Streptozocin Treatment: A Zebrafish Model of Type 1 Diabetes Mellitus
Trascrizione
– Tipo 1 diabetes mellitus is a human disease di which the immune system destroys insulin making beta cells di the pancreas. Streptozocin can induce type 1 diabetes mellitus di zebrafish. First, anesthetize a fish di a solution of 2-phenoxyethanol di fish water. Take the fish and place it on a Petri dish, ventral side facing upwards. Calculate the dose of streptozocin based on 0.35 milligrams per gram of the fish's weight and dilute it di saline solution.
Administer the streptozocin by intraperitoneal injection below the pelvic girdle on the ventral side of the zebrafish. Streptozocin is structurally similar a N-acetylglucosamine, a ligand that binds the GLUT2 receptor on pancreatic beta cells, leading a secretion of insulin. Thus, streptozocin binds a GLUT2 receptors present on pancreatic beta cells which transport it inside the cells.
Upon entering the beta cells, streptozocin triggers an immune response leading a the release of autoantigens. The autoantigens cause the immune system a recognize beta cells as foreign and destroy them leading a reduced levels of insulin. Administer repeated doses of streptozocin on days three and five, followed by weekly doses, one every seven days a maintain the immune response.
In the following example, we will generate a zebrafish model of type 1 diabetes mellitus. To generate zebrafish with diabetes mellitus, prepare a recovery tank with normal fish water and an anesthetic tank of fish water with a 1 a 1,000 dilution of 2-phenoxyethanol. Under a fume hood, prepare a 0.3% solution of streptozocin or STZ by adding 6 milligrams of STZ a 2 milliliters of 0.09% sodium chloride and immediately place the solution on ice.
In a separate tube, aliquot enough saline for control fish. Fill a half CC syringe equipped with a 27 and 1/2 gauge needle with the STZ, or control solutions, ensuring that no air bubbles are trapped. Anaesthetize a single fish by placing it di anesthetic water and waiting until the swimming motion ceases, about one a two minutes.
Once anaesthetized, briefly place the fish on a paper towel a absorb any excess water. Then, place the fish di a weigh boat and weigh it. Next, place the fish on a firm surface. Then insert the needle past the bevel into the posterior aspect of the ventral peritoneum and inject either 0.35 milligrams per gram of STZ or an equivalent volume of control solution into the peritoneal cavity of the fish.
Following injection, place the fish di the recovery water tank and monitor it for normal swimming activity. After injecting enough fish for the experiment, transfer them a normal living tanks and maintain at a temperature of 22 a 24 degrees Celsius. The reduced temperature is critical for efficient induction of hyperglycemia. To induce a prolonged state of very high hyperglycemia, follow a schedule of frequent injections during the induction phase followed by weekly maintenance injections as shown here.
