Histology and Cytology PDF

1 HISTOLOGY Histology or microscopic anatomy is the study of the fine and minute details of the organism with the help of a microscope. It is the study of the tissues of the body. The term histology was derived from the Greek terms, ―histos = tissue‖ a...

1 HISTOLOGY Histology or microscopic anatomy is the study of the fine and minute details of the organism with the help of a microscope. It is the study of the tissues of the body. The term histology was derived from the Greek terms, ―histos = tissue‖ and ―logos = study‖. The etymologic derivation of the English word ―tissue‖ is from the French tissu, which means texture or weave. This word was introduced into the medical sciences by French gross anatomist and physician, Marie F. X. Bichat, in the late 18th century. He is considered as the father of histology. It is interesting to note that this brilliant anatomist did not use a microscope to found this discipline. The term histology was coined by AFJK Mayer (1819). Histology can be subdivided into (1) General histology that deals with the structure of fundamental tissue types (epithelium, connective tissue, muscular tissue and nervous tissue) and (2) Special histology or organology or systemic histology that deals with the fine structure of organs. A sound knowledge on histology is essential for understanding physiological changes occurring in cells and for understanding the changes during different pathological conditions. CYTOLOGY Cytology is the study of cell. Cell is the smallest structural and functional unit of the living body. The word ―cell‖ was introduced by Robert Hooke (1665). Cytology now employs the term protoplasm to denote the entire living substance of the cell which is the physical basis of life. This includes the cell body and its extensions and the nucleus. The substance of cell outside the nucleus is called the cytoplasm and the substance of the nucleus is the karyoplasm or nucleoplasm. The cytoplasmic organelles and inclusions are suspended in cytoplasmic matrix or cytosol. The cell theory explains that all the organisms are made up of cells and all the cells come from pre-existing cells. Cytology or cell biology correlates the structure of cells with their functions and all the life processes. In multicellular organisms, certain cells become specialized to perform specific functions and these cells having common origin and performing similar but specific functions form a tissue. In other words, tissue is defined as a group of more or less similar cells and their extracellular products that perform a specific function or a spectrum of related functions. Several types or a group of such tissues collectively carry out a specific task to form an organ and several similar inter-related organs constitute an organ system. All the new of an organism develop from a pre-existing cell and is capable to give rise to a new individual. This is known  2 as ‖totipotency‖. Each cell is made up of several organelles and cell performs all its functions through the organelles present in the cytoplasm. Thus the activities of an organism are present in primary form in each and every cell. Hence cell is the basic structural and functional unit of life and of an organism. Historical background Robert Hooke (1665) observed a honey comb like pattern in a very thin slice of a cork which he called cellulae or cells. Anton van Leeuwenhock (1683) was the first to observe free cells with the help of a microscope that he developed. It is Alfonso Corti (1772) to report first the living substances inside the cells. In 1831, Robert Brown reported the presence of small sphere in the cells of orchid root, later called as the nucleus. Subsequently, Hugo Von Mohl (1840) and Thomas Huxley (1839) discovered the existence of jelly like fluid in cells, the protoplast or protoplasm (first life). It is the combined work of Mathias Schleiden (1838) and Theodor Schwann (1839), German Botanist and Zoologist brought about the first part of cell theory that all organisms are made of cells. Rudolf Virchow (1863) and Nagali were the first to explain that all cells come from pre-existing cells of similar origin. The cells vary in their shape, size, structural modification and in functions. As against the unicellular organism, in muticellular organism, there is unique co-ordination of cells and tissue systems and have a dual existence. Even if some cells die, others can multiply and replace them. The cells of multicellular organism can differentiate to extreme level of specialization even though they are from a single zygote and cells exist as undifferentiated cells (stem cells or meristematic cells). Differentiated cells are post mitotic cells (RBCs) and dedifferentiated cells which are capable of reversing the differentiation (wound healing tissues). Cell Organelles and Discoverers Protoplasm - Felix Dujardin discovered in 1835 Cytosol - Albert Claude, Christian de Duve and George E. Palade in 1935 Nucleus - Robert Brown in 1833 Nucleolus - In 1781 Felice Fontana described the nucleolus after finding it in the slime from an eel's skin. Nucleoplasm – Edouard Adolf Strasburger "New cell nuclei can only arise from the division of other nuclei." and gave the terms cytoplasm and nucleoplasm in 1882  3 Golgi body Camillo Golgi (1843-1926) while working as a physician developed a silver-osmium technique, the "reazione nera" (black reaction), for which he was awarded the Nobel Prize in 1906. In the late 1890's, 25 years after the publication of his black reaction he noticed a fine internal network in only partially silver-osmium-blackened Purkinje cells of cerebellum. Golgi published the discovery, called the "apparato reticolare interno", in 1898, which is now considered as "Golgi apparatus". Mitochondria The first recorded study ever made in the mitochondria was made by Richard Altman in 1840. He came to know about them and called them bioblasts. Albert von Kolliker discovered the existence of mitochondria around 1857. He was studying human muscle cells when he noted strange granules in them. Carl Benda coined the word mitochondria in 1898. He got it from the Greek word mitos, meaning thread, and chondrios, meaning granule. Endoplasmic reticulum - Albert Claude in Belgium and Keith Porter at Rockfeller Institute discovered endoplasmic reticulum in 1945. Claude derived the term reticulum from the Latin word for "fishnet", (membranous network within cells), and he was among the first scientists to use an electron microscope for cellular study. Ribosomes (Animal cell) – George Emil Palade Early electron microscopy (EM) was troubled by, as George Palade put it, ―the perennial and arduous question of artifact versus reality.‖ Stains and fixatives could precipitate— Keith Porter referred to this as ―the coagulating action of the fixative‖—and produce structures that were not present in the original sample. But when Palade noted a particulate component of the cytoplasm, he confirmed its presence using two different fixatives, and described its particular abundance in embryonic, rapidly proliferating, and glandular cells (Palade, 1955). Thus were born the particles of Palade, later known as ribosomes. Cytoskeleton - Nikolai K Koltsov in 1903 proposed that the shape of the cell is determined by a tubular network, the cytoskelton. However, the word was introduced and coined by Paul Wintrebert in 1931 (in French, cytosquelette) i. Microtubules - De Robertis and Franchi disovered microtubules in 1953 in nerve cell and later Sabatani, Bansch, Barnette in 1963 explained the structure of microtubule. ii. Microfilament/Actin Filaments - Edward David Korn discovered microfilament in 1968 in Acanthamoeba castellanii.  4 iii. Intermediate Filaments - The group of Howard Holtzer in 1968 discovered intermediate filaments. Lysosomes - Discovered by Christian de Duve in 1949 Vacuole - Antony van Leeuwenhoek is credited with discovery of vacuole when he was studying bacteria in late 1500s or early 1600s. Centriole and Centrosome - Discovered by Edouard Van Benden in 1883 and was described and coined by Theodor Boveri in 1888. Astral rays and spindle – Discovered by H. Fol in 1873 Cilia - the oldest known cellular organelle, first described in 1675 by Anton van Leeuwenhoek in protozoa. He described them as ‗incredibly thin feet, or little legs, which were moved very nimbly‘. The term ‗cilium‘ (Latin for eyelash) was probably first coined by Otto Muller in 1786. Physiological / Vital / Biologic properties of protoplasm 1. Irritability: It is the basic property of protoplasm that enables it to react to stimuli. In nerve cells, this property is greatly developed. 2. Conductivity: It refers to the ability of the protoplasm to transmit a wave of excitation from the point of stimulus. This property is well developed in nerve cells and muscle cells. 3. Contractility: Refers to the ability of protoplasm to alter in such a way that the cell becomes shortened in one direction. This property is highly developed in muscle cells. 4. Absorption and assimilation: Cells can take in certain dissolved substances into their protoplasm. The capacity of absorption is highly developed in intestinal epithelium. Assimilation is the ability to breakdown and utilize the ingested material. 5. Secretion: Some cells can produce new products eg. gland cells. 6. Excretion: It is the ability to extrude waste products. 7. Respiration: Take in oxygen which is used for oxidation of food substances. From this reaction energy is obtained. 8. Growth and reproduction: All cells are capable of growing and reproducing cells of same type. 9. Motility:  5 a) Protoplamic streaming: Internal streaming occurs in immobile cells eg: plant cells. b) Amoeboid movement (Phagocytosis and pinocytosis): Through pseudopodia eg: Amoeba, WBC, macrophages.. c) Ciliary and flagellate lashing: eg. Ciliated epithelia, spermatozoa, flagellate protozoans etc. d) By contractility: eg. Muscle fibres. Unicellular organism can generally exhibit all the above properties. Physical Characteristics of Protoplasm: Protoplasm is a semifluid, viscous and more or less transparent substance. It is a mixture of crystalloids and colloids. A colloid is an aggregate of atoms or molecules that is dissolved in but is seperatated from the solvent phase and contains particles of size sufficient to prevent their passing through semipermeable membranes. eg. Glucose. Chemical Composition of Protoplasm Protoplam contains a high proportion of water with proteins, carbohydrates, fats, vitamins, enzymes and inorganic salts. Chemical analysis shows the presence of carbon, hydrogen, oxygen and nitrogen as the major constituents. Potassium is the primary protoplasmic cation where as phosphate, bicarbonate and sulphate are the primary protoplasmic anions. Sodium and chloride are the primary extraprotoplasmic ions. Heavy metals like iron are also seen in traces. All these are associated in watery medium. Certain tissues contain more concentration of one element eg. Ca in bone, iodine in thyroid gland and Fe in RBCs. Nucleic acids, proteins, carbohydrates and lipids are the most commonly occurring biochemical substances of the protoplasm. These can occur as macromolecules which are composed of smaller monomeric subunits. 1. Nucleic Acids: There are two kinds of nucleic acids: a) Deoxy Ribo Nucleic Acid (DNA): Found in nucleus. b) Ribo Nucleic Acid (RNA): Found both in nucleus and cytoplasm. 2. Proteins: They are distributed in the body as structural proteins, enzymes and some hormones. Proteins of biological importance are a) Simple protein (albumins and globulins) b) Conjugated protein (Nucleoproteins, mucoproteins and lipoprotein).  6 3. Polysaccharides: eg. Glycogen and important mucopolysaccharides which are present in the intercellular substance (eg. Chondroitin sulfate and hyaluronic acid.). 4. Lipids: Most of the lipids occur as droplets of neutral fat stored in adipose cells. Certain cells contain complex lipids such as sterols and phospholipids. Structure of cell The cells in the body vary greatly in size, shape and structure. These various adaptations are for the different functions which the cells perform in various tissues and organs. However most of the cells retain a number of features in common. Cytoplasm / Cytoplasmic matrix / Cytosol: It is a structureless solution of proteins, polypeptides, solutes, enzymes, ions, water and various other substances. It is homogenous and translucent in living cells, cytoplasm contains a number of formed bodies embedded in it. These formed bodies fall into two groups. 1. Organoids/organelles: are localized specializations of the cytoplasm (composed of differentiated cytoplasm) which perform some special function related to metabolic or other activities of the cell. These are the permanent constituents of the cell. 2. Inclusions: metabolic products are ingested substances not playing direct part in the metabolic activity of the cell.  7 CYTOPLASMIC ORGANELLES Organelles are small structures whose particular organization gives them a specific function in the metabolism of the cell. 1. Plasma membrane / Cell membrane / Plasmalemma / Cytolemma It covers the surface of the cell. It is a semipermeable membrane through which the interchange of materials between the cell and its environment occurs. It measures 8-10 nm (80 to 100 Ao) The plasma membrane contain about 35% lipid including the phospholipids and cholesterol, 60% protein and a small amount of carbohydrate.This observation as well as physical measurement of membrane thicknesss, birefringence, X-ray diffraction and surface tension lent support to a model of membrane structure proposed by Danielli and Davson in 1935 known as bilaminar theory or unit membrane model or Amphipathic model. This model depicts the plasma membrane as a double layer of bimolecular leaflet of lipid sandwitched between the two protein coats. The phospholipid molecules of the membrane have hydrophobic (non-polar) ends, where the fatty acids are located and the hydrophilic (polar) ends, where the phosphate groups are attached. It is  8 assumed that the hydrophilic ends lie next to the enveloping protein layers. The location of protein in this model proved to be erroneous. It seems that a membrane with continuous lipid leaflets could not provide for all the diverse functions that membrane known to perform. It is now believed that there are scattered sites which provide for special membrane activities. (eg. The attachment of cytoplasmic filaments or the transport of sugars and amino acids across the membrane). At these sites, protein components apparently penetrate the lipid interface of the membrane. This is better explained by the newly introduced fluid mosaic theory of Singer & Nicholson (1972). Fluid-Mosaic model of Singer & Nicholson (1972) This is the modern universally accepted theory described by Jonathen Singer and Garth Nicholson. This model presents the membrane with a continuous lipid bilayer having integral protein molecules. Here the membrane is semifluid and dynamic. Lipids and protein molecules help in trans-membrane transport. The peripheral proteins are loosely connected to the membrane and is easily removed by aqueous solutions. The other proteins are integral proteins and some of such protein molecules project on both sides of membrane and help to transport water soluble substances. These are channel proteins. Long chained protein molecules that traverse the whole membrane and project at both surfaces form the transmembrane proteins which on the surface facing the exterior (cytosolic face) carry oligosaccharides and form glycocalyx. In intestinal absorptive cells, glycocalyx is particularly dense and contains various enzymes that enhance absorption. The plasma membrane also has receptor sites carrying receptor proteins or intrinsic proteins for different types of chemicals such as hormones and neurotransmitters. The extrinsic proteins or resistance proteins may be covalently attached to fatty acids or non-covalently attached to transmembrane proteins. Functions: 1. Acts as a limiting membrane for all cells. 2. It is a protective envelope for the cell. 3. It serves as selective gateway and regulates inflow or outflow of materials. 4. It contains a variety of enzymes. 5. Permits osmosis. 6. Takes in useful substances by endocytosis and gives out wastes and cell products. 7. Specific surface receptors mediate diverse functions such as antigen recognitions, phagocytosis and antibody production.  9 8. Transmission of impulses in nerve cells and muscle cells. 2. Endoplasmic Reticulum (ER) The word ―Endoplasmic Reticulum‖ is coined by Keith Porter (1953). It is a network of double membranes irregularly distributed in the cytoplasm. It is absent in prokaryotic cells and mature mammalian erythrocytes. Two varieties 1. Rough ER (rER) or Granular ER (This has attached ribosomes). 2. Smooth ER (sER) or Agranular ER. Morphologically ER may exist in three forms. 1. Lamellae or cisternae: They are groups of flattened and unbranched sacs arranged in parallel stalks. They are mostly granular. 2. Vesicles: These are rounded or oval sacs. 3. Tubules: They are branching and interconnecting channels. Vesicles and tubules are mostly agranular. ER is believed to originate from nuclear envelope. Granular membranes are believed to appear first and later on they give rise to agranular membranes. The membranes of ER have lipoprotein composition and fluid-mosaic organization. Aggregates of rER appear as basophilic regions within the cells such as nerve cells and pancreatic acinar cells. These regions are referred to as ergastoplasm or chromidial substance. Main function of rER is biosynthesis of proteins. Thus rER is abundant in protein secreting cells such as pancreatic acinar cells and plasma cells which secrete antibodies which is a protein. Another function of rER is glycosylation of proteins to form glycoproteins. The sER (ribosomes are absent) participate in a variety of functions: 1. Steroid hormone synthesis (in testicular interstitial cells, cells of CL and those of adrenal cortex). 2. Synthesis of complex lipids and drug detoxification in hepatocytes. 3. Lipid resynthesis in the intestinal absorptive cells. 4. Release and capture of Ca++ ions in striated muscle cells and concentration of chloride ions in gastric parietal cells for Hcl production (there is an extensive network of sER which is known as sarcoplasmic reticulum in striated muscles).  10 Ribosomes Ribosomes are tiny granular particles of ribonucleoprotein (RNA and protein). With the exception of mature RBC, they are found in all prokaryotic and eukaryotic cells. Some of them are freely dispersed in the cytoplasm and some others remain attached to ER and outer nuclear membrane. Ribosomes take their origin from nucleolus. In the free form these can be seen as clusters attached on single RNA called polysomes or polyribosomes or arranged in circular fashion known as rosette ribosomes. The ribosomes are basophilic in reaction due to RNA. In some cells like nerve cells, large quantity of ribosome and rER are seen throughout the neuroplasm and these are known as Nissl substance or Nissl‘s bodies or chromatophilic substance. Ribosomes are composed of two subunits, a larger subunit and a smaller subunit. The larger subunit contains three molecules of rRNA and approximately 45 different proteins and smaller subunit consists of one molecule of rRNA and 33 different types of proteins. Prior to protein synthesis, these two subunits exist as separate entities. Ribosomes are factories of protein synthesis. Golgi Complex (GC) or Internal reticular apparatus When stained with silver salt or chromium, the GC appears as a black network of 4 to 8 cisternae more often situated above the nucleus. In routine preparations, it may be visible as a lighter stained region called the negative Golgi image. GC exixts in three forms. 1. Lamellae / cisternae 2. Vesicles 3. Tubules. It has a concave face known as trans face or mature face and a convex face known as cis face or immature face. (In some cells, GC is seen continuous with ER, especially rER. Then that ER is termed as GERL or Golgi Associated ER). Functions 1. It acts as a packaging unit. The protein synthesized in the rER are stored, chemically processed and packed for secretion when it gets concentrated. Then membrane vesicles arise from the concave face of GC, packed with the secretion, detached to the cytoplasm move to the cytoplasmic membrane, then it will fuse with cytoplasmic membrane and secreted out. ie. storage, chemical processing and packaging of secretory products. 2. Synthesis of phospholipids and glycolipids  11 3. Formation of acrosome of sperm and cortical granules of ovum. Lysosomes Lysosomes are membrane bound vesicles without any internal structural organization. Also known as the digestive apparatus of the cell or ―suicidal bags‖. They contain various hydrolytic enzymes (more than 50). Lysosomal membrane (single limiting membrane) is highly resistant and impermeable to them. The membrane contains stabilizers eg. Cortisol, cortisone, cholesterol etc. which make the membrane stable and impermeable; and labilizers, eg. Vitamin A, B & K. progesterones, testosterones etc. which destabilize the membrane and make it permeable to enzymes. Lysosomes are polymorphic structures. Two types, primary lysosomes and secondary lysosomes. Primary lysosomes are inactive and contain nothing but hydrolytic enzymes, whereas secondary lysosomes are the result of the fusion of primary lysosomes with a variety of membrane bounded substance ie. phagosomes or phagocytosed bodies. Phagosomes + primary lysosome = secondary lysosome. Lysosomes are highly concentrated in phagocytes and cells of reticulo-endothelial system. Functions 1. Heterophagy – digestion of extracellular materials 2. Autophagy – self digestion Mitochondria Discovered by C. Benda. They are the centres of cellular respiration and energy production. They cannot be demonstrated by routine methods. In the light microscope, they can be demonstrated with the stain, Janus Green B (supravital stain). They are double walled fluid filled bags either rod- or sphere-shaped. The dense central core or matrix is enveloped by two membranes. The space between these two membranes is known as intermembrane space. The inner membrane has numerous hollw infoldings into the matrix known as mitochondrial crests or cristae. Inner surface of the inner membrane presents numerous stalked particles known as elementary particles / F1 particles / Fernandes Moran subunits / Electron Transport particles / ETP. Each particle has three parts: 1. the basal plate. 2. Stalk / stem. 3. Head. These particles are the store houses of enzymes of oxidative phosphorylation and ATP production. Mitochondrial matrix contains several kinds of enzymes of TCA  12 cycle, 2 to 6 circular DNA molecules (viral type), small amounts of mRNA, tRNA and a few ribosomes. This is the only place where DNA is located other than the nucleus. Mitochondrial DNA can as an extrachromosomal gene known as plasma gene (capable of replication). Functions Cellular respiration and ATP production. So it is known as the power house of the cell. Depending on the physiological activity of cell, the structure of mitochondria may vary. Less active (orthodox) ones have large area of matrix. In active (condensed) ones, cristae are more randomly distributed. They can act produce own protein within the organelle. Cell centre / Centrosome / Cytocentre This occupies a central position in the animal cell. (Sometimes it is displaced by synthesized cytoplasmic products or nucleus). Located near the nucleus. Within the centrosome, a pair of deeply staining particles are seen. They are the centrioles which are placed at right angle to each other. Centriole is the only cytoplasmic organoid that reproduces itself exactly (self replicating). EM shows that each centriole is a hollow cylinder closed at one end and consists of nine tubules arranged like a loosly assembled barrel. Each tubule is made up of three subtubules. Functions Centrioles are actively invoved in cell division and formation of cilia and flagella. Peroxisomes These are membrane-bounded spherical organelles containing fine granular materials which are particularly abundant in hepatocytes and cells of PCT. They are a major site of oxygen utilization and are rich in catalysts and hydrogen peroxide. Cytoplasmic Inclusions They are non-living bodies seen in cytoplasm. Common inclusions are 1. Nutritive substances. eg. a. Glycogen: It is the major storage form of carbohydrates and is abundant in liver and skeletal muscle cells. It can be selectively demonstrated by PAS (periodic acid- Schiff‘s) reaction or Best‘s carmine stain.  13 b. Lipid: It is stored primarily in adipose cells. It cannot be demonstrated by routine methods because most techniques involve the use of fat solvents like xylene or benzine. Special techniques used are osmic acid fixation, Sudan-III stained frozen sections, Oil-Red-O etc. c. Protein: Sometimes free protein can be seen in the cytoplasm (in addition to structural components). 2. Pigments: eg. a. Melanin: Occurrs primarily in eye and integument (dark brown to black). It is synthesized by melanocytes. b. Haemosiderin: Formed by the degradation of Hb following phagocytosis of RBC in spleen, liver, bone marrow etc. Golden brown in colour. c. Lipofucsin (wear and tear pigment): It is golden brown but stained only with fat stains. Commonly found in cardiac muscle, liver etc. and composed of indigestible residues of phagocytosis. Foreign substances are seen in the cytoplasm which may be ingested particles like micro-organism, carbon particles, cellular debris etc. Cytoskeleton I. Microfilaments: eg. 1. Actin. 2. Myosin in muscle cells. 3. Neurofilaments in nerve cells and neuroglial cells. Functions: 1. Help in cell movement. 2. Help the cell to change its shape. II. Microtubules: Microtubules are hollow tubules of protein known as ―tubulin‖. They form centrioles and cilia. Cytoskeleton form complex mesh work that maintain cell shape and stability, cell movement with the help of filaments and intracellular transport. Microtubules have a vital role in the movement of organelles, (eg. Movement of chromosomes during cell division). These tubulins can be disturbed with chemicals such as colchicines which will arrest the movement). III. Tonofibrils: Tonofibrils (intermediate filaments) are made up of durable protein. They are present in cytoplasm around the nucleus or intercellular junctions. Nucleus Nucleus was discovered by Robert Brown (1833). Nucleus is the most distinct part of the cell staining well with basic stains like haematoxylin. It is absent in bacteria  14 and mature mammalian erythrocytes. Shape of the nucleus varies. In rounded and cuboidal cells, the nucleus is spherical. In columnar and spindle shaped cells, it is elongated; in neutrophils, it is lobulated; in monocytes, it is kidney shaped. Number of the nucleus also varies. Most of the cells are mononucleated. Example for binucleated cells are the epithelial cells of urinary bladder and some hepatocytes (parietal cells of stomach are sometimes multinucleated). A nucleus has two periods of life namely non-dividing phase and dividing phase. During non-dividing phase, it is known as interphase nucleus. During dividing stage, it looses some of its parts and seizes to exist as a descrete cellular component. Parts of interphase nucleus: 1. Nuclear envelope 2. Chromatin 3. Nucleolus Nuclear envelope: Consists of two concentric membranes separated by a perinuclear space. The outer nuclear membrane may be studded with ribosomes and it is continuous with the membranes of ER. The nuclear envelope is interrupted by nuclear pores. Plugged into each pore is a hollow cylinder or ring known as annulus. The nuclear pore and its annulus together form a nuclear pore complex. These represent channels through which the nucleus communicates with the cytoplasm. Chromatin: It represents chromosomal material and is composed of DNA, basic proteins called histones, other non-histone proteins and varying amounts of RNA. Chromatin occurs in two forms: heterochromatin and euchromatin. Heterochromatin consists of tightly coiled portions of chromatin that lie entangled within the nucleus. Here genes are repressed and transcription does not take place. Heterochromatin is thus the predominant form of chromatin in relatively inactive cells (condensed, deeply stained and proportionately smaller nucleus). Uncoiled portions of chromatin is known as euchromatin in which gene transcription takes place and remain essentially unstained. Thus euchromatin is abundant in relatively active cells (large vesicular nucleus). Most mammalian nuclei are sexually dimorphic because one of the paired ‗X‘ chromosomes remains heterochromatic during interphase in female and this sex chromatin is named as Barr body (appears either as spherical structure underneath the nuclear envelope of all the cells or as a prominent nuclear appendage in neutrophils). Nucleolus: It is a dense well defined spherical body in the nucleus. There is no limiting membrane for the nucleolus. Nucleolus is absent in the neuroglial cells but distinct in  15 neurons. Structurally nucleolus is composed of RNA and associated protein of cytoplasmic origin (these form the pre-ribosomal RNP (ribonucleoprotien) particles). The final assembly of these particles into ribosomes occurs in the cytoplasm. Thus nucleolus plays an important role in the formation of ribosomes. Nucleolus consists of 1) granular part ie. pars granulose, 2) fibrillar part ie. pars fibrosa and 3) nucleolar matrix. Surface modifications of cells Cells have different kinds of functional surfaces. A. Free surface :- Either exposed to air or bathes in fluids B. Lateral surface :- Contact with other cells C. Basal surface or Attached surface:- Rests on a basement membrane. A. Free surface (apical surface) modifications: The primary apical modifications are microvilli, cilia and flagella. 1. Microvilli:- are slender cylindrical cellular processes which are oriented perpendicular to the apical free or luminal surface. Prime function is to increase the surface area of absorption of the cell. They are better developed in epithelia. They are 1-1.5 µmlong and 0.1 µm in diameter. Functions:- 1. Microvilli are most abundant in cells whose principal function is absorption and they increase the surface area (eg. epithelial cells of small intestine). 2. Some of the glycoprotein of the thick glycocalyx of the intestinal microvilli has hydrolytic enzymes such as alkaline phosphatase, disaccharide lactase and sucrase which further enhance absorption. Microvilli are observable with LM as a distinct striated border of the cells (eg. epithelial cells of small intestine, active cells of the PCT of kidney). With EM they are seen to be slender cylindrical process. They contain core of actin filaments which interact with myosin filaments causing the shortening of the microvilli. 2. Cilia and flagella :- Ciliated cells are commonly found in the respiratory system for the movement of mucus film and in the male and female reproductive system (propulsion of spermatozoa and oocytes). Several hundreds of cilia may be found in the single cell. They are 5- 15 µm long and 0.2 µm in diameter. A cilium is a slender thread of cytoplasm enclosed by the plasma membrane. Thicker and longer than microvilli. Anchored in the cytoplasm. Stereocilia: Non-motile (epididymis, vas deferens). Stereocilia do not posses the complex organizational  16 pattern. Hence they may be considered as extremely long and branched microvilli). Kinocilia: motile cilia. Flagellum: eg. Mammalian spermatozoon flagellum. It makes undulatory movement for the populsion of the sperm. Flagellated cells also occur in kidney, adenohypophysis, neurohypophysis, various locations in CNS, Pancreatic islets etc. B. Lateral surface modifications Lateral surfaces are modified to hold cells together, serve as diffusion barriers, permits diffusion between the component epithelial cells and permits intercellular communication. 1. Intercellular cement: Intercellular space of some cells contains mucopolysacharide. The main function is to hold the adjacent cells. 2. Junctional complexes or intercellular junctions which include a. Tight junction (Zonula occludens) b. Adhering junctions - there are three types which occur in the cell boundaries near the free surface d. Gap junctions (Nexus, Close junction, Communicating junction) These complexes may be seen together or independently. 1. Tight junction (Zonula occludens) Zonula = band, belt or girdle It is the most commonly encountered component of junctional complexes and it is the most intimate attachment between adjacent cells. Found primarily between columnar epithelial cells. It serves as a barrier preventing the passage of substances from the lumen to the intercellular space. The outer laminae of the adjacent cell membranes are fused to form network of multiple interconnecting ridge). These are separated by small islets in which apposing cell membranes are separated by an intercellular space of 2nm width. 2. Adhering junctions (Desmosomes) Desmosomes are numerous and distributed at regular intervals over the lateral surface of the epithelial cells. There are three types of adhering junctions: a. Belt desmosomes or Intermediate junctions or Zonula adherens It surrounds epithelial cells in a belt-like fashion. The two plasma membranes are separated by a space of 15-20nm width which is filled with granular material (a  17 polysaccharide adhesive) of low electron density. On their cytoplasmic surface, the membranes are coated with electron dense material whose filaments converge. b. Spot desmosome or macula adherens: They are discontinuous button like structure consisting of two dense plaques on the opposing cell surface separated by an intercellular space of 250 0A width. The intercellular space is filled with a fine granular material that forms an intermediate dense line in the center. Internally each contains an electron dense layer of amorphous material and bundles of the fine cytoplasmic filaments that converge and terminate on this layer. This arrangement effectively links the cytoskeleton of adjacent cells, especially the epithelia, and enable them to resist mechanical stress and maintain their integrity. c. Hemi-desmosomes: Consist of only one half of a desmosome and they connect epithelial cells to their basal lamina (it is an example for basal surface modification of cells). 3. Gap junction (Nexus) Here the intercellular space is narrowed to 20 nm and the space is traversed by tubular channels composed of integral membrane proteins of the apposed cell membrane. They make a direct communication between the adjacent cells permitting passage of ions and substances of low molecular weight. Cell web Little structures that extent across the free end of the cell. They are concerned with the binding the cells together and are connected with intra-cytoplasmic fibrillar material. EM shows it as a fibrillar cytoplasm extending across the free end of the cells under microvilli. Functions :- 1. Delicate fibrils give internal support. 2. Serve as anchorage for microvilli and for rootlets of cilia. Terminal Bars EM shows that it is only a condensed web material associated with Zona adherens. It is a ring like band encircling the top of simple cuboidal and columnar cells. It lies beneath the free surface. Under LM they appear as bars, dots or polygonal rings. Interlocking membranes Epithelial cells have their facing surface thrown into ridges and grooves. They may interlock and fit together. eg.Between the cells of PCT (indistinct). C. Basal Surface modifications: eg. Hemidesmosomes.  18 GENERAL HISTOLOGY BASIC / FUNDAMENTAL / ELEMENTARY TISSUES OF THE BODY 1. Epithelium 2. Connective tissue 3. Muscular tissue 4. Nervous tissue 1. EPITHELIUM The main functions of epithelium are protection, secretion, absorption, excretion and formation of barriers. Epithelia shed cells continually, such desquamated cells examined in smears of appropriate fluid (sputum, uterine, gastric etc.) for signs of malignant change ie the technique of exfoliative cytology. Classification of epithelia 1. Based on the number of layers: a. Simple epithelium – single layer of cells. It is adapted to absorptive and secretory roles. b. Stratified epithelium or compound epithelium– more than one layer of cells. Protect against mechanical and chemical action. c. Pseudostratified epithelium – in this type actually there is only a single layer of cells resting on a basement membrane. But the cells are of different sizes and therefore the nuclei are located at different levels. So it gives the appearance of stratification and hence termed as pseudostratified epithelium. 2. Based on the shape of cells: a. Squamous or pavement epithelium – cells will be thin, flat and scale–like. On surface view, the cells have an irregular shape with a slightly serrated border. Nucleus is centrally located and spherical in shape, causing a bulging of cytoplasm. b. Cuboidal epithelium – here the cells will have almost equal width and height. They appear as squares in cross section but more hexagonal when seen from surface. c. Columnar epithelium – consists of tall, narrow cells with greater height than width. Usually the nuclei are oval and are located near the base of the cell. Types of Simple Epithelium 1. Simple squamous epithelium – there will be a single layer of squamous cells. The cells rest on connective tissue. Distribution: 1) This epithelium lines the serous membranes like peritoneal, pleural and pericardial cavities. Here it is called mesothelium. 2) It lines the heart, all blood  19 vessels and lymph vessels and here it is called endothelium. 3) Lines membraneous labyrinth of internal ear, portions of kidney tubules and the alveoli and respiratory bronchioles of lungs. 2. Simple cuboidal epithelium: consists of single layer of cuboidal cells. It lines mainly the small ducts of exocrine glands; covers choroid plexus of brain ventricles, the ciliary body and anterior epithelium of lens of the eye. 3. Simple columnar epithelium: single layer of columnar cells. Generally simple columnar epithelium lines organs that perform absorptive function and secretion. eg. – Intestine, glandular stomach, uterus, gall bladder, vesicular gland etc. 4. Pseudostratified columnar epithelium: in this type all cells rest on basement membrane but all do not reach the surface. Those cells that reach the surface are either ciliated or non-ciliated or goblet cells (unicellular mucous glands). Basal cells are attached to basal lamina but do not reach the surface epithelium. eg. in respiratory tract and some regions of male reproductive tract. The mucus and cilia comprise an important protective mechanism called mucociliary apparatus. Types of stratified epithelium 1. Stratified squamous epithelium: consists of several layers of cells, the most superficial being squamous in shape. Lower layers can be of different type. There are two types of stratified squamous epithelium – 1) keratinized form which has cells on the surface layer that have lost their nuclei and filled with keratin. 2) non – keratinized form where the superficial cells retain their nuclei. 3 to 5 distinct cell layers are present in stratified squamous epithelium. They are  20 1) Stratum basale or stratum cylindricum or stratum germinativum: the deepest layer of cuboidal or columnar cells. It is named as stratum germinativum because it is the chief region of mitosis (stem cells). 2) Stratum spinosum: next layer; (number of layers vary). Polyhedral cells adhered by desmosomes. Spiny processes of tonofibrils radiate from the surface of cells. These processes (spines / prickles) are actually cellular processes joined by desmosomes. This appearance gives rise to the name stratum spinosum or spiny layer or prickle cell layer. Shrinking of cells by processing techniques demonstrate cellular spaces with numerous processes from adjacent cells. Stratum basale and stratum spinosum together known as Stratum Malpighi or Malpighian layer. 3) Stratum granulosum: this is a layer of granulated cells, which contain keratohyaline granules. These are precursors of keratin i.e. prekeratin. This layer is present only in keratinized form. Also contain lamellated bodies/ Odland bodies/ membrane- coating granules which release their contents into intercellular space to form the intercellular lipid component of the stratum corneum barrier. 4) Stratum lucidum: (lucidum = clear); occurs only in non–hairy skin regions. This layer of flattened keratinized, dead or dying cells with indistinct nucleus and agranular cytoplasm has a translucent appearance because it contains ‗eleidin’, a protein similar to keratin but with a somewhat different staining affinity. Stratum lucidum is only found in specific areas of exceptionally thick skin and in hairless regions (eg. Snout region of pig, plantar and palmar surfaces, planum nasale etc.) 5) Stratum corneum: present in keratinized form. Flat dead keratinized squamous cells without nucleus. Cytoplasm is filled with keratin. Keratin is a water resistant protein that forms a protective barrier. The most superficial layers of stratum corneum are flat horny plates, which are desquamated constantly (sometimes called stratum disjunctum). Keratinised cells are surrounded by a plasma membrane and a thick submembranous layer that contains a protein, involucrin. This is synthesized in stratum spinosum and cross-linked in stratum granulosum by an enzyme that makes it highly stable. Thus involucrin provides structural support to the cell, thereby allowing the cell to resist invasion by micro-organisms. Occurrence: a) Keratinized:- 1) covers the entire surface of the body ie. the skin, 2) it lines the mucous membrane of digestive tract from mouth to the secretory portion of stomach ie. ruminant fore-stomach (keratinized) and the anus. b) Non-keratinized:  21 Anterior epithelium of cornea, bulbar conjunctiva, mouth, pharynx and oesophagus, vaginal vestibule and glans penis. 2) Stratified cuboidal epithelium: in this type mostly there will be 2 layers of cells with the superficial layer being cuboidal in shape. eg. Excretory ducts of some glands like sweat glands and cells lining antrum of ovarian follicle (membrana granulosa). 3) Stratified columnar epithelium: there will be 2 or more layers of cells, the superficial being columnar in shape. Seen in the distal portions of the urethra and excretory ducts of salivary glands, in lacrimal sac and duct and palpebral conjunctiva of horse and dog. 4) Transitional epithelium: (plastic epithelium; plastic=changing) this type is multi- layered or stratified. The shape of cells change depending on the pressure exerted and the cells are capable of gliding over one another. This type is restricted to urinary system. ie. urinary bladder, uretors and initial parts of urethra. So it is sometimes referred to as urothelium. The shape of cells depends on the degree of organ distention at the time of fixation. When the bladder is full, cells become flattened and when it is empty, the cells become cylindrical. In the relaxed state; it may consist of as many as 6 or 7 layers. Basal cells are polyhedral. Cells of intermediate region are polyhedral or pear–shaped. Luminal or surface cells are cuboidal (umbrella-like or dome- shaped or pillow-shaped). At the time of distention as few as 2 or 3 layers may be apparent. Function: 1. Urothelium functions as a lining that is capable of stretching so that luminal volume is increased. 2. It is an effective barrier to the movement of water. It prevents movement of water from the connective tissue space to the lumen that is occupied by hypertonic urine. Softness and plasticity of epithelia are best developed in transitional epithelium. The cells are held together by viscous cement, which permits gliding over of cells on one another. Basement Membrane / Membrana Propria: this is a sheet of variable thickness interposed between epithelium and subjacent connective tissue. It consists of a ground substance of connective tissue, condensed gel-like and continuous with the ground substance of the underlying connective tissue. Embedded in this condensed ground substance are delicate reticular fibres, which are continuous with those of underlying connective tissue. Basement membrane in ordinary preparations is distinguishable in some location (eg. trachea). In other locations (beneath stratified squamous epithelium and columnar epithelium of intestine), it is not easily seen. Basement membrane serves  22 for attachment of epithelium to the underlying connective tissue. Fibroblasts may or may not be present. Mucous Membrane / Mucosa: this lines all the canals and cavities of the body, which connect with the exterior, ie. they line alimentary tract, the respiratory passages and the genitourinary tract. The essential parts of the mucosa membrane are surface epithelium, basement membrane and a stratum of connective tissue called the lamina propria. 2. CONNECTIVE TISSUE (Supportive tissue) The primary functions are to connect other tissues, to provide a framework and to support the entire body. Unlike epithelium, all types of connective tissues are mesodermal in origin. The connective tissue is composed of cells, fibres and amorphous ground substance or matrix. Classification of connective tissue 1) Embryonic connective tissue a) Mesenchyme b) Gelatinous or mucous connective tissue 2) Adult connective tissue a) Loose connective tissue b) Dense connective tissue 1. Irregular connective tissue 2. Regular connective (collagen and elastic) c) Reticular connective tissue d) Adipose tissue - white & brown e) Cartilage 1. Hyaline cartilage 2. Elastic cartilage 3. Fibrocartilage f) Bone 1. Embryonic connective tissues a. Mesenchyme: It is composed of irregularly shaped mesenchymal cells, which posses numerous cytoplasmic processes. The processes may be in contact with adjacent cells and thus form a three dimensional network. They undergo numerous mitotic cell  23 divisions and continuously change their shape. The intercellular space is filled with amorphous ground substance. During early development, mesenchyme does not contain fibres. During later stages fibres start appearing. Mesenchyme is the precursor for other tissues, gives rise to various types of adult connective tissues like cartilage, bone, muscle, blood and blood vessels. b. Gelatinous or mucous connective tissue: It is found primarily in umbilical cord and embryonic hypodermis (subepidermal region). In the umbilical cord it is termed, as ―Wharton’s gelly‖. Gelatinous connective tissue is characterized by star shaped cells with numerous cytoplasmic processes, which form a meshwork and these meshes are occupied by viscous gel-like amorphous ground substance and delicate collagen fibrils. In the adult animal, gelatinous connective tissue occurs in the core of (in the lamina propria of) omasal laminae, reticular folds and in the bovine glans penis and in the comb and wattles of gallinaceous birds. 2. Adult connective tissue Consists of fibres, cells and ground substance. Connective tissue fibres 3 types of fibres: Collagen, reticular and elastic fibres. a. Collagen fibres: Collagen fibres are most numerous of the fibres encountered in connective tissues. They are the secretary products of fibroblasts. Fresh collagen fibres are white, and in histological preparations, they stain with acid dyes. (Red / pink in H&E, red in van – Gieson‘s, blue in Mallory‘s and Masson‘s triple stain). Structurally collagen is a protein (which on boiling yields gelatin). Collagen fibres are seen extracellularly. Intracellular collagen fibrils have not been seen, except in certain tumors of mesenchymal tissue. Collagen fibres do not branch and are made up of parallel fibres of 10- 300 nm in diameter and of indefinite length. Each fibre is made up of a large number of microfibrils (made up of tropocollagen molecules), which have characteristic cross striations at regular intervals (at 67 nm intervals). The repeating cross-striational pattern is likely  24 formed by rows of collagen molecules arranged in an end-to-end fashion with each molecule overlapping the adjacent molecule by approximately one quarter of its length. So, under LM, collagen fibres appear as wavy fibres. They are characterized by high tensile strength and poorly stretched (only 5% of initial length). Consequently, they are found wherever high tensile strength is required such as in tendons, ligaments and organ capsules. The basic / fundamental unit of collagen is tropocollagen molecule. 5 types of collagen are there viz. type I, II, III, IV and V. Type I is the most widely st distributed type. (1 formed collagen is type III). Collagen molecules can be broken down by the enzyme collagenase. Vitamin C (scurvy factor) is, in man and guinea pig, a dietary factor essential for collagen formation. b. Elastic fibres (Yellow elastic fibres) They usually occur as individual, branching and anastamosing fibres and the ends are curved. (Spiralling and kinking of broken ends caused by elastic recoil in spread preparations). The fibres are thicker than collagen (0.5-12 µ in diameter). Structurally they are formed of the protein elastin (does not yield gelatin on boiling), they can be stretched 2-2 ½ times the original length. They are found in regions, which require great elasticity such as ligamentum nuchae, external ear, vocal cords, trachea, lungs and major blood vessels. The elastic fibres are lightly stained by ordinary methods but deeply stained by orcein and resorcine-fuchsin (Verhoeff‘s elastin stain). Elastase, a pancreatic enzyme will digest elastic fibres. Differences between collagen and elastic fibres Collagen fibres Elastic fibres 1. White 1. Yellow 2. Lesser diameter (10-300nm) 2. Thicker than collagen (0.5-12 µ ie. 500- 1200 nm) 3. Occur as parallel fibres 3. Usually occur as individual branching and anastamosing fibres with curved broken ends  25 4. Cross striations at regular intervals 4. No cross striations (64nm interval) 5. Formed of collagen, which on boiling 5. Formed of elastin that does not yield yields gelatin gelatin 6. Poorly stretched 6. Can be stretched 2- 2 ½ times 7. Found in regions where high tensile 7. Found in regions, which require great strength is required elasticity 8. Digested by collagenase enzyme 8. Digested by elastase c. Reticular fibres (argyrophilic fibres) They can‘t be demonstrated by routine staining techniques. These fibres can be seen only with silver impregnation technique, (thus the term argyrophilic or argentaffin fibres). These fibres are actually individual collagen fibrils (type III) coated by proteoglycans (contain 90% carbohydrate and 10% protein) and glycoproteins (contain 40% carbohydrate and 60% protein), which posses the affinity for silver salts. Reticular fibres are actually collagen fibres that have undergone minimal polymerization. This reduced polymerization with its associated availability of reactive groups may account for the argyrophilia. (When individual fibres are bundled to form collagen fibres, the coating is supposedly displaced and the argyrophilia disappears). Reticular fibres are immature form of collagen. They form delicate network around capillaries, muscle fibres, nerves, adipose cells, hepatocytes etc. and also take part in formation of capsules and framework of various organs especially lymphoid organs. Ground substance The fibres and cells of connective tissue are embedded in amorphous ground substance. This is predominantly composed of mucopolysaccharides, which are currently referred to as glucosammoglycans and proteoglycans (include hyaluronic acid and chondroitin sulfate). They show affinity for basic stains.  26 a. Loose connective tissue (Areolar C. T.) This is the most widely distributed type of connective tissue in the adult animal. It is present around blood vessels and nerves between muscle bundles and layers of smooth muscles. It makes up the interstitial tissue of most of the organs. The functions range from purely mechanical support to more sophisticated functions such as participation in tissue repair and inflammation. In loose connective tissue, cells are relatively more than fibres. Cells of loose connective tissue: 1) Fixed cells: 2) Wandering / free cells: a) fibroblasts a) macrophages b) pericytes b) mast cells c) adipose cells c) plasma cells d)chromatophores 1) Fixed cells a) Fibroblast: – This is the most common type of connective tissue cells. They are large stellate cells with extensive cytoplasmic processes. Cell membrane is delicate and usually not seen. Nucleus is spherical or oval, large, pale and may contain 1 or 2 nucleoli. Cytoplasm is light staining, homogeneous and basophilic. Fibroblasts are present in all connective tissues except reticular connective tissue. They are responsible for the formation of fibres (that is why the name fibroblast). Fibrocyte is the name given to inactive fibroblast / adult fibroblast and are inactive metabolically. They are elongated or spindle- shaped with a condensed cylindrical nucleus with dense chromatin and very little acidophilic cytoplasm, and is not motile / remains fixed. Fibrosis is the formation of fibrous connective tissue after injury. b) Pericytes / mesenchymal cells or cells of Rouget:- These are elongated pericapillary cells. Cells have fusiform nucleus surrounding which there will be a thin rim of cytoplasm. A number of slender cytoplasmic processes are present. They are pleuripotential, undifferentiated cells, which serve as reservoir cells / stem cells that can differentiate into other types of connective tissue cells as needed and is the precursor of most connective tissue cells. c) Adipose cells / fat cells / adipocytes: - These cells are spherical or oval in shape, most of the cell is occupied by a single large lipid droplet. Cytoplasm is reduced to a thin  27 peripheral layer. Nucleus is flattened and located at one end of the cell (eccentric). 2) Wandering / Free cells a) Macrophages / histiocytes / clasmatocytes: - They develop from the monocytes of blood. The monocytes migrate into the loose connective tissue and transform to macrophages. (Mesenchymal cells and reticular cells may give rise to macrophages. Blood monocyte and macrophage are considered identical cells. Within the blood vascular system (intravascular) this is referred to as monocyte. Once it enters the connective tissue compartment (extravascular), it is referred to as macrophage). In routine histological sections, fixed macrophages also called as histiocytes. In LM, it is difficult to distinguish it from fibroblasts. The cytoplasm is coarsely granular and shows vacuoles (studied by vital staining). These cells are distributed in fascia, stroma of various organs, sinusoids of liver, lymphoid organs and bone marrow. These cells are highly phagocytic and show amoeboid movement and therefore they are concerned with disposal of bacteria and other foreign tissue and cellular debris. They comprise the RE system. The nucleus of macrophage is the key to identify this cell. It is bean– shaped / kidney–shaped. It is smaller than fibroblast nucleus and darker. They play an important role in removing damaged RBCs. Macrophages may fuse to form foreign–body giant cells with many nuclei when faced with a large object for digestion. Milky spots seen on mucous membranes are dense accumulations of macrophages. b) Mast cells: - Sometimes referred to as tissue basophils. (German word ―masten‖ = fattened). They are common in loose connective tissue especially along the course of blood vessels. They are large oval or spherical cells containing numerous large metachromatic (basophilic) granules (when stained with metachromatic dyes like toluidine blue, thionine etc. they appear as pink / magenta granules). Under EM, the granules are membrane-bounded and have crystalline, lamellar or fine granular characteristics. These granules contain histamine and heparin (in mouse and rat, serotonin also) and take part in allergic and anaphylactic reactions. c) Plasma cells (10 µ in diameter): - These are spherical or oval cells with eccentric nucleus. The chromatin is radially arranged in a regular manner, which gives a ‘cart– wheel’ (clock–face or spoked–wheel) appearance to the nucleus. Due to the presence of numerous ribosomes, the cytoplasm will give intense basophilic reaction. Plasma cells develop from B- lymphocytes and are active in humoral antibody production. Plasma cells contain spherical acidophilic inclusions known as Russel bodies, which  28 are located within the dilated portions of endoplasmic reticulum. These bodies give a postive reaction for immunoglobulin (secretory or degenerative significance). d) Chromatophores (melanophores):- These are large cells with a number of branching cytoplasmic processes. They store pigments in the cytoplasm. They occur in uterine caruncles of sheep, dermis, choroid, iris and meninges. Pigment is mainly melanin. Primary function of these cells is to protect the body from excessive exposure to UV radiation. Melanin is synthesized by melanocytes. Melanocytes are able to invade epithelial layers and pass their pigments to other cells. These passive recipients of this pigment are termed as melanophores / chromatophores. b. Dense connective tissue In this fibres are more abundant than cells. Depending on the arrangement of fibres, it is divided into dense irregular and dense regular connective tissue. 1. Dense irregular connective tissue: - Bundles of collagen fibres cross each other at varying angles. This arrangement helps to withstand stretching forces in any direction. Seen in capsule of various organs like liver, spleen, kidney, testis etc. and the fascia, aponeurosis, pericardium etc. 2. Dense regular connective tissue: - This occurs in two varieties. (1) collagen tendons and ligaments and as (2) Elastic ligaments. In both, the fibres are arranged in the same plane and direction, according to specific functional requirements. a) Collagen tendons and ligaments: They consist of bundles of collagen fibers arranged parallelly and bound together by a thin layer of loose connective tissue known as peritendineum. Between these bundles, fibrocytes are seen. Tendon cells are fibrocytes, which are elongated seen in between bundles of collagen fibres and are arranged in rows. Aponeurosis: They have the same structure as tendons but are broad. Fibres run in superimposed layers, those of one layer at an angle to adjacent layers. The layers may interweave. b) Elastic ligaments: Consist of branching and interconnecting elastic fibers surrounded by loose connective tissue. eg. ligamentum nuchae and ligamentum flava.  29 c. Reticular connective tissue Composed of stellate reticular cells and a complex three- dimensional network of reticular fibers. Distributed in the stroma of all lymphatic organs (spleen, tonsil, LN etc), haemopoitic organs and bone marrow. d. Adipose tissue It is a specialized type of connective tissue. Two types of adipose tissue, white fat and brown fat distinguished by differences in colour, vascularity, structure and function. Large deposits of fat found in subcutaneous connective tissue is known as panniculus adiposus. Adipose tissue is also abundant in kidney region, in mesenteries and mediastinum and in cervical, axillary and inguinal regions. In white fat, cells are large with a diameter of 130 µm and are spherical or oval in shape and the cytoplasm contains a large single fat droplet. Nucleus is eccentric. Therefore white adipocytes are known as unilocular/univesicular adipocyte. Surrounding each fat cell, there is a delicate reticular connective tissue framework. This accommodates capillaries and fine nerve fibers. Brown fat, differ from white fat in that (1) the cells are smaller (2) cytoplasm contains a large number of small lipid droplets – multilocular/multivesicular adipocyte. In routine preparations, the spaces resulting from loss of lipids give a foamy appearance. (3) Nucleus is centrally placed. (4) They have a better capillary network and nerve supply than white fat. (5) More mitochondria than white fat cells. High concentration of cytochrome oxidase system in the mitochondria is primarily responsible for brown colour. Brown fat is particularly abundant in rodents and hibernating mammals. In others it has a limited but specific distribution. It develops during prenatal period and is confined generally to axilla, interscapular region, mediastinum, mesenteries and perirenal area. It helps the young animal in resisting postnatal cold extremes. Function of adipose tissue – (1) participation in fat metabolism, (2) synthesis of intracellular lipids, (3) serves as thermal and mechanical insulator, (4) In brown fat, oxidation of stored fat generates heat (rather than ATP) causing a rise in body temperature in arousing hibernating animals. RETICULOENDOTHELIAL SYSTEM (RE system) Coined by ASCHOFF in 1924. This name is given to the system more on physiological and pathological considerations than on anatomical structure. The cells of this system do not form true endothelium. All highly phagocytic cells of the body, except leucocytes belong to this system. The only certain method of identification is by injection  30 of non-toxic inert particulate matter like the carbon particles or china ink into the living animal. Injection of trypan blue or lithium carmine into living animal is called vital or intra-vital staining. The macrophages engulf the particulate matter like carbon particles. After vital staining, the cells containing the dye are found in the loose connective tissue, in the reticular tissue or in the blood sinuses of certain organs. In the connective tissue, they correspond to the histiocytes. Kupffer cells of liver, dust cells of lungs, reticular cells of lymph nodes, spleen and bone marrow, cells lining the sinusoids of spleen and microglia of CNS are examples of this system. Since this system includes reticular and endothelial lining cells of certain organs, it is named as RE system or Mononuclear phagocytic system. e. Adult Supportive Tissues 1. Cartilage Cartilage is a specialized type of connective tissue consisting of cells (chrondrocytes), fibers and amorphous ground substance. Cartilage forms the temporary skeleton of embryo and provides a model from which most bones develop. In the adult body, it covers articular surfaces of bones and forms the sole skeletal support of larynx, trachea, bronchi etc. Cartilage does not posses any blood vessel of its own (avascular), but vessels supplying other tissues occasionally pass through it. Cartilage is covered by a connective tissue covering, the perichondrium. Exchange of substances between cartilage cells and blood vessels of perichondrium is mediated by the tissue fluid of the cartilage ie. the bound water of the matrix. According to the nature and visibility of fibrillar elements, cartilage is subdivided into three. 1. Hyaline cartilage 2. Elastic cartilage 3. Fibrocartilage 1. Hyaline cartilage (glass-like) (Hyalos, Gr. = glass). This appears as bluish white translucent mass when fresh. Found in tracheal rings, larynx, bronchi, ventral extremity of ribs, nose etc. It forms the entire appendicular skeleton in the embryo. Composed of cells and matrix. (a) Cells: Mesenchymal cells differentiate into the cartilage forming cells of the body known as chondroblasts. These chondroblasts envelop themselves with their own secretions and eventually become isolated in small spaces called lacunae (lacuna =  31 hole). At this point, the cells are referred to as chondrocytes. Cells conform to the shape of lacunae in fresh state. Fixation and dehydration result in the retraction of cells from wall of lacunae and in section the cells appear as stellate. Nucleus is centrally placed with one or two nucleoli. Cytoplasm is granular and contains fat, glycogen and sometimes pigment. Growth in hyaline cartilage occurs in two ways: 1) Appositional / perichondrial growth by recruitment of fresh cells (chondroblasts) from the inner cellular layer of perichondrium. Here cells are smaller. 2) Interstitial growth by mitotic division and deposition of more matrix around already established chondrocytes (occurs towards the central region of cartilage). In bone, only the appositional growth is possible. Isogenous groups or Cell nests: Maturation of the chondroblasts into chondrocytes is accompanied by a cellular hypertrophy, an enlargement of the lacunae and a change in lacunar shape to an ovoid / angular configuration. Chondrocytes also cluster as isogenous groups or cell nests in the central portion of cartilage. These represent the daughter cells of a single parent cell or chondrocyte that undergoes mitosis during the growth process. Further, these nests indicate that interstitial growth has been accomplished. Cartilage is avascular and no blood vessels serve the matrix. Therefore nutrients and waste materials must diffuse through the matrix. This diffusion may break down and various degenerations then occur. eg. calcification of the matrix. This is prompted, organized and made use of in the process of endochondral ossification. (Cartilage canals may convey blood vessels through the matrix to non–cartilaginous regions). (b) Matrix: Bluish–white, homogeneous and translucent. It contains fine collagen fibrils in amorphous ground substance containing chondroitin sulphate, hyaluronic acid etc. The matrix is intensely basophilic in H & E, reacts positively to PAS and to metachromatic dyes. Because the fibrils have same refractive index as that of amorphous ground substance, matrix appears homogeneous. Surrounding each lacuna is the territorial matrix. (This contains a functionally special region immediately adjacent to the chondrocyte ie. pericellular matrix. The chondrocytic plasmalemma and pericellular matrix function together in the polymerization of collagen and proteoglycans). The area outside this is the interterritorial matrix, which stains lighter than the former because of increased numbers of collagen. Hyaline and elastic cartilages are surrounded by perichondrium (analogous to capsule of organs). Perichondrium consists of outer fibrous portion and inner cellular portion.  32 2. Elastic cartilage It is seen in external ear, epiglottis, eustachian tube etc. This appears yellow and more opaque than hyaline cartilage. In addition to all the structural components of hyaline cartilage, this possesses a dense network of elastic fibers that are visible in H & E preparations. Diagnostic features: (1) presence of elastic fibers (2) isogenous cell groups may be more frequently observed. 3. Fibrocartilage Of the three, this occurs less frequently. It is found in intervertebral discs, menisci of stifle joint, pubic symphysis, glenoid and cotyloid ligament, round ligament of hip joint etc. Fibrocartilage is a combination of dense collagenous fibers and cartilage cells. Diagnostic features: 1) Predominance of coarse collagenous bundles in parallel and/or V- shaped orientation (Classical appearance of this tissue is the herring bone configuration in which the collagen bundles are oriented to each other in the form of a ―V‖). 2) Limited amount of matrix between bundles. 3) Frequently the cells lie in rows between which are dense wavy bundles of collagen. 4) Lacks a perichondrium and is not seen as discrete pieces, ie. it is a transitional tissue between bone and bone or tendon and bone. 2. Bone Bone is a connective tissue with cells and fibers embedded in a hard, unbending substance well suited for supportive and protective functions. Hardness and rigidity is due to the presence of large amount of calcium salts deposited in the matrix. Organic material constitutes 35% and mineral part 65%. So before sectioning the bone, it has to be rendered soft by removing calcium salts and this process is known as decalcification. Example for decalcifying agent is nitric acid. Organisation of bone tissue Grossly two types of bone may be recognized (1) spongy or cancellous or trabecular bone and (2) dense or compact bone. The head or epiphysis of long bone when cut longitudinally has a spongy appearance. The shaft when cut seems to be formed of thick plates of bone known as compact bone. Histologically they differ in the degree of porosity. Differences between bone and cartilage 1) Bone matrix contains mineral component – amorphous calcium phosphate and hydroxy apatite crystals.  33 2) The cartilage cells are embedded within a matrix without contact with each other, but the bone cells (osteocytes) are in contact with each other through cellular processes located in the canaliculi. 3) Whereas cartilage is avascular, bone is an extremely vascular tissue. 4) Mineralized matrix, unlike normal cartilaginous matrix, is minimally expanded, therefore, is unable to grow by the interstitial mechanism. But cartilage grows by appositional and interstitial growth. Structure of bone Bone is traversed by longitudinal canals called as Haversian canals or central canals which are connected with each other and with the free surface by Volkmann’s canals or perforating canals or transverse canals. These canals lodge blood and lymph vessels and nerves. In cross section, Haversian canals can be seen surrounded by circularly arranged (4-20) or concentric lamellae. Within these lamellae, lacunae are seen which accommodate osteocytes which are the principal cells of mature bone and this type of arrangement is known as Haversian system or osteon which is the fundamental structural unit of compact bone. Numerous delicate channels known as canaliculi extent at right angles from lacunae and join adjacent lacunae. The osteocyte resides in the lacunae and it possesses numerous cytoplasmic processes which extend into the canaliculi and may come in contact with adjacent cells. Canaliculi of one Haversian system are not in contact between adjacent Haversian system. They are separated. Each Haversian system is marked or limited by cement lines. Organic matrix of the bone is termed as osteiod which is secreted by osteoblasts. Osteoblasts are bone producing cells some of which are trapped in lacunae and then become osteocytes. As osteoblasts mature, the Golgi complex and ER reduce and lysosomes increase. Osteoclasts are another type of cells responsible for resorbing the organic matrix and minerals. They are commonly multinucleated and larger than osteoblasts. Resorption tunnels through the bone or eaten out hollows or lacunae are known as Howship’s lacunae. Osteoclasts help in remodeling osteons. Three types of lamellae are identified in bones. Those surrounding the Haversian canals are the concentric lamellae. Between the osteons are many irregular lamellae known as interstitial lamellae. Third type of lamellae is the circumferential lamellae. Cocentric lamellae encircling the medullary cavity are known as inner cicumferential lamellae. Beneath the periosteum is the outer circumferential lamellae  34 that run parallel to the surface. Adjacent lamellar systems are sharply delineated from each other by a dark staining thin layer of modified matrix, the cement line. Most bones are invested by periosteum except at the articular surfaces. It has two layers; the outer fibrous layer made of collagen fibres and inner vascular and cellular layer known as osteogenic layer. The periosteum is attached firmly to the bone by bundles of coarse collagen fibres that have been incorporated into the lamellae of the bone. These fibres are called perforating fibres or Sharpey’s fibres. The marrow cavity and Haversian canals are lined with a delicate layer of squamous cells known as endosteum. Bone Marrow Bone marrow is a soft tissue which occupies the medullary cavity and larger Haversian canals and also between spaces between trabeculae of spongy bones. Structurally it has a delicate reticular connective tissue in which there are many types of cells. There are two types of bone marrow. Red marrow and yellow marrow. Red marrow predominates in foetus and young animals. During foetal and growing stages, the red marrow is osteogenic. As age advances, fat will be deposited in the bone marrow and becomes yellow marrow. In adult animals, red marrow is limited to sternum  35 and hip bones, vertebrae, ribs, cranial bones and epiphysis of long bones. Bone marrow is the chief blood forming organ of the adult body being the normal source of RBCs and granular leucocytes. In foetus haemopoisis occurs in liver, spleen and bone marrow. Red Marrow It is composed of stroma of reticular cells and fibres with free cells located in the mesh work. Majority of free cells of bone marrow are the immature developmental stages of the granulocytes and erythrocytes. Granulocytes: The stages of granulocyte development in the order of differentiation are haemocytoblasts, myelocytes and granular leucocytes. 1. Haemocytoblasts (myeloblasts): They are large upto 15 µ. The nucleus is large and rounded and the cytoplasm is deeply basophilic. They constitute normally 0.3-0.5% of the cells of marrow. 2. Myelocytes: These constitute 12% of the marrow cells. These cells show an increase in the granules, a decrease in the density of chromatin of the nucleus. According to the variety of granulocytes they are going to differentiate, the myelocytes show neutrophilic, eosinophilic or basophilic granules. At a very early stage in the transformation of haemocytoblasts into myelocytes, the cytoplasm is basophilic and shows less granules. The cells in this stage are called promyelocytes. When myelocytes transform into leucocytes, they show a transition stage in which the nucleus is kidney shaped. This stage is a metamyelocyte stage. Erythrocytes: They constitute 20-30% of marrow cells. The various stages in the differentiation of RBCs are: proerythroblasts, erythroblasts, normoblasts and erythrocytes. 1. Proerythroblasts or rubriblasts: (Basophilic erythroblasts)-they are smaller than myeloblasts; nuclear chromatin is coarser. 2. Erythroblasts: There is increase in haemoglobin in successive stages so that erythroblasts at successive stages show less of basophilia and more of acidophilia. Hence these cells are called polychromatophil erythroblasts. Cells are smaller than proerythroblasts. 3. Normoblasts: More haemoglobin. Cells slightly larger than RBCs. The cells extrude their nucleus to become RBCs. The youngest erythrocytes show a delicate reticulum and so it is called reticulocyte. The reticulum disappears before reticulocytes leave the marrow.  36 Megakaryocytes: Some haemocytoblasts become large megakaryoblasts (30-100 µ) and show lobulated nucleus. These are giant cells or megakaryocytes which give rise to the platelets. Lymphocytes and monocytes: Develop from germinal centres of lymphoid organs by differentiation of haemocytoblasts. BLOOD AND LYMPH Blood is a connective tissue (connection between deep lying tissues and atmospheric oxygen) consisting of free cells and a fluid intercellular substance, or plasma. The structural elements of mammalian blood are not all true cells but formed elements. They include red blood corpuscles (erythrocytes), white blood corpuscles (leucocytes) and blood platelets. Erythrocytes: They are highly differentiated specialized for the function of transporting oxygen. In lower vertebrates, the erythrocyte is nucleated but in mammals, it loses its nucleus, Golgi complex, mitochondria and centrioles before entering blood stream. They are acidophilic biconcave discs and round in all mammals except camellidae family (camel and llama) where they are biconvex elliptical. (Birds, reptiles and fishes have elliptical nucleated erythrocytes). On surface view, the erythrocytes are circular in outline and the central depression appears as a lighter area depending on the focus. The average diameter in microns of the erythrocyte is as follows: Elephant 9.0; man 7.5; dog 7.3; rabbit 6.5; cow 6.1; cat 6.0; horse 5.7; sheep 5.0; goat 3.7; musk ox 2.5. (Largest RBC – Elephant, among domestic animals – dog). Number (in million per cubic m.m.) Goat – 17.3 Sheep – 11.5 ± 1.8 Cat – 7.2 ± 1 Horse – 7 ± 0.7 Dog – 6.1 ± 1 Ox – 6 ± 1.3 Swine – 5.6 ± 0.7 Man – 5.0 Guinea pig – 5.0 Rabbit – 4 to 6 Chicken 2.9 ± 0.54.  37 The erythrocyte is elastic and can change its shape. Another characteristic is the tendency of the corpuscles to adhere to each other along their concave surfaces thus forming rows or rouleaux like piled up coins. Leucocytes: (White Blood Corpuscles) are nucleated (true cells) and show amoeboid movement. They wander and thus form the wandering cells of connective tissue. They may be divided into non-granular and granular varieties. The cytoplasm of granular leucocytes shows numerous granules. Number of leucocytes in thousand per cubic millimetre Chicken: 19.8 ± 9.6 Cat: 17.2 ± 6.6 Swine: 14.7 ± 4.5 Goat: 12 Dog: 11.3 ± 3.3 Sheep: 9.2 ± 3.1 Horse: 9±1.6 Man: 9 0x: 8 ± 2 Non-granular leucocytes or agranulocytes: - These include the lymphocytes which are small, and a group of larger cells, monocytes which have more cytoplasm and a more indented nucleus. The granulocytes are comparatively undifferentiated and can reproduce by mitosis. Lymphocytes: The size varies from 6 to 10 microns, the majority being 7 to 8 microns. The nucleus is large and stains intensely. The basophilic cytoplasm forms a narrow rim around the nucleus. Monocytes or large mononuclear leucocytes are large cells (12 to 15µ) but they may occasionally be up to 20 microns. It is the largest leukocyte. The nucleus is ovoid, kidney or horse-shoe shaped and eccentrically placed. It stains less intensely than lymphocytes. Cytoplasm is basophilic and abundant. In connective tissues they are phagocytic and are called macrophages. Granulocytes: These are divided into three classes according to the nature of the granules present as neutrophils, eosinophils and basophils. They are further characterized by the presence of many lobed (polymorphism) nucleus, hence the name polymorphonuclear leucocytes. The lobes of the nucleus are connected by fine chromatin strands. The cells are highly differentiated and cannot reproduce by mitosis. Neutrophils: These are large cells, 8 to 12 microns and constitute 60 to 70 percent of the total leucocytes. The nucleus shows 3 to 5 lobes connected by fine strands and is in  38 the form as an ‗S‘ or horse-shoe. Granules are fine and neutrophilic, ie they have affinity towards the neutral stains of the Romanowsky compound. (In the rabbit, guinea pig and chicken, they take acid stain. In these species staining reaction is different. Hence these cells are also called heterophils). They are highly phagocytic and show great amoeboid movement. In man, dog, horse and pig, the neutrophils are in majority. In ruminants, chicken and laboratory rodents, lymphocytes are in majority. Eosinophils: These constitute 2 to 4 percent of the leucocytes. They are larger than neutrophils, about 10 to 14 microns in diameter and contain course highly refractile granules which stain with eosin and other acid dyes. The nucleus is polymorphous, generally with only two lobes. Basophils: These are rare being only 0.5 per cent of leucocytes except in the chicken which has 2 per cent. Their size is 8 to 10 microns. The nucleus is large and irregularly polymorphous. The coarse granules are basophilic and are soluble in water. They are therefore not usually seen in the ordinary preparations or appear blurred. Functions of the leucocytes: Lymphocytes – These can develop in the connective tissue, into plasma cells and monocytes and from the latter into macrophages. They are also concerned in the production of antibodies. Monocytes are highly phagocytic. Neutrophils – After migration from blood stream the neutrophils phagocytose bacteria and other small particles. They have been called the microphages, in contrast with the macrophages, which are larger cells that engulf larger particles. The neutrophils also elaborate powerful proteolytic enzymes which may act outside the cell body. Eosinophils: Their number increases in parasitic infections. Basophils. They are phagocytic in function. Blood Platelets: They are colourless, about 3 microns, biconvex discs but appear spindle shaped when seen in edge. They have no nucleus. They show a central granular mass which stains deeply with basic stains and a peripheral hyaline zone. Birds and other lower vertebrates have true thrombocytes which are nucleated and have basophilic cytoplasm. Lymph: Lymph, like blood consists of a fluid plasma in which are suspended various corpuscular elements. Red blood corpuscles and platelets are absent. The chief cellular elements are lymphocytes with few granules.  39 3. MUSCULAR TISSUE Three basic types: Smooth muscle, Skeletal muscle and Cardiac muscle. SMOOTH MUSCLE It is formed of spindle shaped cells, the centre being enlarged and extremities narrow. It is non-striated and involuntary in action. Nucleus is located in the centre and surrounded by large amount of cytoplasm or sarcoplasm. Plasma membrane is known as sarcolemma. Cells are arranged in such a way that narrow portion of one cell is in contact with wider portion of adjacent cell. Diameter ranges from 3-8 µ and the length from 15-200 µ and can adjust to variations in shape of the organ. In pregnant uterus, the length may reach upto 500 µ. Cytoplasm is homogenous and contains myofibrils. Occurrence: Seen on the wall of alimemtary canal from terminal part of oesophagus to anus, urinary bladder, ureter, urethra, uterus, vagina, blood vessels etc. and muscles like arrectorus pilorum, retractor penis and in the ciliary body and iris. SKELETAL MUSCLE Skeletal muscle fibre is formed by the fusion of many myoblasts during embryonic stage. So each cell is multinucleated and the nuclei are located at the periphery of the cell. Characteristic feature is the presence of cross striations due to the presence of parallelly arranged bundles of myofibrils, each myofibril containing approximately 1500 thick and 3000 thin myofilaments. Individual muscle fibre is surrounded by reticular connetcive tissue known as endomysium. Groups of muscle fibres form primary bundles known as fasciculi by collagenous covering known as perimysium. Several primary bundles join to form a muscle. So the entire muscle is surrounded by epimysium which is also made up of collagenous connective tissue. All these connective tissue networks are continuous with each other. This carries blood vessels and nerves.  40 Plasmalemma of skeletal muscle is the sarcolemma; the endoplasmic reticulum is the sarcoplasmic reticulum and the mitochondria are the sarcosomes. Cytoplasm contains myofibrils. Structure of Myofibril Each myofibril consists of alternating light and dark bands. Light bands are known as isotropic bands or ‘I’ bands. This is made up of actin filaments or thin filaments which has a diameter of 6nm. Actin is a globular protein. Actin filaments are in the form of double helix. In the thin filament, in addition to actin, are other proteins like actinin, troponin and tropomyosin. (skeletal muscle proteins include actin, myosin, actinin, troponin, tropomyosin and M-line protein). Centre of the I band shows ―Z‖ line on  41 which the actin filaments are attached. The other end is free and interdigitate with the myosin filaments. Dark or A band consists of myosin filaments which are parallely arranged. Each myosin filament or thick filament has a diameter of 15nm. Myosin filament is ‗golf club‘ shaped. It has a HMM (Heavy Meromyosin) portion or arm portion and a LMM (Light Meromyosin) portion. These two are joined by a hinge joint. Arm has two portions, the arm and the head. The arm is connected to the head by another hinge joint. Muscle contraction is by sliding filament theory, ie. the actin filaments slide over myosin filaments. The line through the centre of A band is ‘M’-line (M stands for ‗mittel‘ (German word) = middle, intermediate). Here, each myosin filament is connected to 6 adjacent myosin filaments by M-line filaments/protein so that in cross section, they appear hexagonal. M-line forms the central line of H band (Helles band, Helles is German word that means clear). Here only myosin filaments are there. Towards the periphery of the A- band, the actin and myosin interdigitate. The region of myofibril between two adjacent Z lines is known as sarcomere. It is the contractile unit of skeletal muscle. Length of sarcomere is 2.5µ in length in relaxed state. ie. A band = 1.5 µ. I band = 1.0 µ. During contraction, the length is reduced to 2.0 µ or less. The length of A band remains constant. The actin filament slides over the myosin so that width of I-band is reduced and so also the H-band becomes narrow since the actin filaments are moving towards the M-line. Other than the myofibrils, the sarcoplasm contains sarcoplasmic reticulum. These are arranged around the myofilaments in a longitudinal manner. They form dilated portions towards the terminal region known as terminal cisternae. There is a second set of sarcoplasmic reticulum known as transverse tubule system or T tubules. These are formed by the invagination of the sarcolemma towards the interior of the muscle fibre. The arrangement where the T tubule with two terminal cisternae one on either side is named as TRIAD. This is very important in excitation-contraction coupling mechanism. Here the sarcolemma and membrane of the terminal cisternae are in close contact with each other where they form gap junctions which help in calcium ion transport. Innervation Ratio: Ratio of number of nerve terminations innervating skeletal muscle fibres. In highly active muscles like extrinsic muscles of eyeball the ratio is 1:1 (one  42 nerve termination per a skeletal muscle fibre). In others like gastrocnemius, the ratio is 1:1000. CARDIAC MUSCLE It forms the musculature of heart. Cells are cylindrical and striated which are similar to skeletal muscle fibres but the cells branch and anastamose. The single nucleus is located in the centre of the cell. Myofibrils are not so distinct as in skeletal muscle. Intercalated discs are peculiar to cardiac muscle. They are cross bands 0.5-1 µ thick and are strongly refractive in fresh muscle, but deeply stained in fixed material. They often follow an irregular or broken course giving the appearance of ―step- formation‖. EM studies shows that these represent specialized cell junctions including desmosomes and gap junctions. 4. NERVOUS TISSUE This is highly specialized for the expression of the two chief basic properties of protoplasm, irritability and conductivity. The nerve fibre conducts the impulse with a speed up to 10m per second. Other than this function, some neurons are secretory ie. neurosecretory neurons (eg. Herring bodies in posterior pituitary, in pineal gland, subcommissural organ etc.). Nervous tissue consists of neurons and supportive cells and neuroglia. Neurons Neurons or nerve cells are the structural and functions units of nervous system. Mammalian nervous system contains approximately 100 billion neurons. Human cerebral cortex contains about 14 billion neurons. The neuron is composed of a cell body together with its processes, the dendrites and axon. The axon conducts impulses away from the cell body and is usually single. The dendrites conduct impulses towards the cell body. Neuron doctrine Neurons are the genetic, morphologic, functional and trophic units of the nervous system. This statement of neuron doctrine recognizes that (1) each neurons is derived from an embryonic stem cell, a neuroblast and contains all the necessary coded information to fulfil its functions (2) each neuron is a separate and dinstinct structural unit that makes contact with other units but have structural continuity (3) chains of these cells comprise the conduction mechanism for the nervous system (4) each neuron is responsible for nutrition, metabolism and maintenance of the component parts.  43 Classification of neurons I. Based on the number of processes extending from the cell body a. Unipolar neurons- Only one axon eg. Neuroblast. b. Pseudounipolar neurons: These are derived from the original bipolar cell. Seen in sensory dorsal root ganglia of cranial and spinal nerves. Physiologically the single pole is formed by fusion of the axon and the dendrite. Because of the single pole formation with the cell body on top and processes diverging at almost right angles, the pseudounipolar cell is frequently referred to as a ‗T‘ cell. c. Bipolar neurons: These are the most primitive and are found most commonly in the embryo. In the adult, they are located in sensory neuroepithelia of the eye, olfactory epithelium and within the ganglia of vestibulocochlear nerve. Pseudounipolar and bipolar neurons are sensory neurons. d. Multipolar neurons: These are the most common type of neuron found within the CNS of higher mammals. In the peripheral nervous system, this type of neuron is located in the autonomic ganglia. The most outstanding physiological advantage of a multipolar neuron is that many dendrites permit more axodendritic synapses. e. Anaxonic neurons: eg. Amacrine cells of retina. Seen in the inner plexiform layer of retina and connects bipolar neurons with ganglion cells. II. Based on the size of the cell body: Diameter may vary from 4µ (granule cells of cerebellum – the smallest neurons in the body) to 150 µ (cell bodies of the lower motor neurons within the ventral horn of the spinal cord). III. Based on the length of axon: a. Golgi type I neurons: Those with extremely long axons (feets / metres). eg. neurons contributing to the sciatic nerve (In man, >3ft long from spinal cord to foot; in giraffe, 15ft or more). b. Golgi type II neurons: Neurons having extremely short axons. Eg. Cells in the gray matter of the cerebral and cerebellar cortices, interneurons etc. IV. Based on fibre diameter or rate of impulse conduction: a. A fibres: Largest in diameter (10-20 µ): Most heavily myelinated fibres to skeletal muscle and sensory fibres from muscle-tendon receptors (speed: 100-120 m/sec) b. B fibres: 5-9 µ in diameter- lightly myelinated - eg. Preganglionic autonomic fibres (slower rate of conduction).  44 c. C fibres: 1.3-2 µ in diameter – non-myelinated- eg. Postganglionic autonomic fibres and small non-myelinated sensory neurons of peripheral nerves and dorsal roots (very slow rate of conduction). V. Based on function / With regard to the direction of nerve impulse travel: a. Afferent / sensory neurons: (eg. cell bodies in the dorsal root ganglia) b. Connector / internuncial neurons (cell bodies and the terminals of their processes are located in the gray matter of spinal cord or brainstem) c. Motor / efferent neurons: Cell bodies lie in the gray matter of neuraxis. VI. Based on the shape of cell body a. Stellate cells b. Pyramidal cells (with apical and basal dendrites) c. Purkinje cells (with flask-like body and dendritic tree at one end) Structure of a multipolar neuron 1. Nucleus: large, spherical, vesicular nucleus and central in position. In some cells they are eccentric. Nucleus is pale staining (more euchromatin, so very active), but the nuclear membrane is dark. Nucleolus is single, large and very prominent (Owl‘s eye or bull‘s eye appearance).  45 2. Neuroplasm a. Neurofibrils: These are fine cytoplasmic fibrils which form bundles in the cell body. Their arrangement is irregular in the cell body and parallel in the processes. b. Nissl bodies or tigroid bodies or chromidial substance or chromophil substance: (Named in honour of Franz Nissl who studied this material). These are rER and ribosomal aggregations in the neuroplasm which are characteristic of nerve cells. These are basophilic. They are seen in dendrites also, but not in axon and axon hillock. These are classified as small, medium and large Nissl bodies. It is quite labile and may dissolve when the cell body and / or its processes are injured (chromatolysis). Nissl bodies can be demonstrated by crystal violet. Other organelles are Golgi complex, mitochondria and inclusions like lipochrome and melanin. 3. Dendrites: They are extensions of cell body and thus increase the surface area. They are highly branched and tree-like. They contain neurofibrils, Nissl bodies and mitochondria. 4. Axon or axis cylinder: Axon is always single and arises from a special conical par