Anatomy and Human Biology 2214M. Hall

July 27, 2009

CARDIOVASCULAR SYSTEM LAB

Objectives.

Find and recognize the blood vessels in a tissue section.
Recognize the layers of the walls of arteries, veins and the heart
Distinguish the following structures on your slides:

muscular arteries / venules = small veins
tunica intima / medium-sized veins
endothelium / vena cava
tunica media / heart
smooth muscle cells / atrium
tunica adventitia / ventricle
internal elastic membrane / endocardium
external elastic membrane / subendocardial layer
elastic arteries / aorta / myocardium
arterioles = small arteries / coronary artery
capillaries / heart valves
pericyte / Purkinje fibers

Arteries

Arteries have to cope with the pressure pulses generated by the heart. The large arteries close to the heart contain very large amounts of elastica in their media, which allows them to function as a temporary pressure reservoir. These include the aorta and some of the first arteries that branch from it. Their walls have dozens of sheets of elastica arranged circularly. Further down the vascular tree the arteries are dominated by smooth muscle in their media layer. These arteries are called muscular or distributing or, simply, medium sized arteries. Their main function is to regulate the amount of blood that flows to various parts of the body. The still smaller arteries are called arterioles or small arteries. Their function is also to distribute blood at controlled rates to specific regions of tissue. They also regulate blood pressure, since the small arterioles are the ones that provide the greatest resistance to blood flow. There are no concrete criteria for distinguishing the smallest distributing arteries from arterioles.

Veins

Three categories of veins are distinguished: small, medium and large. Veins with diameters less than 2 mm are referred to as small veins or venules. These have two main layers: an intima and adventitia. A few smooth muscle cells may be present as a media in larger small veins. The adventitia consistsof loose connective tissue with thick bundles of longitudinally oriented collagen and elastic fibers.

Medium-sized veins (2-9 mm) possess an intima consisting of an endothelium and an inconspicuous connective tissue layer. The media usually is visible but much thinner than in corresponding arteries. It consists mainly of circularly arranged smooth muscle and collagenous fibers. The adventitia is thicker than the media. It contains connective tissue with thick longitudinal collagenous bundles and elastic fibers.

Large veins have an intima similar in structure to that of the medium-sized veins. The media also is similar in structure to the medium-sized veins but with more smooth muscle. The adventitia is well developed. It contains prominent longitudinal layers of smooth muscle. It also has thick elastic fibers.

The most obvious structural difference between arteries and veins is the thickness of the tunica media.

Slide descriptions

D-121. Pancreas, Monkey (Masson Stain). This section contains a typical medium sized, muscular ( = distributing) artery. Hold the slide up and locate the artery off to one side as a hole about a millimeter across ringed in blue. Examine it through the 10X objective with which the entire artery should fit into view. The staining procedure colored collagen blue-green, elastica white and cells red, in particular the muscle cells in the artery. The radial lines in the arterial wall are artifact where the section of tissue has buckled instead of lying flat on the slide. Such problems are not uncommon for tissue with a lot of elastica.

The tunica media of the arterial wall is instantly recognizable because it is composed mainly of red-colored smooth muscle with some collagen. It ends internally in a colorless sheet of elastica, the internal elastic membrane. That structure and the traces of tissue inside of it constitute the intima. Along one side of the artery the intima is pathologically thickened. Smooth muscle cells have invaded the intima to form an arteriosclerotic lesion.

The tunica media is surrounded by the tunica adventitia, composed of dense connective tissue. The adventitia is about 1/4 - 1/2 the width of the media. The whole artery runs in a seam of (blue) connective tissue between bright red lobules of pancreatic parenchyma.

Examine the individual layers of the wall more closely at high (40X) power. The smooth muscle cells of the media are circularly arranged, as are the collagen fibers. This layer is very dense. The internal elastic membrane is prominent. It has puckered up the inner surface but if the artery were distended that surface would be smooth. In most places the intimal surface is well preserved and you can see the flattened nuclei of the endothelial cells resting on a minimal layer of connective tissue.

The boundary of the tunica adventitia with the tunica media is characteristically very sharp. The outer edge of the adventitia is less obvious but still recognizable. Let me reiterate an important structural generalization about arteries and veins: the components of the media layer are arranged circularly while those of the adventitia run mainly longitudinally.

Disregard that arteriosclerotic plaque. Your large arteries are probably riddled with plaques such as this. Eventually some will grow so large as to impede the flow of blood. If this happens in your coronary arteries you may welcome a bypass operation. Better push your plate away from a second helping of Australian mutton stew and learn to like bread without butter.

Before leaving the slide go back to low power and scan around for the large duct in the middle of the section. It too is a hollow tube running in a seam of connective tissue. A glance at high power immediately distinguishes it from an artery. It is lined with tall epithelium instead of very squamous endothelium.

D-22 Aorta (elastic stain) The largest arteries in the bodies are of the elastic variety. Their media is modified by having many sheets of elastica wrapped around. Smooth muscle and collagen lie between these sheets. The aorta is the prime example, stained here to reveal the elastica. Its resilient elastic wall is quite thick, but in proportion to the size of the lumen it is thinner than the wall of muscular arteries. The elastic stain readily distinguishes the three tunics in the wall.The tunica media contains an especially large amount of elastica. The intima and adventitia, with less elastica are distinctly paler and redder

Go to 40X magnification.The concentric, fenestrated elastic membranes show up well. Most of the cells are smooth muscle cells also arranged circularly. The outermost elastic sheet marks the boundary between the media and the adventitia. The adventitiais composed primarily of thick, longitudinally oriented bundles of collagen. The elastica here is also mostly in the form of longitudinal fibers.These show up as black dots and dashes at 40X.

The intima is less satisfactorily presented on these slides. Its endothelium has been lost from large regions of the aorta.

D-23 Aorta (H&E). Compare the aorta stained with familiar (H&E) with that on the last slide. First distinguish the intima, media and adventitia layers. Then examine the media at higher magnification. You should be able to discern the elastic lamina which appear as blank pale wavy bands. The collagen appears as irregular wavy wisps accompanied by spaces resulting from tissue shrinkage. The important point is to recognize that this wall looks substantially different from that of a muscular artery even when stained with H&E.

D-162 Scalp. The scalp is a good place to look for small blood vessels. Determine which edge of the section is the skin surface (it is covered with epithelium) and which is the internal edge (the ragged edge where the connective tissue holding the skin to the underlying tissues was separated). Look for blood vessels close to the internal edge. Start of by picking out a small, but not tiny, arteriole with several layers of smooth muscle in its wall. The intima consists of the flattened endothelium, an all but invisible trace of connective tissue and the internal elastic membrane.There should be a discernible adventitia, although the outer extent of that layer may be indistinct.

Next, pick out several venules. They will be in various stages of collapse. Their thin walls are mostly adventitia. The endothelial cells should show up well along the edge of the lumen. The somewhat elongated nuclei farther out belong to fibroblasts. You may find a smooth muscle cell, but probably not unless the venule is quite large. Conveniently, arterioles (and arteries) often are accompanied by comparably sized venules, making the comparison between the two easy.

Now look at the smaller vessels. A variety of characteristics can help to distinguish small arterioles from venules. The most definitive is the presence or absence of smooth muscle cells in the media. Small arteries and arterioles have them, small veins and venules don’t. Large venules may have a scattering of smooth muscle cells but MANY fewer than arterioles of comparable size

Finally you might (or might not) be able to recognize alymphatic vessel. Its wall will be much thinner and its lumen much larger than a venule. Lymphatic capillaries usually are collapsed and their endothelial cell nuclei look like scattered fibroblasts lying between collagen fibers.

By the time you finish with this slide you should be able to identify most of the blood vessels that you come across..

D-2 Adipose tissue Adipose tissue is well supplied with smallarteries and venulesand is especially rich in capillaries. The capillaries consist of a single layer endothelial cells, sometimes with an associated pericyte. They often run where three fat cells come together. You can find nice examples of longitudinal as well as transverse sections. Capillaries are large enough in diameter for only one red blood cell to pass through at a time.

Arterioles, of course, have one or more layers of concentrically arranged smooth muscle cells wrapped around a simple endothelium. You will see few here, so do not get your hopes up.

D- 27 Vena cava (Masson). The vena cava is the largest vein in the body. Its tunica media is easily visible as a relatively narrow layer of smooth muscle (red) and collagen (blue) arranged circumferentially. Like all veins, the adventitia of the vena cava is much thicker than the media. Here it is composed mainly of bundles of smooth muscle arranged longitudinally. Connective tissue fills in between the bundles. There is a pathologicalthickening along one side. Either ignore it or note that it is formed by circularly arranged smooth muscle cells invading into the inner layers of the vessel.

D-19 The heart. Hold up this slide as shown in the picture below and then rotate it 90 degrees counterclockwise. Now it is in its ordinary orientation. The atrium is at the top and the ventricle below. The outside of the heart is to the left and the lumenof the chambers, lined with endothelium is to the right. The heart valve is the thin line of tissue extending downwards. It is a thin sheet of connective tissue, and in this section shows up as a narrow band. Note that it attaches to the wall of the heart at the junction between the atrium and the ventricle.

It is obvious that a substantial ring of connective tissue lies between the upper and lower chambers of the heart. The prominent round structure is a section of a coronary artery. This artery is unusual in that it has two discrete layers of smooth muscle in its media. Its intima also has yucky pathology. (I told you to stay away from Australian mutton).

Begin by examining the wall of the ventricle under the 10X objective. The main layer is the myocardium, which corresponds to the tunica media of an artery. It is a thick band of darkly staining cardiac muscle. The endocardiumis a thin layer, consisting of an endothelium and some underlying connective tissue. Check to see that it varies in thickness from place to place, and is more developed in the atrium than in the ventricle.

The epicardium is a fairly thick layer of loose connective tissue. It has a large number of fat cells, making it stain palely. (Fat cells are a dead giveaway for distinguishing the epicardium from the endocardium.)A single layer of mesothelial cells covers the free surface of the epicardium

Examine the atrium. Note that it has a much thinner myocardial layer and a thicker endocardium than does the ventricle. Examine the valve and note that is covered on both sides by endothelial cells—it is an extension of the endocardium. It consists of fairly dense fibroelastic tissue.

D-18 Left atrium (elastic stain).The atriumhas the same three layers as the ventricle. However, the myocardium is thinner and the endocardium thicker. Examine the endocardium. Itis very well developed. In fact, its lower part is often recognized as a "subendocardial layer". This is where most of the elastica of the atrium is found.

D-20 Purkinje fibers. (H & E). This slide has the interventricular septum cut horizontally. It demonstrates a specialized type of cardiac muscle cell, the Purkinje fiber. Purkinje fibers form the conduction system of the heart. Bands of these cells run from the atrium down along the interventricular septum to the apex of the heart to make the ventricles contract from the bottom up. Because the interventricular septum is within the heart, both sides of the septum you see are covered with endocardium. The Purkinje fibers, cut transversely, appear subendocardially on both sides. They may be distinguished from normal cardiac muscle cells by the following criteria :

They are wider and lighter staining than cardiac muscle cells.
Except for the nucleus, the central part of the Purkinje cell appears empty. This is because it contains a lot of glycogen, which doesn’t stain well with H & E.
The few myofibrils are confined to the periphery of the cell.
They are often arranged in groups of 4 or 5 cells, surrounded by connective tissue
A substantial number of Purkinje cells have two nuclei

D-121 Pancreas (H & E)D-22 Aorta (Elastic Stain)

D-23 Aorta (H & E)D-162 Scalp (H & E)

D-2 Adipose Tissue (H & E)D-27 Vena Cava (Masson Stain)

D-19 Heart (H & E)D-18 Left Atrium (Elastic Stain)

D-20 Purkinje fibers (H & E)