Preparation of Ringer's solution
Dissolve 6.5 g of sodium chloride (NaCl), 0.14 g of potassium chloride (KCl), and 0.20 g of sodium bicarbonate (NaHCO3) in 800 mL of distilled water. Also, dissolve 0.16 g of hydrated calcium chloride (CaCl2.2H2O) and 0.39 g of hydrated magnesium sulfate (MgSO4.7H2O) in 100 mL of distilled water. Add the 100 mL cautiously to the 800 mL, stirring vigorously. Then make the final volume equal to 1 liter by adding more distilled water and filter if necessary.
Sacrifice the frog
The frog species to be used in this exercise is Rana pipiens. The preferred method of sacrificing the frog is known as the double pith procedure. The frog is held with the thumb and fingers securing the limbs and the index finger depressing the snout so that the animal's head is at an angle to its body. A dissecting needle is run down the middle of the frog's head until, about 2 to 3 mm behind the posterior border of the eardrum, a depression is felt. This depression marks the location of the foramen magnum, a large opening in the skull (see attached figure, Part A). The dissecting needle is then forced through this opening and the needle is twisted to destroy the brain. The dissecting needle is then forced into the spinal column and pushed down until the legs extend and relax.
Excision of the sciatic nerve and gastrocnemius muscle
Lay the double-pithed frog with its ventral side up. Using forceps, raise the skin at the midline of the body between the forelegs and then cut the skin all the way around the body just below the forelegs. Tightly grasp one forepaw with one hand and, with the other, strip the skin down and off the hindlegs with one pull (see attached figure, Part B). Wash the frog in Ringer's and lay it ventral side up. Cut through the abdominal muscles and remove the organs to expose the three roots of the sciatic nerve on either side of the vertebral column. Turn the animal over and separate...