FASTING CONFINEMENT EFFECTS ON SHARKS 611 



experimental group, and tag numbers identified individual animals. Seventy- 

 eight animals were used in this experimental series. All were mature females, 

 49-91 cm in standard length; several were gravid, bearing pups estimated to 

 be between 3 and 15 months in utero. Fewer than 16 animals were held in a 

 pen at any one time. Squalus generally refuse to feed in captivity (Sargent, 

 Gatten, and Mcintosh 1971, 1972; Murdaugh and Robin 1967). Attempts to 

 induce feeding were unsuccessful, and the sharks fasted until they were 

 sacrificed or died in the experimental enclosure. 



In 1973 a record was kept of the water temperature, and water samples 

 were taken at intervals to detect any alterations in salinity. 



Four groups were utilized: 



I. 1971; 20 animals— Blood samples of 1-3 ml were taken at intervals of 

 3 to 8 days from time of capture to time of death. At random intervals, 

 animals were removed and sacrificed for tissue samples. 



II. 1973; 16 animals— Blood samples were taken from each animal at 

 5-day intervals up to the time of death. 



III. 1971, 1973; 11 animals— Sharks were held undisturbed; individuals 

 were randomly sacrificed for tissue and blood samples. 



IV. (1971, 1973; Controls) 



a. Wild population: Animals were caught and immediately sacrificed 

 (within 5 min) for blood and tissue samples (19 animals). 



b. Experimental population: Animals were placed in the pen and left 

 undisturbed until they expired naturally (12 animals). 



Blood samples were drawn from the caudal vein using sterile, disposable 

 syringes with 18- or 20-gauge needles. The volume of blood per sample was 

 approximately 3.5 ml. Hematocrit determinations and colorimetric assays 

 for serum urea, cholesterol, protein, glucose, and total lipid were performed 

 on fresh sera. Aliquots of sera (1-1.5 ml) were frozen for transport to 

 Cornell University; after thawing, these were assayed for serum sodium, 

 potassium, chloride, total osmolarity, urea, iron, and calcium. 



Total serum osmolarity was determined on an Advance Instrument Wide- 

 range Osmometer. Serum sodium and potassium were analyzed by flame 

 photometry, and serum calcium and iron levels were assayed by atomic 

 absorption spectrophotometry. Serum chloride levels were determined using 

 an IL-279 chloride analyzer. Glucose and urea were assayed following the 

 procedure of Hanok (1969) with the aliquot for urea determination diluted 

 1:10. Serum cholesterol was analyzed by a modification of the Lieberman- 

 Burkhardt technique (Huang et al. 1961). Total serum lipids were assayed by 

 the turbidimetric method (Davidsohn and Wells, 1965) and the units con- 

 verted to mg% lipid by the conversion factor used in mammalian serological 

 assays. Total serum protein was determined by the biuret method. 



For blood smears, 0.5 ml of blood were drawn from the caudal vein into a 

 heparinized syringe. Smears were made upon arrival in the laboratory, air- 

 dried, and stored. These were subsequently stained with Wright's stain. When 

 tissue samples were desired, following the removal of a blood sample the 



