OSMO-REGULATION 135 



titles. Such emiryos as are available may be segregated into stages and if there are suf- 

 ficient numbers, some may be deprived of their Jelly and vitelline membranes. So far as 

 possible test the covered and nude eggs against the same media, using identical and suf- 

 ficient numbers in each medium so that the results may be considered statistically valid. 

 It is of utmost importance that all conditions of relative volirme of medium per egg, 

 temperature, etc. are identical. 



Experimental Procedure : 



Prepare 2 finger bowls for each of the media, each of which should receive 

 50 cc. of the solution to be tested. Place 25 eggs or embryos in each finger bowl, 

 all of the same stage of development. The ratio must be 2 cc. of medium per egg or 

 embryo. With Urodele material, which Is less abundant, the volume of medium need 

 not be reduced but the number of embryos per container may be less than 25 but must 

 remain the same for all. The environmental conditions of temperature, light, etc. 

 must be identical for all so that the only variable is the medium being tested 

 against embryonic development. 



Eggs and embryos that are to be deprived of their jelly and vitelline membranes 

 should first be placed in the test medium and then denuded. 



OBSEHVATIONS AM) EXPERIMENTAL DATA : 



There are three sets of observations to be made: 



1. The survival of eggs or embryos allowed to remain in the various media, begin- 

 ning at different stages of development. These data will have significance 

 not only in relation to Isotonicity but also to tolerance, at the various 

 embryonic stages. Duration of observation: one week. 



2. The developmental rate in anl so tonic solutions as compared with the controls. 

 These data will be significant only under the condition that the temperature 

 is very accurately controlled. 



3. The diameter readings, or swelling and shrinking of the eggs and embryos. 

 These readings can be taken directly by placing fine-lined graph paper beneath 

 the finger bowl containing the eggs and determining the relative values against 

 the graph paper. Beadings should be taken along two axes, at right angles to 

 each other, since some eggs may be ovate. The average of the two readings 

 would then be taken as the value for any particular egg. 



This type of observation has relative value only, and can be used on body 

 cavity eggs, aged eggs, and early cleavages up to but not including the gas- 

 trula stage. Gastrulatlon Involves the development of internal cavities which 

 would render these readings invalid. 



If time permits, two additional sets of observations should be made: 



k. The aged uterine egg of the Anur'an shows cortical breakdown, beginning after 

 about 5 days at 10°C., and earlier at the higher temperatiures. Such an egg 

 should react more with increasing age toward anisotonlc media, eventually re- 

 sponding like any non-living osmometer. Readings may be made over rather short 

 intervals of time. 

 5. The tolerance of the extremes in osmotic pressure by the various critical 

 embryonic stages exposed for brief periods can be tested. For Instance, re- 

 cently Inseminated eggs or eggs about to gastrulate might be exposed for a 

 limited time to glass distilled water, or to double Standard Solution, or to 

 both In succession before returning them to the control medium. 



BEFERENCES : 



Andreassi, G., I939 - "Effect of saline solutions on the development of eggs of Ambly- 



stoma." Boll, Soc. Ital. Biol. Sperm. 1U:195. 

 Bachmann, E. L. & J. Euunstrom, I912 - "Der osmotische Druck wahrend der embryonalent- 



wlcklung von Eana temporarla." Pfluger's Arch. f. Ges. Physiol. li*-l4-:287. 

 Eoltfre.ter, J., 1914-3 - "Properties and functions of the surface coat in amphibian embryos." 



Jour. Exp. Zool. 93:251. 

 Krogh, A., E. Schmidt-Nielsen, & E. Zeuther, I938 - "The osmotic behavior of frog's eggs 



and young tadpoles." Zeitschr. f. vergl. Physiol. 26:230. 



