MicroanatomyLesson 1 Microscope & Tissue PreparationAnatomy the scince dealing with form and structure of living organisms.2kinds of anatomy: Gross Anatomy the study of the structure of the body and its parts without the use of the microscope; As seen by the naked eye. Microscopic Anatomy also known as Histology Department of anatomy dealing with the minute structure, composition & function of tissues You see in the microscopic level.Embryology - branch of biology that studies thee formation and early development of living organisms; Where it came from.Etymology:Histos Greek tissuesLogia study/ knowledgeHistology study of tissues both in the plant and animals.Organology study of organsHistology study of tissuesCytology study of cells.Why Study MicroAna? Help understand gross anatomy. Foundation of Pathology (to know which is abnormal) Structural basis for Physiology Helps understand that: Examining a structure will help us know its function. If we know the function of an organism/ tissue, we can deduce its microscopic features.

Important considerations:1. Kind of microscopea. Light microscopeb. Microscopes with built-in lights2. Preparation of the tissue suitable for viewing with the microscopea. Quality of the slideImportant persons:Robert Hooke Father of modern MicroscopyHans Lippershy - telescopeSacharias Jansen telescopeAnton Van Leeuwenhoek Father of MicrobiologyFirst microscope 1595 ciscaMicorscopy classification:1. Type of light sourcea. Visible Light Usual; used to view the general structure (overview)b. Beam of electrons Electron microscope; to view the more detailed structure in cells (organelles)c. UV light (Ultraviolet) - to view the more detailed structure in cells (organelles)2. Usefulness of the microscopea. Ability to magnifyb. Ability to resolve detail1500x useful magnification in Light microscopeResolving Power the capacity of miscroscope to clearly separate 2 close together. Limited by wavelength of light and Numerical aperture. Numerical aperture light gathering capacity of objective (lens) 0.2 micrometer ( ) light microscope Advanced technology = Resolving power = quality of imageMicroscope components: Mechanical Parts holds the optical, moves stages Stage - shelf to support the material examined The hole in the center of the stage serves to let the light from illuminator and condenser shine upon specimen. Stage Clips securely holds the specimen (slide) using 2 knobs below the right side of stage, we can move the specimen Left and Right &Forward and Backward. Base, Pillar, Arm holds essential optical parts Body tube The principal lenses are attached to. Revolving nosepiece holds the objectives. Coarse Adjustment knob moves the body tube up or down RAPIDLY; Used for scanning and LPO Fine Adjustment knob moves the body tube up or down SLOWLY; Used for HPO. Mirror collects/reflects light into the lenses. Iris Diaphgram controls light amount admitted by the condenser. Substage Condenser directs maximum light; well focused. Optical Parts objectives and lenses Objectives holds the lenses of the microscopeNosepiece mounts the objectives Scanning (Overview)4xRed LPOLow Power Objective 10xYellow HPOHigh Power Objectice40xBlue OIOOil Immersion Objective 100xWhite/Black Eye piece / Ocular magnification 10x Diaphragm regulates the amount of light reaching the specimen.Objectives & Focusing:1. Start with the lowest magnification2. OIO lens is for maximum magnification ex. Looking at bacteriaPower of Magnification:TOTAL Magnification = Magnifying power of ocular (eyepiece) x Magnifying power of objectivesMICROSCOPES TYPES:Light microscope: Simple - composed only of 1 lens Compound composed of 2 or more lenses

Visible Light Source Microscopes: Polarizing Microscope For studying crystalline forms Birefringence optical property of Polarizing; Double Refraction Uses 2 Prisms ( below condenser & above objectives) Rotation of polarizer elicits birefringence Also used to view Muscles, Fibers (Connective Tissues) & Fat droplets Examples: Uric acid crystals Calcium phosphates

Phase- contrast Microscope For studying Living cells Purely optical B&W/ Grayish Refractive Index speed with which object is traversed by light wave. Interference Microscope Ability to retard Light Dark Field Microscope Used for transparent Makes everything black; you only see the cell dust in beam of sunlight effect Ex: Chylomicrons SiphyllisBright-Field Microscope Non Visible Light Source Miscroscope: UV Microscope Better resolution 0.1 micrometer quarter lens Fluorescence Microscopy (brighter) Can use dye Electron Microscope Uses electron beams. Looking for organelles in details (ex. Inside mitochondria)

PREPARATION OF TISSUESSTEP

1FIXATIONFixativesFormalin for CadaversAlcohol (Etyl)

2DEHYDRATIONDehydrating AgentsAlcoholAcetoneremoving excess water; dried enough but not too dry

3CLEARINGClearingXyleneChloroformRemoves excess dehydrating agents; de-alcoholization

4IMPREGNATIONPut waxing materials

5EMBEDDINGEmbedding materialsParrafinCelloidinAllows to solidify

6TRIMMING

7SECTIONINGCutting the sections3 10 micrometerMicrotome instrument used to cutSections are mounted on slides; put in Water bath to prevent wrinkles.

8STAININGH&E (most common)HematoxylinEosinRemoval of Paraffin

9MOUNTINGMounting mediumBlasamCover glass to protect stained/ processed tissue

Cassettes carries tissue Duration:12 hours (usual time)5 hours Cryostat (shortcut method) uses 24C.

Stains:ACIDIC (Red/Pink)BASIC (Blue/Purple)

Eosin Stains the cytoplasm

Hematoxylin Blue Basophilic Dyes the Nucleus 9nucleic acid) Natural dye From Mexican trees heartwood Hematin active components from oxidation of Hematoxylin

Micro AnatomyLesson 2: Cells

The Cell functional unit of living organismsSingle cells: Bacteria a dn AlgaeMulticellular has extracellular matrix (matrox of the bone)Diffferentiation - cell assumes specialized structure and functionWell differentiated = better function = more matureEukaryotic CellProkaryotic Cell

Larger Nucleus Cytoplasm Well defined nuclear membrane Ex: animals Smaller Nucleus Cytoplasm No nuclear membrane or envelop NOT CLEARLY DISTINGUISHED Ex. Bacteria unicellular

Eukaryotic Cell: 2 basic components: Nucleus Cytoplasm Contains all organelles Nucleus largest Surrounded by cell membrane/plasma membrane/plasmalemmaNucleus headquarters Control center Distinct Nuclear Membrane Contains DNA (in chromosomes responsible for division) Geneyic Material Building proteins Cell reproductino Found in all Cells except RBC & platelets Has Nucleolus/Nucleoli controls RNAChromosomes 46 chromosomes/23 pairs Rod-shaped/ threadlike Structures made up of masses of heterochomatinXY male XX female Chromatin DNA & Proteins Histones - on dividing; becomes active in cell division Parts of ChromosomesTELOMERES - ENDEuchromatin orange (active)Telomeres regukates/stopes dividingTelomerase cuts telomeres; no more cell division

Cell Division = Cutting TelomereLonger Telomeres = more Cell DivisionPremature aging of cells shorter telomeresEuchromatin lose network of chromatin fibrilsHeterochromatin condensed network of chromatin that appears as course of granules

CYTOPLASM matrix in which are embedded several organelles plus deposits of carbohydrates. Lipids and proteins. Plasmamembrane (plasmalemma)- Separates it from extracuricullar environment A.k.a. cytosol (solution inside the cell) Not just H20 Cytoplasm Eosin Nucleus HematoxylinSTRUCTURES FOUND IN THE CYTOPLASM:Organelles All eukaryotics little organs (enzymes) Metabollically active Membrane-bound Permanent part of cytoplasmMitochondria All eukaryotic Powerhouse ATP Production Tend to accumulate in parts of the cytoplasm where metabolic activity is more intense Cristae MatrixRibosomes Types: Free and Attached Dense, dark colored granules Dense (dark colored) granules Big subunit 1 Large molecule of RNA 30 small proteins Larger subunits (adjacent to membrane) 2 molecules RNA 40 associated proteinsFree protein synthesis for cells; singly scattered in cytoplasm; Synthetically active cellsClusters of 10 or 20 Polyribosomes/PolysomesAttached protein synthesis for export; partner of Golgi BodyEndoplasmic ReticulumSmooth Endoplasmic Reticulum Site of: Steroid synthesis Lipid synthesis Carbohydrate synthesis Hormone synthesis Oxidation Conjugation of substances Detoxyfying Noxius substances poisonus substances Ex: Liver has more S.E.R for detoxifyingRough Endoplasmic Reticulum Segregate proteins designed for export or intracellular utilization or intracellular utilizationGolgi Apparatus Involved in secretory activity Stack of flattened membranous sacs Modify and package proteins from R.E.R. Secretory vescicles Carries proteins and phospholipids to become part of cell membraneVescicles pinches part of Golgi for export (secretory vescicles) trafficking substances in or out of the cells relation with Lysosomes: Vescicles attaches to Lysosomes digestion ex. proteinsLysosomes Primary: no digestive, metabolizing, inactive small vescicles; transfers into secondary when needed Secondary involved in enzymatic activities, vacuoles for digestion, activeResidual bodies ex: pigments Phagocytotic vacuole Phagolysosome (secondary, active)Peroxisomes or Microbodies Spherical Smaller than mitochondria Has enzymes Uses Oxygen to detoxify harmful substances; removes alcohol formaldehyde Disarms free radicals Free Radicals H2Os H2O Free Radicals has only one electron so it gets a partner from other pair of electron (snatched the partner). Antioxidants provides electrin to free radicalsCentrioles For mitotic division For spindle fibers Schort cylindrical structures composed of microtubules Center of activities associated with cell dividion Self duplicatingInclusions Leftovers Ex: Pigment Crystals Secretory granules Stored carbohydrates, proteins and fats (Globules) Appear as vacuoles, graules, globules Temporary componenets of cell Accumulation of metabolites or cells pigments = undigested substancesCell Membrane Plasmalemma Plasma membrane Outer boundary of all eukaryotic cells Composed of lipids Phospholipids Cholesterol Good Bad too much causes disease. Goes to cell membrane, not really bad Proteins (weak structure if less) Gives shape Regulates passage of substances in or out of the cell Provides means for cellular attachment Has antigenic molecules that plays a role in recognition and tissue specificity Has ion pumps to regulate intacellular environment Has receptor for hormones Can generate messenger molecules to facilitate the cells response to stimuli Cell to cell communication: Hormones attach to receptors Androgens/testosterones attach to partner receptors send signal to nucleus decides to be masculine (mustache, oily skin) Drugs/medications Lipid bilayer stabilizes Hydrophillic Hydrophobic H2O swelling Proteins Integral & Peripheral Cell coat or Glycocalyx EXTERNAL COVRING; CONTAINS Carbohydrates for absorption Ex. small intestine has lots of glycocalyx.SPECIALIZATION OF THE CELL MEMBRANE:A. Junctional complex- cell attachments4 types of cellular attachments:1. Macula adherens strongest; ex. skin2. Zonula adherens celltocell communication a.k.a. Intermediate junction, Fascia adherens3. Zonula Occludens Tight junctions4. Nexus/Gap Junction in neurons (communication)Macula Adherens/Desmosomes Prevent cells under mechanical stress from being pulled apart Ex: mouth, esophagus, vagina, skin Site of attachments of cytoskeleton to the cell surfaces and sites of cell to cell adhesion Anchoring junction (button-like thickness) PROTECTIONZonula Adherens Intermediate Junction Fascia adherens Collar-like ring of attachment which increase the Surface area of contents of cells Larger intercellular space within striations Intercalated disc of cardiac muscle HOLDS FIBERZonula Occludens Tight junctions The cell coat is absent and the cell membranes between two cells have fused. Forms a barrier that prevents free passage across the epithelium example: lining of the urinary bladder, GIT BARRIERNexus/Gap junction Regions of low electrical resistance for cell to cell propagation or excitation- contraction impulse conduction. Communicating junction found in epithelial muscular and nervous tissues Significant for cell-to-cell communication GAPS/COMMUNICATIONB. INVAGINATIONS1. Infoldings increases the cell surface2. Vesicular - droplike invaginationEndocytosis take ina. Pinocytosis fluid, smaller particlesb. Phagocytosis large particlesc. Receptor-mediated endicytosis hormones or drugsExocytosis take outC. MICROVILLI finger-like projection of absortive epithelial cells on freee surfacesas of cell membrane1. Striated border more in lining of GIT2. Bruch border fiund in PGT of kidneys

D. STEREOCILIA long process in the apical region of cell membranes, non mobile e.g. lining in the epididimis (cilia) in fallopian tube respiratory passagesCell Types1. Labile cells always dividing. Ex. Skin cells, digestive tract, respiratory tract, stem cells2. Stable Cell do not multiply continuously but can do so when necessary. Ex. GO, liver, proximal tubules of the kidney, endocrine3. Permanent Cells incapable of multiplication in the adult. Incapable of regeneration. Ex. RBCs, mervous tissues, cardiac myocyte, lens of the eye.CELL CYCLE G0 resting Interphase G1 S G2 M-phase P M A T CytokinesisInterphase in between DNA replication; no cell division; DNA are active but not dividingM- phase nuclear division mitosisInterphase gap phaseG1 DNA synthesizesG2 after resting, before DNA synthesisS DNA synthesisMeiosis -1 product (gametes), basal bodies and eggMitosis 2 daughter cellsParenchyma function Principal or characteristic of functional cells of the organ Form main bulk of the hunan body Ex: skeletal muscle cell skeletal muscle Liver cells hepatocytes liver Nerve cell neurons; spinal cord bone cells osteocytes bone Cartilage Chondrocytes Blood RBCs red blood cellsStroma structural support Other cells and fibers of an organ which forms the framework or support cells and tissues of organs Ex. stroma of spinal cord meninges and neuroglial cells Stroma of skeletal muscles connective tissue covering enndo, peri and epimysium Stroma of liver interlobular Connective tosue Glissons capsuleIntercellular Spaces Spaces between characteristic cells or parenchyma of an organ Contents:A. Tissue fluids Derived from plasma, found throughout the bodyB. Ground substances/.matrix Derived from charafteristic cells in specific organs Extracellular matrix solid, liquid, semiliquid, semi solidC. Cells other than parenchymaa. Pigment cellsb. Phagocytic cellsc. Plasma cellsd. Fat cellse. Blood cellsD. Intercellular fibersa. Collagenb. Elasticc. Retricular fibersE. Blood VesselsF. NervesG. Lymphatic vesselsKuffer cells macrophage in livers intercellular spaceWBC large, spherical, fewCLASSIFICATION OF CELLSA. SIZE RBC index (7-13 micrometer) Small smaller than RBC Medium almost same size with RBC Large bigger than RBCB. NUMBER OF CELLSAbsolute terms exact number of cells in area per unit volumeEx. WBC 5,000 per cubic mm. Few Moderate NumerousC. NAMING CELLSa) Fundamental Type they form Muscle cell musclular system Nervous cell nervous systemb) Organs where they are found Liver cell in liverc) Person discovered Horizontal cell of Cajal Dr. Cajald) Characteristic histologic feature Acidophil affinity to acid dyeD. SHAPE OF CELLSa) Flattened or Squamousex. endothelialb) Cuboidalex. thyroid follicular cellsc) Columnarex. epithelium lining the GITd) Stellateex. nerve celle) Pyramidalex. parietal cells of Stomachf) Spherical or globularex. leukocytes or WBCg) Plygonal/polyhedralex. liver cellsh) Disk Shapedex. goblet cellsi) Cylindrical/Elongated with Bluntex. Skeletal muscle cellj) Disc Shapeex. RBCk) Cylindrical/Elongated with Branchingex. cardiac musclesl) Fusiform/ Spindle Shape/ Elongated w/tapering endsex. smooth muscle cellm) Irregular ShapeE. ARRANGEMENT/ DISTRIBUTION OF CELLSa) Sheets or Layersex. epidermisb) Bundles or fascicles ex. skeletal c) Cords or rows or columns ex. liver cellsd) Concentric ex. osteocytes in compact bonee) Nodular ex. lymphocytes in l. folliclesf) Isogenous group ex. cartilage cellsg) Irregularly or haphazardly arranged ex. nerve cells or spinal cord

TORIO, CAMILLE ANGELIN T.