CHAPTER 2 :BLOOD CIRCULATION and THE CARDIOVASCULAR SYSTEM

Blood serves as a transport medium.Specific functions

Carries: nutrients from the digestive tract to the tissues; end products from cells to the excretory organs; carbon dioxide from tissues to lungs; secretions of endocrine glands throughout the body.

Regulate body temperature Maintain constant concentration of water and electrolytes in the cells; Regulate body’s hydrogen ion concentration; Defend against microorganisms.

Homeostasis is the maintenance of uniformity and stability in this extracellular fluid through physiological processes of diffusion, pressure gradients, concentration gradients, active transport and regulatory mechanisms controlled by the nervous and endocrine systems.

BLOOD CELLS, PLASMA and SERUM

Three classes of blood cells (corpuscles):

1. Erythrocytes (red blood cells)2. Leukocytes (white cells)3. Thrombocytes (platelets)

Hemoglobin is the pigment that contributes to the red color of the blood contained in the erythrocytes.

Plasma is the yellow to colorless fluid in which all these cells are suspended. When examined as a thin film, it is always colorless. Bilirubin is the pigment that contributes to the color of plasma, although carotene and other pigments are contributing factors.

In coagulation, blood lost from the body becomes a gelatinous mass, after which the blood clot retracts, forcing from the clot a clear, watery fluid called serum – similar to plasma except that fibrinogen and other clotting factors have been removed.

Plasma may be obtained by adding to whole blood an anticoagulant to prevent clotting and letting the cells settle out, as they are heavier than plasma. Centrifuging the blood hastens the settling of cells to obtain plasma more readily.

ANTICOAGULANTS

FUNCTION: Anticoagulants are used to obtain blood samples free from clots for transfusions and for analytical work.

Commonly used anticoagulants in transfusions:

1. Heparin, a conjugated polysaccharide, naturally produced by basophils (a kind of leukocyte) in the blood and mast cells (part of the connective tissue surrounding capillaries in the lungs and other organs) throughout the body.Concentrations of this anticoagulant:0.2 mg of heparin per ml of blood

1 mg of heparin of 100 – 500 ml of blood at 0˚C & 10 – 20 ml at room temp.

2. Sodium citrate,as the citrate combines with calcium ions of the plasma forming an insoluble calcium salt.Caution: One must be careful not to give too much citrate because citrate can combine with sufficient calcium ions to produce tetany, and thus interfere with the functioning of nerves and of skeletal and cardiac muscles.Concentrations of this coagulant:0.2 – 0.4 percent of blood

Similar salts are also used exceptpotassium salts because of the possibility of producing a heart block.

Other useful coagulants in analytical work:

1. Sodium, potassium and ammonium salts of oxalates and fluorides.2. Chelating compounds such as ethylenediaminetetraacetic acid (EDTA)

Note: Ammonium salts are not recommended when compounds containing nitrogen are being determined quantitatively, because of nitrogen in the anticoagulant.

Anticoagulants and their effects on cell size:

Analytical work:

Heparin or EDTA keep the size of erythrocytes constant, are important in calculating mean corpuscular volume (MCV), mean corpuscular hemoglobin (MCH) and mean hemoglobin corpuscular concentration (MCHC) as diagnostic aids.

Ammonium salts increase the size Potassium salts decrease the size 6 mg of ammonium oxalate&4 mg of potassium oxalate inhibited coagulation of 5 ml blood &

kept size constant (Heller & Paul (1934))

ERYTHROCYTES

Erythrocytes are biconcave, circular discs varying in diameter and thickness according to species and nutritional status that are capable of undergoing changes in shape while passing through capillary beds.

Appearance /structure in animals:

Mammals – nonnucleated, nonmotile cells Dog – markedly concave Cat & horse – slightly concave Ruminants (cattle, sheep, goats) and pigs – discoid Most animals below mammals – elliptical and has nuclei.

Origin

Yolk sac produces nucleated red blood cells in early fetal development, later involving liver, spleen and lymph nodes.

Bone marrow, place where the formation of red blood cells (erythropoiesis) takes place after birth.

Under pathological conditions in postnatal life – liver, spleen and lymph nodes may again assume erythropoiesis.

Erythropoiesis (in bone marrow) goes on continuously and are poured into the blood stream at a rate to balance the destruction of red cells.

TRANSPORT IS THOUGHT TO HAPPEN IN TWO WAYS

The entrance of the newly formed corpuscles into a bone marrow capillary is more of penetration – no stoma or opening is necessary. Erythrocyte being nonmotile, enter the capillary by diapedesis( the passage of cell through the unruptured blood vessel wall).

According to another view, erythrocytes develop intravascularly, as there are two kinds of capillaries in the bone marrow:collapsed one which are erthrogenic , and open ones through which the blood flows. The young ones are forced into the blood stream as the erythrogenic capillaries become active and patent.

Metarubricyte, is the nucleated bone marrow cell which non nucleated erythrocyte is formed . The nucleus is lost by extrusion or absorption before the corpuscles enter the blood stream.

Normally 1 – 3 percent of erythrocytes are reticulocytes(young red blood cells that contains the remains of nucleus).

Pathological condition that increase their number in blood:

Hemorrhagic, hemolytic & other anemia – when bone marrow is more active in producing a great number of erythrocytes.Exception: horse

Composition:

In adult

62 – 72 percent water 35 percent solids (most part – hemoglobin; others: proteins, lipids, fat, vitamins, glucose,

enzymes and minerals)

Cations and anions (within & outside cell) help in establishing and maintaining electrical gradients across cell membranes by the sodium pump, active transport and diffusion (physiological processes that help in maintaining balanced electrolytes in cells).

Sodium is the principal cation in extracellular fluid.Potassium is the principal cation of erythrocytes of domestic animals except dog, cat, cattle, goats and sheep.

Erythrocyte and potassium values:

Dog – 8.7 and 107 Cat – 5.9 and 103.7 Cow – 21.8 and 79.8 Goat – 18.4 and 93.2 Sheep – 18.4 and 83.5, 64.2 and 15.6, 58.1 and 46.0 (depending on breed) Pig – 99.5 and 10.8 Chicken – 97.3 and 7.1 Turkey – 99.5 and 9.7

Size and hemoglobin content

There are many shortcomings in the measurement of the diameter of erythrocytes, making it less important than cubic volume (which should be used to measure cells.

The cells in the dry smear ( but in the moist state but losing water) are smaller Very few cells are measured from the blood sample taken The depth of the cell or third dimension is left out of account

The following formulas will give mean corpuscular volume, hemoglobin, and hemoglobin concentration in erythrocytes.

These formulas are an aid in diagnosing various types of anemias.

Iron deficiency anemia in mammals aremicrocytic type (very small cells)

MCVprovides the average size of cell size in cubic microns.

MCH expresses the average weight of hemoglobin present in the erythrocytes. MCHC gives the average percentage of the MCV which the hemoglobin occupies.

Pernicious anemia in man is macrocytic, but this anemia does not occur in animals.

Note: Error in counting erythrocytes contribute to variations in MCV and MCH.

Number

The number of red blood cells varies within species.

Plasma fluids are constantly being shifted across capillary walls, cell counts vary between arterial and venous blood samples.

Factors that affect erythrocyte counts, hemoglobin concentration, PCV & concentration of other blood constituents:

Age, sex, exercise, nutritional status, breed lactation, pregnancy, egg production, stage of estrous cycle excitement (release of epinephrine), blood volume (hemodilution or hemoconcentration), time of day, environmental temperature, altitude and other climatic factors.

Surface area

The surface area has great importance to the respiratory or gas-transport function of the blood.

Total erythrocyte surface can be estimated when blood volume, erythrocyte count, diameter, and thickness are known.

The estimated mean surface area per erythrocyte was calculated first from the dimensions reported on dry films. This value was increased by 20 percent to arrive at the estimated wet erythrocyte surface area.

The average thickness was 2 µ.

The figures for erythrocyte surface area compared to the body surface area – A man of average size has less than 2 sq m of body surface area, and a 500 kg cow somewhat less than 5.

Tonicity

Rule: In order to keep erythrocytes constant in size they must remain in the environment with the same osmolarity as blood plasma. When the osmotic pressure of plasma is lowered sufficiently, hemolysis (laking) of erythrocytes results.

3 Types of solutions:

1. hypotonic– solutions osmotically weaker than plasma, causing hemolysis of cells.Plasma osmotic pressure may be lowered by adding hypotonic salt solutions or water to blood. In such cases, water passes into the cells through osmosis through its semipermeable

membrane, causing the cell to swell. This results in the stretching and eventual mechanical rupture of its membrane, with hemoglobin passing through the surrounding medium.

2. Isotonic – solutions similar to plasma in crystalloid osmotic pressure, into which erythrocytes may be placed without resulting in cell volume changes.

3. Hypertonic – solutions that exert a higher osmotic pressure than blood plasma. They cause water to pass from the corpuscles by osmosis, and shrinking of the corpuscles results. Such erythrocytes are said to crenated.

Physiological salt solution

also known as physiological saline the isotonic solution of greatest interest. an aqueous solution of sodium chloride usually containing 0.85-0.9 percent sodium chloride.

Minimum resistance(osmotic resistance of the weakest corpuscles)

indicated by the concentration (%) of sodium chloride solution at which hemolysis begins.

Maximum resistance(resistance of the strongest corpuscles)

indicated by the concentration at which complete hemolysis.

It Is not known whether hemolysis caused by lowering the osmotic pressure of the plasma plays a part in the destruction of erythrocytes in the body under normal conditions. However, knowledge of osmotic resistance is of practical importance in preparing solutions for intravenous injection. Large amounts of water may be injected into the blood stream at low rates without producing any significant amount of hemolysis.

Processes that cause hemolysis of erythrocytes:

freezing and thawing stirring and agitation high temperatures substances that lower surface tension (saponins, soaps and bile salts) alcohol, ether, chloroform and acetone.

It is possible to calculate what percentage solution of a compound is isotonic with erythrocytes by using the following formula:

Life Span

Length of life for erythrocytes:

man – ranges from 90-140 days, averaging 120 days. For several small laboratory animals- life span is much shorter

The half-life of erythrocytes is 17 days. The half-life of autologous (individual’s own erythrocytes) transfusions are longer than homologous (same specie) transfusions.

Erythrocytes are destroyed in great numbers daily. The total number of erythrocytes in the body of a 450 kg animal with a blood volume of 8 percent of body weight is 300 trillion. If the average life of individual erythrocytes is 100 days, then 3 trillion must be destroyed (and formed) in the body every day, or about 35 million every second.

Pathological conditions:

Iron-deficiency anemia – erythrocytes become smaller in size Microcytic type of anemia – the result of younger cells not being released into the circulating

blood in sufficient quantities to replenish those being lost.

Fate of erythrocytes

Reticuloendothelial cells that destroy the old, exhausted erythrocytes:

Histiocytes Macrophages Clasmatocytes

Reticuloendothelial cells include the stellate or Kuppfer cells found in the liver, spleen, bone marrow and lymph nodes.

As erythrocytes are destroyed, the iron-containing moiety of hemoglobin is conserved and the pigmentary type is converted into bile pigment, an excretory product.

Liver and spleen are important storehouses of the iron that is not immediately used in the production of new hemoglobin.

The reticuloendothelial cells in different organs are important in the destruction of erythrocytes. Red bone marrow is the principal place or erythrocyte destruction ( and bile pigment formation of dogs). Spleen for man; liver for rabbit, guinea pig and birds.

Hematopoiesis

Hematopoiesis (the formation of red blood cells) is a continuous process.

Nutrients essential for this process:

Vitamin B12 (cyanocobalamin) – functions in the maturation of erythrocytes folic acid (pteroylglutamic acid)

Both vitamins act as coenzymes in the synthesis of nucleic acids or their constituents, purine and pyrimidine bases.

Other vitamins that aid are:

pyridoxine, riboflavin, nicotinic acid pantothenic acid, thiamine, biotin ascorbic acid

When these vitamins are deficient, growth and development of erythrocytes are impaired.

Pyridoxine deficiency in swine produces a microcytic, hypochromic anemia.

Other essentials:

minerals and amino acids water and energy

The minerals most commonly needed:

Iron – built into the hemoglobin molecule Copper – a coenzyme or catalyst in hemoglobin synthesis Cobalt – dietary essential for ruminants and is needed for bacterial synthesis of vitamin b12 in

the rumen.

Note:Cobalt given in excess will cause polycythemia. Though animals, if placed in high altitudes may also show polycythemia because of the decreased oxygen pressure (PO2).

Pathologic conditions that do not elicit the response for greater red cell production.

Nongenerativeanemias like Trichostrongylosis in cattle Anemia associate with chronic infection Neoplasia or other cachetic diseases

Pathologic conditions that elicit the responsefor greater red cell production.

Hemorrhage Hemolytic diseases Blood-sucking parasites Nutritional deficiencies

In hypoxic conditions (tissues are not supplied with sufficient oxygen) caused by inadequate number or improper functioning of erythrocytes) resulting in the release of erythropoietin, hemopoetin or erythrocyte maturing factor to blood plasma that stimulates hematopoiesis.Thekidney has been cited as producing this, but hypoxic cells in general may produce it.

PACKED CELL VOLUME

The volume of cells in the circulating blood is usually less than the plasma volume.

Hematocrit – test utilizing a centrifuge in which is placed a small tube containing a sample of blood with the appropriate kind and quantity of anticoagulant.

In the past

The Wintrobe hematocrit tube is centrifuged at 3000 rpm for 30 minutes; then the percentage of packed red cells is read from a scale or calculated.

As a rule in normal animals the volume of packed red cells (PCV) is directly related to the erythrocyte count and hemoglobin content.

With this rule, it became a common practice in some laboratories to allow the hematocrit tube containing blood to stand vertically for an hour before centrifugation in order to obating the erythrocyte sedimentation rate.

In Recent years

The microhematocrit centrifuge and tubes (with or without heparin) have been used widely to save time, values being obtained in 5 minutes.

The PCV is expressed as a percent volume of packed cells in whole blood after centrifugation.

Hemoconcentration due to dehydration, asphyxia, or excitement causing release of erythrocytes in the spleen can result in abnormally high PCV values. In excitement, the epinephrine causes splenic contractions.

HEMOGLOBIN

Hemoglobin, the pigment in erythrocytes, is a complex, iron-containing, conjugated protein composed of a pigment and a simple protein.

Globin – the protein, a histone. Heme – causes the red color, a metallic compound containing an iron atom.

Biosynthesis of hemoglobin starts in the erythroblasts and continues in the subsequent stages of development.

Four heme molecules unit with oneglobin to form hemoglobin.

The molecular weights of hemoglobin from most species are reported to vary from 66,000 to 69,000. Based on the iron content of hemoglobin as 0.334 percent and the atomic weight of iron being 55.84, a value of 16,700 as the minimal molecular weight of hemoglobin is obtained.

Hemoglobinuria occurs when erythrocytes are hemolyzed in the blood stream by protozoa, toxins or chemical agents, releasing hemoglobin into the plasma that is permitted passage by the basement membrane of the kidneys.

Fetal and Adult hemoglobin

At a given oxygen tension, fetal hemoglobin will take up more oxygen than adult hemoglobin.

The passage of oxygen from the mother to the fetus is by diffusion, the tension of oxygen in the blood of the umbilical vein is same as the venous blood on the maternal side of the placenta.

The production of fetal hemoglobin depends upon presence of amino acids at site of formation or origin of erythrocytes.

Infants may be born with one half to three fourths of their hemoglobin of the fetal type. By the end of the first year, fetal type of hemoglobin may reach 1 percent. The animal retains the ability to make fetal hemoglobin in adult life.

Amount

The amount of hemoglobin in the blood is expressed as g/100 ml of blood.

As a rule in most mammals normal blood hemoglobin values are between 13 and 15 g/100 ml. Exception: lactating cows. Cold-blooded: usually lower, being 12-13.

Excitement elevates erythrocyte numbers per unit of volume, easily demonstrated with anesthesized dog after injection of epinephrine.

The hemoglobin concentration in the avian blood is more difficult to determine. A method has been devised to determine hemoglobin in bloof of chickens and values obtained are similar to those when iron method is used. Normal hemoglobin concentration in chicken ranges from 6.5 to 9 g/100 ml by this method.

Conditions that lower the oxygen content of blood, like elevated barometric pressure, cause an increase in the production of hemoglobin and the number of erythrocytes and vice versa.