Dissection Techniques and Histological Sampling of the Heart in Large Animal Models for Cardiovascular Diseases

The structural differences in the heart between species and individuals, as well as discrepancies in the literature necessitate a thorough understanding of heart anatomy and adherence to strict methodology. This technique can improve observer uniformity and increase the comparability and the reproducibility of experimental cardiovascular disease research. A person who is unfamiliar with this technique may have difficulty identifying all of the heart's components.

My recommendation is to spend some time getting acquainted with the heart's anatomy. Because the heart's complex anatomy is difficult to comprehend, a visual presentation of the method is required. To begin, weigh the subject animal before placing it on the necropsy table on its left side.

Then perform an external examination of the subject by inspecting and palpating the cutis and mucosa, noting all changes in color, consistency, and volume. Next, using a knife, make a sharp seven-centimeter incision in the right axilla. After locating the coxofemoral junction, open the capsule with a knife and cut the ligament of the head of the femur.

Extend the skin incision in the axilla cranial to the mandibular symphysis and caudally to the peroneum. Remove the skin with the scalpel, exposing the thoracic and abdominal wall from the ventral midline to the level of the vertebral transverse processes. Next, using the scalpel and forceps, create a flap through the abdominal wall, ventrocaudal to the right costal arch.

Then, following the caudal aspect of the right hypochondrium, open the abdominal cavity with scissors. Continue extending the incision caudally along the right paralumbar area until the external iliac crest. Afterward, extend the incision until the pubic insertion of the linea alba and briefly examine the abdominal organs in situ to note any change in their color, consistency, volume, and position.

Next, using the scalpel and forceps, puncture the diaphragm at the highest point. Then using bone-cutting forceps, remove the right ribcage by first sectioning the parasternal cartilaginous segment and then the dorsal segment of the ribs proximal to the costovertebral joints. Examine the heart in situ to note its size, color, position and connections with adjacent tissues.

Also, look for any change in consistency by palpation. Then, using scissors, open the pericardial cavity with a longitudinal section from the base to the apex of the heart. Examine the pericardial cavity and the epicardium for any change in color, consistency and volume.

Next, using scissors, release the heart by sectioning the superior vena cava at least one centimeter above the entrance into the right atrium, preserving the crista terminalis and yielding the transverse sections of the aorta, pulmonary trunk, and both vena cavae. Examine the external contour of the organ, the appearance of the epicardium, that of the myocardium by the transparency of the epicardium, and the external appearance of the large vessels. To perform the inflow-outflow method for cardiac dissection, place the heart with the atrial face upward, and using scissors and toothless forceps, broadly cut from the caudal vena cava into the right atrium.

After cutting the heart to the right auricular appendage to expose the crista terminalis or the sinoatrial node, inspect the tricuspid valve from the dorsal view. Then, by cutting a section from the right atrium to the junction between the right ventricle and intraventricular septum, display the right atrioventricular valve. Next, section the right ventricle free wall along the intraventricular septum till the origin of the pulmonary trunk.

After thoroughly examining the right atrioventricular valves, including the chordae tendineae and the papillary muscles, cut the chordae tendineae. Next, place the forceps along the pulmonary trunk and cut the trunk longitudinally to examine the pulmonary outflow tract, pulmonary trunk origin, and semilunar valves. To inspect the bicuspid valve from the dorsal view, cut the left atrium from the pulmonary veins to the tip of the left auricle.

Next, place the heart with the interventricular septum facing downwards and make a large incision in the ventricular free wall from the ventricle base to the cardiac apex. Then, cut the chordae tendineae of the cusp of the left atrioventricular valve. Finally, open the aorta by placing forceps through the left ventricular outflow tract.

Use saline to remove all blood clots and weigh the heart. For taking samples for histology, first, make two longitudinal parallel cuts, approximately five millimeters apart, with a scalpel involving the right atrium, tricuspid valve, and the right ventricular free wall. Next, make two similar parallel cuts involving the left atrium, mitral valve, dorsal papillary muscle, and left ventricle.

Within the dorsal upper third of the heart, transversally section the base of the heart following a line, including the interventricular septum, right atrium, aorta, aortic valve, and tricuspid valve. And make two parallel longitudinal cuts involving the base of the aorta and the interventricular septum. Then, trim the right atrium around the crista terminalis.

To obtain six pieces of the sinoatrial node, make several parallel sections about three to four millimeters apart, oriented transversally on the longitudinal axis of the crista. Next, trim the obtained tissues in histological cassettes and immerse the tissues in 10%neutral buffered formalin for at least 48 hours. To sample the coronary arteries, use forceps and a scalpel to dissect the coronary arteries from the paraconal groove and fix them in 10%neutral buffered formalin overnight.

If needed, decalcify the tissue in a one-to-one mixture of 8%formic acid and hydrochloric acid. Next, make several transverse cuts about three millimeters apart. Place the sections in a histological cassette and put the cassette in 10%neutral buffered formalin for embedding.

After fixing the heart in 10%neutral buffered formalin for at least 48 hours, place two toothless straight dissection forceps. One through the cranial cava, right atrium, tricuspid valve, and right ventricle. And the other through the pulmonary vein, left atrium, mitral valve, and left ventricle.

Utilizing the space between the forceps as guidance, use a cutting blade to cut from the base to the apex of the heart. About five millimeters apart, cut another longitudinal section of the cardiac segment that includes the aorta. Then, completely separate the cardiac base from the apex.

After placing each tissue segment in a histological cassette, place the cassettes in 10%neutral buffered formalin until embedding. This protocol enabled successful visualization of the external features and internal structures of the heart. Using this technique, heart tissue samples could be successfully collected for histological studies from different animals like dogs and cats.

Always be cautious while releasing the heart because it may damage some structures of interest. And also, during dissecting, harvesting, and embedding the sinoatrial node. Although this method was created to diagnose arrhythmia modeling, hypertrophic cardiomyopathy, myocardial infarction and cardiac intoxication, it can be used to diagnose any cardiac condition.