292 INVERTEBRATE PHYSIOLOGY 



ventional physiology, this seemed quite unlikely. All of the known physio- 

 logical processes (Belehradek, 1935), e.g., rate of embryonic development, 

 rate of heart beat, metabolic rate, rate of muscle contraction, and even the 

 conscious estimation of time in man (Hoagland, 1933) are accelerated 

 about two to three times for each 10-degree C rise in temperature. If a 

 metabolic clock were present, could it actually have this most unique prop- 

 erty of temperature independence of the frequency of the daily cycles it 

 regulated ? 



Starting with the temperature question, I commenced a series of studies 

 in which I was assisted by a number of my present and former graduate 

 students. Among those who have worked with me upon this problem are 

 H. M. Webb of Goucher College, M. I. Sandeen of Duke University, M. 

 Fingerman of Tulane University, G. C. Stephens of the University of 

 Minnesota, M. F. Bennett of Sweet Briar College, M. N. Hines of Wooster 

 College, W. J. Brett of Millsaps College, and R. O. Freeland, C. L. Ralph, 

 Miss J. Shriner, and R. A. Brown at Northwestern University. 



Initially for this study we wanted an animal which was cold-blooded, 

 i.e., one whose body temperature varied with that of its surroundings, and 

 which had a definite and easily measured daily behavior pattern. The 

 common fiddler crab, which possesses a very striking daily variation in 

 body color, appeared to be an ideal animal for the work. It becomes pale in 

 the early evening as a result of movement of black pigment in each of the 

 numerous highly branched pigment cells of its skin to the centers of the 

 cells. At about daybreak the animals grow dark in color as the pigment 

 commences to disperse into the branches of the pigment cells. They remain 

 dark throughout the day. A quantitative method was developed for deter- 

 mining the exact stage of dispersion of the pigment at any given time of 

 day. By placing a large group of animals in a photographic darkroom at 

 constant temperature and then sampling a group from time to time, it was 

 possible to follow the persisting daily variations in the pigment cells. The 

 rhythmic change was studied continuously for about two months. During 

 this time the rhythm not only did not weaken, but rather became stronger 

 and stronger for about two weeks, and then continued unabated for the re- 

 mainder of the time. 



In the two months of observation, there was no measurable tendency 

 for the rhythm to get out of phase with the outside day-night rhythm ; in 

 other w^ords, the clock could not have gained or lost more than a few 

 minutes in the two-month period. This rhythm continued to remain syn- 

 chronized with the outside day-night cycles, whether the animals were in 

 a darkroom at 26° C, 16° C, or even 6°C (Brown and Webb, 1948). 

 If, however, crabs which were in a photographic darkroom were left six 

 hours in sea water chilled to within a degree or two of freezing, and then 



