histology a microscopic study of human body...

Lab Exercise: Histology (Revised Spring, 2012), of 17

HISTOLOGY A Microscopic Study of Human Body Tissues and Mitotic Cells

Introduction: Histology is the microscopic study of plant and animal tissues. Although all organisms are composed of at least one cell, we will be concentrating on observing cells and tissues of the human body. All organisms are composed of cells. Human body cells are grouped by their similarities in structure and function into tissues. There are more than 200 different types of tissues of the human body and all of these may be categorized into one of four groups: Epithelial tissue, Connective Tissue, Muscle tissue and Nervous tissue. In order to view these tissues, samples were taken from organs. Organs are macroscopic structures which are composed of more than one tissue type and perform a specific function for a multi-cellular organism. Because you will be observing sections of organs, you will see several different tissues in each slide so it is important to scan the slide in order to find the tissue of interest. The thinly sliced sections of organs were mounted and stained with dyes (commonly hematoxylin and eosin) or stains that provide blue and red color to the tissues. In some cases, differential stains are used to stain specific structures. As you view the different slides, it is to your advantage to view all of the slides within each group before observing slides of a different group. For example, you may start with connective tissues so you would view all of the slides or tissues that are grouped within this category identifying common characteristics before moving on to the epithelial tissues. Note: As a novice microscopist it is important to remember a few things! You may only have one slide at your bench at a time. Your single slide should always be placed on the stage and never on your bench or in your notebook. No slide is ‘created equal’. Each specimen on every slide is unique so if you have problems with one slide after a few minutes—review the histology photo album and/or return that slide and get a different slide with a different label. If that does not work, be sure to ask your instructor for advice! Each slide is labeled with a tissue you can find on the slide. However, the specimen that provided the sample often includes several different tissues—so you must scan the slide in order to find the tissue identified on the label.


Activity 1: Microscopic Observation of Epithelial Tissue Epithelial tissue covers body surfaces, lines the lumens of body tubes and lines most of the body cavities. As a cover, there is ‘free’ space on one side of the tissue and other cells located on the other side of the epithelial tissue. As we look at these tissues under the microscopic, one key observable characteristic is to find the white or empty (usually) space on one side and then identify the specific epithelial tissue but the cell shape that is on this apical surface.

The name of the epithelial tissue describes its general structure: the number of

layers and the individual cell shape. The cell shape on the apical surface (next to the free space) is used to identify the tissue without regard to the deeper cell shape. Cells are described as squamous (flat), columnar (tall), cuboidal (roughly cube-like or spherical). Microscopically the different epithelial cell shapes can be identified by the amount of cytoplasm surrounding a stained nucleus. The squamous cells have very little cytoplasm and look very similar to an egg ‘sunny side up.’ Cuboidal cells will have about the same amount of cytoplasm from the nucleus to the edge of the cell membrane in any direction. While, columnar cells commonly have a lot of cytoplasm between the nucleus and the apical surface. Simple epithelial tissues include a single layer of cells, while stratified indicates that there are many layers of epithelial cells. Pseudostratified indicates that the cells appear many layered, but are actually a single layer of cells. Individual cells may be squamous (flat), cuboidal or columnar. Individual cells may also have cellular structures that contribute to their function, such as, cilia or microvilli. Epithelial tissues are attached to an underlying connective tissue by a basement membrane. The basement membrane is composed of substances which are secreted by the epithelial cells and connective tissue cells forming a thin layer marking the margins (boundaries) between the two tissues.

Stratified squamous

squamous

columnar

cuboidal


Procedure: 1) Before Lab, complete the definitions and complete the table in your lab notebook identifying common

locations and functions for each of the following tissues. You will need to use your textbook and/or lecture notes to help you complete this portion of your lab notebook.

2) Before Lab, update your histology atlas in your lab notebook by (pencil) drawing a sketch of the following

epithelial tissues you will be observing in lab.

a. simple squamous epithelial tissue b. simple cuboidal epithelial tissue c. simple columnar epithelial tissue d. pseudstratified ciliated columnar epithelial tissue e. stratified squamous epithelial tissue

3) Use your textbook and the Histology Photo Album On-Line to draw each epithelial tissue as listed below.

Include the following labels for each epithelial tissue within your histology atlas: (You may use a pencil to label any drawings).

a. title of the tissue b. apical surface c. nucleus d. basement membrane e. cell membrane f. cytoplasm g. Other cellular features shown in bold-faced type as you read through the lab manual description of

the tissue should also be labeled on the corresponding drawings.

4) Note: Coloring your drawings of the histology atlas is considered optional. 5) Note: There is space provided in your histology atlas for Notes. This area is for you to make notes to yourself

about the tissue you are observing. In some cases, you may wish to give yourself directions on what to look for in order to find this tissue at a later date.


6) Observe the Simple Squamous Epithelial slide using your high power objective lens.

Simple Squamous Epithelial Tissue 10X Objective Lens This photograph of a sample of kidney tissue clearly demonstrates the white space which is where to look for epithelial tissues under higher powers.

Simple Squamous Epithelial Tissue 40X Objective Lens Simple squamous epithelial tissue is found making up the capsule that surrounds the glomerulus, a capillary bed. The cells are extremely flat, almost line-like with slightly raised purple bumps of the nuclei. The arrow is pointing to the nucleus of a single simple squamous epithelial cell.


7) Observe the Simple Cuboidal Epithelial slide using your high power objective lens.

Simple Cuboidal Epithelial Tissue 10X Objective Lens This photograph of a sample of kidney tissue clearly demonstrates the white space which is where to look for epithelial tissues under higher powers. Simple cuboidal cells make up tubules within the kidney. The lines of white demonstrate the apical surface of simple cuboidal while other tubules are sectioned to see the lumen lined with cuboidal cells.

Simple Cuboidal Epithelial Tissue The simple cuboidal cells form the lumen of the tubule. Cuboidal cells have approximately the same amount of cytoplasm surrounding their nucleus in all directions. You should be able to identify the line marking the basement membrane of the epithelium. Some cuboidal cells of the kidney have microvilli; fine-brushlike extensions of the cell membrane at the apical surface. Microvilli usually require light adjustment and/or use of the oil immersion lens. The arrow is pointing to the basement membrane of a simple cuboidal epithelial cell.


8) Observe the Simple Columnar Epithelial slide using your high power objective lens.

Simple Columnar Epithelial Tissue The simple columnar epithelium can be seen next to the large free surface. The dark pink nuclei are aligned in a row. The basement membrane separates the single layer of columnar cells from the deeper connective tissues.

Under high power, not only are the simple columnar epithelial cells identifiable but there are goblet cells interspersed in many specimens. Scan the free surface and try to identify a goblet cell on your slide. Goblet cells produce mucous and secrete it toe the free surface.


9) Observe the Pseudostratified Ciliated Columnar Epithelial slide using your high power objective lens.

The slide of a specimen from the trachea is the most common example used for pseudostratified ciliated columnar tissue. The epithelial tissue is always located at a free surface, so scan the outer margins of the specimen to locate it. Pseudostratified ciliated columnar epithelial tissues are an example of simple columnar shaped cells, but appear stratified due to the irregular placement of the nuclei in adjacent cells. Under high power (below), numerous goblet cells and cilia are easily viewed. The goblet secrete mucous that traps inhaled dirt, debris and microbes preventing them from entering into deeper airways. The cilia propel the mucous anteriorly toward the pharynx.


10) Observe the skin slide and find the Stratified Squamous Epithelial slide using your low power and high power objective lens.

At the apical surface is the epidermis, darkly stained stratified squamous epithelial cells show the dermal papillary ridges which are the ridges of your finger prints. (The deeper dermal layer, lighter pink stain is composed of dense irregular connective tissue.)

Under high power, the stratified squamous epithelium is named for the cell shape at the apical surface. Toward the basement membrane, the cells become more rounded but are still part of the stratified squamous epithelium. The epithelial cells nearest the basement membrane are highly mitotic and producing new cells which replace the dead cells that are constantly sloughed off the surface.


Activity 2: Microscopic Observation of Connective Tissue Connective tissues include a wide variety of different tissues that function to protect, support and bind together other body tissues. Connective tissues are found throughout the body and are the most abundant tissue type! There are two general characteristics common to all connective tissue: (1) an extracellular matrix and (2) unique cells. The extracellular matrix is the substance located outside of the connective tissue cells. The ground substance includes water and adhesion proteins that allow the connective tissue cells to attach. Scattered throughout the matrix are proteins (fibers) and include collagen (strength), elastin (stretch and recoil) and reticular fibers (fine collagen fibers). The amount of extracellular matrix and its composition will vary from one connective tissue to another. The unique cells of connective tissue are largely responsible for the production and maintenance of the matrix. In many instances, the cells are separated by large amounts of matrix, while other connective tissue cells lie close together with very little matrix in between. Procedure: 1) Before Lab, complete the definitions and complete the table in your lab notebook identifying common

locations and functions for each of the following connective tissues. You will need to use your textbook and/or lecture notes to help you complete this portion of your lab notebook.

2) Before Lab, update your histology atlas in your lab notebook by (pencil) drawing a sketch of the connective

tissues you will be observing in lab.

a) Your histology atlas should include a labeled diagram of Compact bone Hyaline cartilage Adipose Blood

b) Each connective tissue will have a variety of unique anatomical features to label. Be sure to read the

descriptions closely and label all (bold faced) anatomical features, including cell types and matrix, for each connective tissue slide. Refer to your textbook and the On-Line Histology Photo Album to check your correctly labeling the anatomic features of each connective tissue.

c) Note: Coloring your diagrams is considered optional.

d) There is space provided in your histology atlas for Notes. This area is for you to make notes to yourself



3) Observe the Ground Compact Bone slide using your high power objective lens. Identify the different parts of the tissue as described.

Compact bone looks very much like a cross section of a tree—the rings of the tree correspond to rings of calcium salts and collagen fibers of the matrix. Each ‘tree’ is an osteon—a cylindrical unit of compact bone. At the center of the rings is a space, the central canal, which may have blood vessels and/or nerves passing through. Arranged within the concentric rings of matrix are the bone cells, osteocytes. Each osteocyte occupies its own small space, called a lacuna. Extending from the lacuna are tiny canals, canaliculi, which allow a conduit for exchange between cells and between the osteocytes and its blood supply.

In order to see an osteocyte within a lacuna, it may be necessary to adjust the light (iris diaphragm lever) or use the fine focus and take the slide in and out of focus so that the osteocyte ‘pops’ out

This photo has a different staining technique than you will view in lab, the matrix is stained black. The osteocytes (cells) are located within the lacunae (white space) which are interconnected by small canals/canaliculi.


4) Observe the Hyaline Cartilage slide using your high power objective lens. Identify the different parts of the tissue as described.

Hyaline cartilage is the most abundant cartilage—it contributes a flexible structure to the nose, larynx, trachea, ends of bones and makes up the fetal skeleton. Hyaline cartilage is much more flexible and less rigidly arranged than bone. Hyaline cartilage may be stained with a pink or purple stain. (Refer back to the trachea image for the purple stain). As you view the slide of hyaline cartilage, the cells are irregularly spaced apart. Each cell, chondrocyte, occupies a space within the matrix called a lacuna.

The matrix contains abundant collagen fibers but due to the other components of hyaline, it gives the matrix a ‘glassy’ appearance.

5) Observe the Adipose slide using your high power objective lens. Identify the different parts of the tissue as

described.

Adipose tissue looks like chicken wire microscopically or each cell looks like a signet ring. The cytoplasm of the adipocyte, adipose cell, is filled with a fat vacuole that stores fat for the body. The nucleus is often pushed snug against one side of the cell membrane to allow the fat vacuole to expand. Adipose tissue is a modified areolar connective tissue with all types of fibers between the cells. However, due to the function as long-term fat storage, there is very little space between individual cells so you will not see much matrix between the adipocytes of adipose tissue.

Blood vessel


6) Observe the Blood Smear slide using your high power objective lens. Identify the different parts of the tissue as described.

Blood is the only fluid or liquid connective tissue. The extracellular matrix is composed of blood plasma. The formed elements (cells) of blood include erythrocytes (red blood cells) and leukocytes (white blood cells). The erythrocytes are composed of cell membrane and their cytoplasm is filled with hemoglobin which give them their pink appearance. Upon maturation, erythrocytes eject their nucleus in order to maximize the intracellular space for hemoglobin and oxygen carrying. Leukocytes are only visible when stained. There are a variety of different types of leukocytes found in blood. Leukocytes are identified by the shape of their nucleus, size and presence of granules (stained storage areas). You will note that the number of erythrocytes greatly outnumber leukocytes.

In addition, thrombocytes (blood platelets) are small cell fragments may be seen between the formed elements of blood.


Activity 3: Microscopic Observation of Muscle Tissue There are three different types of muscle tissue—each is specialized to contract (shorten) producing movements. Cardiac muscle is only found in the heart and its contractions are used to pump blood through arteries. Smooth muscle contractions cause the movement of substances within body tubes food through the esophagus or small intestine. Contractions of the smooth muscle making up arterial walls contribute to maintaining blood pressure. Skeletal muscle carries out voluntary contractions of body movement like walking and talking. Procedure: 1) Before Lab, complete the definitions and complete the table in your lab notebook identifying common

locations and functions for each of the following muscle tissues—skeletal, smooth and cardiac muscle. You will need to use your textbook and/or lecture notes to help you complete this portion of your lab notebook.

2) Before Lab, update your histology atlas in your lab notebook by (pencil) drawing a sketch of the muscle

tissues you will be observing in lab.

a) Each type of muscle issue will have a variety of unique anatomical features to label. Be sure to read the descriptions closely and label all anatomical features (bold-faced) for each muscle tissue slide and use your textbook and On-Line Histology Photo Album to help in your identifications.

b) Note: coloring your diagrams is considered optional.

c) There is space provided in your histology atlas for personal Notes and observations.

3) Observe the Skeletal Muscle slide using your high power objective lens. Identify the different parts of the tissue as described.

Skeletal muscle is composed of long cylindrical cells. The individual cells can be as long as the muscle itself. As the muscle length increases, the individual cells get ready to divide, double their cytoplasmic contents including nucleus, but they do not complete cytokinesis thus ending up as long multinucleate cells. By finding an area on the slide of the muscle fiber running longitudinally (lengthwise) you will note that you cannot see the ‘ends’ of the cell.

Running perpendicular to length of the cell you should see striations (stripes) composed of the proteins actin and myosin. These proteins carry out the contraction (shortening) of the cell.


4) Observe the Smooth Muscle slide using your high power objective lens. Identify the different parts of the tissue as described.

The cells making up smooth muscle are spindle shaped, long with narrowed ends. Smooth muscle cells are not nearly as long as skeletal muscle fibers (cells). The photograph shows the darker stained nuclei within the individual smooth muscle cell cytoplasm. Smooth muscle is often arranged in layers—running in different directions. In this photograph, there is just one layer running mostly vertically. Smooth muscle contains actin and myosin but it is arranged in the same fashion as skeletal muscle so smooth muscle cells lack striations

.

5) Observe the Cardiac Muscle slide using your high power objective lens. Identify the different parts of the

tissue as described.

Cardiac muscle cells are much shorter in length and uninucleate (single nucleus). As the cardiac muscle cells are arranged, they form branching patterns. Connecting adjacent cardiac muscle cells are specialized cellular junctions called gap junctions called intercalated discs which are darker pink than the striations seen throughout the cytoplasm of the cardiac muscle cell.


Activity 4: Microscopic Observation of Nervous Tissue Nervous tissue is composed of two general cell types: (1) neurons and (2) glial or support cells. Neurons are specialized to conduct an electrical impulse called an action potential across its length. These action potentials are responsible for our being able to sense our environment, process this information and carry out an appropriate response in order to maintain homeostasis Procedure: 1) Before Lab, complete the definitions and complete the table in your lab notebook identifying functions for the

different parts of a neuron. You will need to use your textbook and/or lecture notes to help you complete this portion of your lab notebook.

2) Before Lab, update your histology atlas in your lab notebook by (pencil) drawing a sketch of the neuron smear

you will be observing in lab and label the anatomical structures (bold-faced) common to neurons. Use your text book to aid you in your identifications.

a) There is space provided in your histology atlas for Notes. This area is for you to make notes to yourself


3) Observe the Neuron Smear slide using your low power objective lens. Identify the different parts of the tissue as described.

Neurons are large cells surrounded by much and smaller and numerous glial cells. The cell body of a nueron includes most of the cytoplasm and nucleus. The nuclei are large, highly visible, and often you will find the nucleolus within the nucleus that looks like an ‘owl’s eye’. Extending from the cell body are long cellular extensions called processes. Different processes are responsible for different parts of the action potential or conduction of the electrical impulse. Neurons, including their cellular processes, can be very long—you actually have a neuron which is up to 3 feet long running from the terminal end of your spinal cord to your big toe! Each neuron has one axon. The axon typically identified because it is the largest process attached to the neuron’s cell body. Dendrites are all of the other cellular extensions. Dendrites are responsible for sensing the environment and starting an electrical impulse which travels toward the cell body. Axons are responsible for continuing the action potential along its length away from the cell body.


Activity 5: Mitosis In order to increase in body size, we need to increase our cell number; or, tissue repair involves replacing cells with new cells. Creating new cells is achieved through cell division. Cell division includes nuclear division (mitosis) and cytoplasmic division (cytokinesis). Mitosis is the type of cell division that creates new cells involved in both growth and tissue repair. Mitosis results in genetically identical cells called clones. These cells have the same number and kind of DNA, as the parent cell. The cell cycle which includes all of the phases a cell undergoes from its formation until it divides. Most of a cell cycle, the cell is in Interphase. Where the cell is undergoing normal metabolic activities—carrying out its designated function, making proteins, synthesizing ribosomes, accessing the genetic code stored in DNA and replicating DNA. The DNA is accessible to nuclear proteins to make cytoplasmic proteins and ribosomes, it appears granular microscopically and is called chromatin. Within the nucleus are darkened condensed areas where ribosomes are being synthesized so the nucleoli are clearly visible. Separating the chromatin from the cytoplasm is the nuclear envelope. Mitosis, nuclear division, describes how the two sets of DNA which is formed during interphase are divided so that each daughter cell has its own complete set of genetic instructions. Mitosis includes the following phases: Prophase, Metaphase, Anaphase and Telophase. Prophase is the stage where the strands of DNA are packaged into chromosomes. During prophase, centrioles are assembled from cytoplasmic proteins, the nucleolus disappears, and the nuclear membrane is disassembled. Metaphase is identified by the alignment of the chromosomes at the cells equator. An animal cells, centrioles have migrated to opposite poles and connecting the centrioles to the chromosomes are long proteins fibers called the spindle apparatus. Anaphase occurs as the chromosomes are moved by the spindle apparatus towards the centrioles. One DNA molecule and it’s replicate (copy) are moved so that each new daughter cell has the same genetic information to begin metabolism immediately upon cytoplasmic division. Telophase is marked by the unpackaging of the chromosomes back into its accessible form, chromatin. The nuclear membrane is reassembled, nucleoli reappear and both the centrioles and spindle apparatus is disassembled.


Procedure: Mitosis 1) Before Lab, complete the descriptions of the cellular events that occur during all stages of the cell cycle. Label the diagram for each mitotic phase. 2) Before Lab, prepare your lab notebook, by drawing interphase and each stage of mitosis for an onion cell

in your histology atlas. Label the anatomical structures that are visible during each phase. 3) During lab, observe an onion mitosis slide has several different root tip specimens on it; each root tip has

many cells which are in various stages of division. So, as you view a root tip microscopically, you will most likely find several different mitotic phases in one field of view.

Using your scanning objective you will note that one end of the onion root tip is pointed while the other is level or torn. The area of mitosis will be best viewed toward the root tip. See the red boxed area.

INTERPHASE The nucleus has a granular appearance where the DNA is called chromatin. In many cases one or more darkened areas of the nucleoli and nuclear membrane. Remember, the cell membrane of a plant cell is also surrounded by a thick cell wall.

PROPHASE The nuclear membrane has been disassembled and the chromatin is condensing into chromosomes.

METAPHASE The chromosomes have aligned at the cells equator and the spindle apparatus can be seen. (Note there are no centrioles).

ANAPHASE The chromosomes are being separated and moving by the spindle apparatus to opposite poles. (On the right hand side are two onion cells in interphase, while the upper left hand cell is in prophase).

TELOPHASE The chromosomes are being unpackaged and losing the linear (noodle) appearance. At the cell’s equator a new cell wall is being laid down, so cytokinesis is a concurrent event.

Filename: 07_Histology_1_ Directory: E:\LabNotebook Template: C:\Users\karen\AppData\Roaming\Microsoft\Templates\Normal.dotm Title: HISTOLOGY Subject: Author: klchambe Keywords: Comments: Creation Date: 4/30/2012 11:10:00 AM Change Number: 7 Last Saved On: 5/15/2012 2:15:00 PM Last Saved By: karen Total Editing Time: 74 Minutes Last Printed On: 5/15/2012 2:16:00 PM As of Last Complete Printing Number of Pages: 17 Number of Words: 3,971 (approx.) Number of Characters: 22,639 (approx.)

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