Abstract
The principle of the method to be described for testing pituitary chromatophorotropic substances is based on the fact that the dermal melanophores of excised skin of Anolis carolinensis respond to pituitary chromatophorotropin 1 by dispersion of the melanosomes.
The use of hypophysectomized frogs and lizards and light adapted frogs involves several undesirable conditions. They are all time-consuming, and in the case of operated reptiles, endocrine imbalance, especially thyroid effects, are involved. The use of excised skin of Anolis as a test object for measuring intensity of effect, is very rapid, entails no difficult operative procedure, requires only small amounts of material, and eliminates the influences of other possible endocrine or neural disturbances. The test has been found to be reliable. Uniform responses are produced on skin of wellfed animals 100% of the time without difficulty. The complete dispersion of melanosomes is easily detected since the color change from green to brown is very striking.
Acetone-dried pituitaries of several vertebrates (man, beef and dog) have been tested and complete assay of pituitaries of Rana catesbeiana has been made by this method. It has been found that frog material is best employed in dilutions of 1 mg in 20 cc cold-blood Ringer's solution. Stronger concentrations react upon the skin too rapidly for accurate observations to be made. The solution is refrigerated 24 hours and then filtered. The filtrate is saved for the test. It has been found that the unfiltered solution shows partial inactivation of the hormone after 24 hours at room temperature. Complete inactivation is accomplished after 48 hours.
Lizards used for such tests should be well-fed and watered daily, since variations in reactions or complete unresponsiveness results when the animals are not fed and watered regularly.
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