First of all, the identification number of the animal should be properly recorded and recorded on the cassettes to be used..
In the case of laboratory animals, the animal should be weighed, as well as all the organs sampled. This will be very useful to calculate the ratio between the weight of the organs and the total body weight. These data are of special interest when the animals come from a toxicopathology project.
- Assessment of the animal's general condition and body condition.
- Record in detail any external lesions, either superficial on the skin or deeper under the skin.
- Examination of mucous membranes and body cavities, appreciating any changes in coloration or other abnormalities.
- Examination of the perineal region for signs of diarrhea, rectal or vaginal prolapse or any other lesions.
- Gently palpate the abdomen for any masses or fluid. If masses are found, make a description of their consistency. If the abdomen is distended and full of fluid, it is advisable to take a sample with a sterile syringe for evaluation.
Incision of the skin and examination of the subcutaneous tissue:
- If a dissection plate is available, the animal is fixed on it in dorsal recumbence and moisten the skin with 70º alcohol.
- An incision in the skin along the midline from the mandible to the pubis is made. In the case of males, the incision will end on both sides of the penis.
- Remove the skin to both sides of the incision, examining the skin and subcutaneous tissue.
- Examine the mammary glands: mice have 5 pairs (3 thoracic and 2 abdominal), while rats have 6 pairs (3 thoracic and 3 abdominal). In females the mammary tissue extends from the mandibular region to the base of the tail, occupying almost the entire ventral area of the abdomen and thorax.
- Examine the superficial lymph nodes, which under physiological conditions will be bilateral, grayish and small in size. The most commonly examined are the mandibular, axillary and/or popliteal.
- Examine the three pairs of salivary glands located on both sides of the cranioventral region of the neck: mandibular, sublingual and parotid. Take samples from all three by gently pulling them from the most caudal portion cranially.
- Examination of the extraorbital lacrimal glands, located in the ventrolateral region of the head, where they appear as a flat grayish-brown glandular tissue.
- In the inguinal region the presence of clitoral glands in females and preputial glands in males is observed. Both are modified sebaceous glands that appear immersed in the subcutaneous adipose tissue.
- Examination of the penis and prepuce in males, which will be removed with the rest of the genital organs.
Opening of the abdominal cavity:
- The cavity is opened along the midline, removing the skin to both sides and the organs are examined in situ, as well as the serous membrane (peritoneum) and the presence of abnormal contents in the cavity..
- Assessment of the state of fatness according to the fat deposits found.
- Located in the left upper abdominal quadrant is the spleen, which is removed by cutting the gastro-splenic ligament. It is an elongated, oval, slightly curved organ with an intense red color.
- Dissection of the anus from the surrounding skin and opening of the pelvic cavity.
- Removal of the gastro-intestinal bundle (abdominal esophagus, stomach, small intestine, large intestine and mesenteric lymph nodes) in a cranial-caudal direction, cutting the insertion of the mesentery as close as possible to the intestine. Separate the intestinal loops carefully.
- The pancreas in rodents is diffuse, with a left lobe located near the spleen and a right lobe adjacent to the duodenum. The left lobe is sampled; the right lobe will be sampled with the duodenum.
- The stomach has two distinct regions: an anterior or proventricular one, with a whitish appearance, and a glandular one, more reddish and with a thicker wall. The opening is made through the greater curvature and a careful washing of the mucosa with saline solution or fixative.
- The duodenum is short in rodents (only one centimeter), while jejunum and ileum are grossly indistinguishable. The large intestine consists of the cecum, colon and rectum. Cross-sectional samples of 2-3 mm are taken from each section of intestine without opening it and then the remaining intestine is opened to assess the mucosa.
- The liver has four lobes: medial, right medial, right lateral and caudate, with a papillary process. There is no gallbladder in the rat. It is a fragile organ that has to be handled with care. In the rat a portion of the left lateral lobe and the right medial lobe will be sampled; while in the mouse a portion of the left medial lobe and the gallbladder will also be sampled.
- The adrenal glands are small, whitish structures located in the perirenal fat, larger, pink and translucent in males; while in females they are smaller, whitish and opaque. Because of their small size they are fixed in toto (complete).
- The kidneys are organs located in the dorsal wall of the abdominal cavity, bean-shaped and reddish-brown in color. Samples are taken in toto from both kidneys or at least a longitudinal portion of one kidney and a transverse portion of the other kidney).
- In males:
- Extraction and examination of the testes in the case of males, located in intra-abdominal position or inside the scrotum.
- Assessment of male accessory glands: seminal vesicles, prostate (two ventral and two dorsolateral lobes) and coagulative glands (located dorsocranial to the prostate).
- Removal of the penis, foreskin and urethra.
- In females:
- Dissect the vulva and vagina free of skin and cut the supporting ligaments of the entire reproductive tract, assessing each of the organs.
- Special importance of the ovaries, which are located within the perirenal fatty tissue, near the caudal poles of both kidneys.
Opening of the thoracic cavity:
- Make two lateral cuts on each side of the rib cage and another transverse cut in the most cranial portion of the sternum to expose the thoracic organs.
- With scissors make a cut of the ventral thoracic musculature, including all those over the trachea.
- Make a transverse section of the trachea near the mandible and pull it in a ventro-caudal direction, cutting the connections of the various cervical and thoracic organs with their supporting tissue, until the entire bundle of trachea, thymus, lungs and heart is removed.
- In some cases, formaldehyde can be introduced with a syringe through the tracheal opening to ensure proper fixation of the tissues, taking special care not to overinflate the lungs.
- Opening of the heart to visualize the valves and the endocardium of the four chambers.
Opening of the cranial cavity:
- Place the cadaver in sternal recumbence and remove the skin and musculature covering the cranial vault, and then proceed to decapitate.
- Using small sharp-tipped scissors make an upward cut in the midline of the cranial vault introducing the tip through the foramen magnum.
- Using forceps remove both halves of the cranial vault leaving the brain exposed.
- It is advisable to introduce the complete skull in formalin, with the encephalon inside the cavity, but exposed to improve the fixation of the tissue.
In particular, toxicology studies should be carried out in accordance with the OECD 407 protocol, according to which a complete, orderly and systematic necropsy of the animals should be performed with special attention to the body surface, natural orifices, as well as the observation of the cranial, thoracic and abdominal cavities and their contents. A thorough study of liver, kidneys, adrenals, testicles, epididymides, prostate and seminal vesicles with accessory sex glands (as a whole), thymus, spleen, brain and heart of all animals is recommended; apart from those moribund or euthanized before the end date of the study. Any adherent tissue should be removed as appropriate for fresh weighing as soon as possible. Alternatively, seminal vesicles and prostate may be weighed subsequent to fixation. In addition to these tissues, fresh weighing of tissues such as the ovaries and uterus (including the cervix) or after fixation, such as the thyroid, can optionally be performed.