exocrine pancreas. Study the serous acini which make up
most of the pancreas. They are typical serous glands with the addition of
centroacinar cell nuclei in their centers. The duct system of the pancreas consists
of long intercalated ducts (low cuboidal epithelium) emptying into small
intralobular ducts with cuboidal epithelium surrounded by a small amount of C.T.
These drain into larger interlobular ducts lined by a columnar epithelium and
surrounded by much C.T. Sketch these structures. b) Endocrine pancreas.
Constituted by small isolated islets of Langerhans surrounded by thin C.T. septa.
The cells within the islets are polyhedral and form cords around capillaries.
Liver (slide 14). Find the capsule of C.T. Identify the following structures: a) The
portal spaces containing a branch of the hepatic artery, portal vein and
interlobular bile duct. b) The central veins (center of the hepatic lobules). c) The
sinusoids and the cords of hepatocytes. Note that the sinusoids open into the
central vein. Identify the bile canaliculi between hepatocytes and the cholangioles
(small ducts linking the bile canaliculi with the interlobular ducts. Make sketches
of these structures.
12
ANATOMY AND CELL BIOLOGY
Introduction to Dynamic Histology
Laboratory 11
Objectives
1.
To study the histology of the kidney
2.
To study the histology of the ureter and urinary bladder
Assignment
1.
Kidney (Electronic Slide 16). L.P. observations: Find at the periphery of the
kidney the capsule and note the separation between the outer cortex and the inner
medulla (the cortex stains more intensely). In the cortex find branches of the
interlobular arteries which delimit the boundary of the lobules. The center of each
lobule usually has a straight medullary ray. H.P. observations: Find a renal
corpuscle and examine the parietal layer of Bowman's capsule, the glomerular
tufts of capillaries containing nuclei of endothelial cells, red blood cells,
mesangial cells, as well as the large light nuclei of podocytes (which correspond
to the visceral layer of Bowman's capsule). Draw a renal corpuscle at H.P.
Find other renal corpuscles which show: a) the junction with the proximal
convoluted tubule; b) the point of contact with a macula densa of the distal
convoluted tubule; c) the point of junction of the glomerular tuft with afferent
and efferent arterioles. Sketch each of these images.
Study the proximal convoluted tubules (which are more numerous than the distal).
The cells are cuboidal and deeply acidophilic and contain round central nuclei.
The lumen is smaller than that of the distal convoluted tubules and is partly
occluded by material. The apical border of the cells has a brush border. Make an
H.P. drawing of a section of this tubule.
Study the distal convoluted tubule. The cells are paler and smaller than in
proximal tubules. There is no brush border and the lumen is wider. The nuclei of
the cells are close to the apical plasma membrane. Draw a section of this tubule.
13
Study the medullary ray which is at the center of each lobule. The main duct in
the ray is the branched collecting tubule. The distal convoluted tubules drain into
it. The duct resembles the distal convoluted tubules, but the apices of the cells
seem to bulge into the lumen. The other tubules in the medullary ray are the
descending and ascending limbs of the loop of Henle. Identify the collecting
tubule, and make an sketch.
In the medulla identify the collecting tubules and the thin ad thick limbs of the
loop of Henle. Note the capillaries between the tubules; these can be confused
with the thin limbs of Henle's loop.
2.
Histology of the ureter. It is a long muscular tube leading from the calyx of the
kidney to the urinary bladder. The lumen is star shaped in cross section and is
lined by transitional epithelium (Section 16 may or may not have a portion of this
organ). The lamina propria is elastic and merges with a submucosa, followed by
the muscle coat made up of 3 layers of smooth muscle (inner longitudinal, middle
circular and outer longitudinal). Find the beginning of the ureter at the hilum of
the kidney.
3.
Histology of the urinary bladder (Slide 10). The bladder is a large muscular bag
for storing urine. It is lined by a transitional epithelium which is capable of great
distension. The smooth muscle coat is irregularly arranged in several layers.
Draw the transitional epithelium under H.P.
14
ANATOMY AND CELL BIOLOGY
Introduction to Dynamic Histology
Laboratory 12
Objectives
1.
To study the histology of the nervous system
Assignment
1.
Spinal ganglion cells (Electronic Slide 22). Study the histology of the spinal
ganglion and find the nerves emerging from it. The spinal ganglion consists of a
collection of the nerve cell bodies, nucleus plus perikaryon of the neurons. These
are the largest cells in the ganglion. The perikaryon is surrounded by a layer of
cuboidal satellite cells which are type of neuroglia. Study the cell body of a
neuron. The nucleus is round, large, located in the center and contains a large
nucleolus. The cytoplasm contains large flecks of basophilic material, Nissl
substance, which is the ergatoplasm (RER) of the cell. The Nissl substance
extends into the larger dendrites, but not into the axon hillock and/or axon. Some
neurons contain dark yellow pigment granules called lipofuscin granules.
Peripheral nerve. Leaving the ganglion is the posterior root of the spinal nerve, a
peripheral nerve containing nerve fibers (myelinated axons) of the neurons of the
spinal ganglion. Define the epineurium, perineurium and endoneurium. Axons
are slightly basophilic. The myelin sheath appears acidophilic and vacuolated.
The larger nuclei, with pale chromatin are the nuclei of Schwann cells. The
thinner, darker nuclei are fibrocytes of the endoneurium. Observe node of
Ranvier.
2.
Cross section of spinal cord (Slide 22). Note the C.T. (pia mater) around the cord.
Outside the pia mater are sections of peripheral nerves (the anterior and posterior
roots of the spinal nerves). Distinguish the gray and white matter. The gray
matter is in the form of an H. It contains the cell bodies of neurons. The white
matter is on the outside and contains the nerve fibers running in the cord. Both
gray and white matter contains nuclei of neuroglial cells. In the center of the cord
15
is the central canal. It is lined by columnar cells, the ependymal cells. In the
anterior horn find the large neurons (anterior horn cells). Make drawings.
In the white matter there are nerve fibers, but the nuclei belong to three types of
neuroglia cells. The large pale nuclei are of astrocytes. The darker nuclei with a
distinctive nuclear membrane are the oligodendrocyte, and the smallest and
darkest nuclei, often irregular in shape are the microglia.
3.
Study the section of the cerebellum (Slide 7). It is a sagital section through the
horizontal convolutions called lobes. Each has numerous secondary folding, each
being a folium. Study one cerebellar lobe; note the folia; find the core of the
white matter in the center of the lobe. Study the gray matter of the cortex. It
consists of three distinct layers. Next to the white matter is the granular layer,
containing numerous nuclei of small neurons. In the middle there is a single layer
of large cells with large dendritic processes extending into the outermost layer
called Purkinge cells. The outermost layer is the molecular layer and it is
composed of numerous neuronal processes and glial cell nuclei.
4.
The cerebrum (Slide 6) shows some convolutions or folds. Identify again the
centrally located white matter and the gray matter on the outside. The cerebral
cortex of gray matter consists of poorly defined layers. An inner layer next to the
white matter, the polymorphic cell layer, consists of neurons of irregular and
variable size. In the middle pyramidal cell layer, the neurons tend to be shaped
like pyramids of differing size (some of these neurons have very large cell
bodies). The outermost layer is the molecular layer consisting of neuronal
processes and glial cells. Note that glial cells are also present in the outer cortical
layers. Also note that the vascular thin layer of C.T. on the outer surface of the
cerebrum is the pia mater and it is the innermost protective membrane for the
brain.
16