The overall goal of this procedure is to gain strict control over the otherwise fluctuating serum estrogen concentrations in female rats and mice. This is accomplished by first removing the ovaries by the dorsal root. Next cel elastic capsules and a Nutella mixture are prepared.
Finally, a capsule is inserted subcutaneously, or the Nutella mixture is administered orally. Ultimately, results can be obtained using radioimmunoassay that shows stable or fluctuating serum 17 beta estradiol concentrations for the capsule and Nutella methods respectively. The main advantage of these techniques over existing methods like slow release pellets, is that they have been proven to produce physiological concentrations of estradiol for prolonged periods of time.
This method can help answer key questions in the estrogen field, such as differences in effects between administration, roots and doses. After anesthetizing the animal, place it in a prone position on a heating plate connected to a rectal thermometer. Apply IGEL and for postoperative relief on the lateral aspect of the animal's Neck subcutaneously.
Administer five milligram per kilogram IL in saline anteriorly from the iliac crest shaver four by four centimeter area. Then disinfect the area with an antiseptic before covering the animal with a four by four centimeter aperture surgery sheet. Make a two to three centimeter midline incision and bluntly dissect the skin from the under underlying facia, one centimeter lateral of the midline.
Make another incision through the fascia. Then as superficially as possible, bluntly dissect laterally under this fascia until the abdominal cavity is reached. Using tweezers, grab the adipose tissue that surrounds the ovary in the abdominal cavity and gently pull it out.
Identify the ovary and ligate the uterine horns and vessels. 0.5 to one centimeter proximally to this structure. Cut out the ovary and ligated adipose tissue and put the remaining tissue back into the abdominal cavity.
Make another incision in the contralateral fascia and remove the other ovary. Close the wounds, and if not immediately administering 17 beta estradiol. Let the animal awake from anesthesia.
Place the animal in a heated cage for at least two hours after surgery. Housed the rat separately for the first few days postoperatively. 24 hours after surgery subcutaneously.
Inject the animals with another five milligram per kilogram IL to administer 17 beta estradiol through scholastic capsules. Thoroughly mix the desired amount in sesame oil. Cut three centimeter lengths of scholastic tubing and five millimeter lengths of wooden applicator sticks for plugging Using forceps.
Squeeze one wooden plug into the scholastic capsule before injecting the 17 beta estradiol sesame oil solution. To fill the entire length, gently cap the scholastic capsule on the open end with another wooden plug. Incubate the capsules in remaining 17 beta estradiol solution overnight before implantation.
After closing the ectomy incisions, shave and disinfect a two by two centimeter square area on the dorsal aspect of the rat's neck. Make a one centimeter incision and cord laterally and bluntly. Dissect a subcutaneous pocket making ample space for the capsule.
Wipe off the scholastic capsule and gently insert it through the incision. Then close the incision with a suture. Ensure that the capsule strains the skin as little as possible, particularly around the wound.
To use the nut cream Nutella as a medium of estrogen administration thoroughly dissolve 17 beta estradiol in sesame oil, and then mix the sesame oil with the nut cream. Prepare a placebo nut cream without the 17 beta estradiol for five days prior to the experiment. Train the rats to eat hormone free nut cream on days one through three.
Train the animals in groups on the fourth and fifth days. Feed them the nut cream in separate cages throughout the experiments. Place the animals in separate cages when they are fed the nut cream physiological serum.
17 beta estradiol concentrations, which the current methods aim to reproduce are approximately five to 140 picograms per milliliter for rats, and five to 35 picograms per milliliter for mice. As shown here. When administered to 300 gram female spread doy rats.
The subcutaneous elastic capsules produce slowly decreasing 17 beta estradiol serum concentrations from around 40 picograms per milliliter on day two to 10 picograms per milliliter by day 35. However, immediately after administration, a serum concentration peak of approximately 1000 picogram per milliliter can be expected falling to the physiological range within eight hours when the capsules were administered to 25 gram female. C 57 black six mice.
Serum concentration slowly decreased from approximately 90 picograms per milliliter on day two to 25 picograms per milliliter on day 35 when administered via the nut cream to 300 grams. Spra do rats. Physiological levels of 17 beta estradiol fluctuated daily between 10 to 70 picograms per milliliter in mice.
The 17 beta estradiol levels fluctuated between 20 and 120 picograms per milliliter Once mastered, the technique of ectomy and capsule implantation can be done in 10 minutes if performed properly. While attempting this procedure, it's important to remember to strictly adhere to hygienic guidelines.
