Histochemical methods offer the advantage, in the study of vitamin deficiency states, that a considerable amount of information can be obtained from each animal. The amount of tissue required for the demonstration of any one enzyme is small; thus the number of enzymes that can be examined is virtually unlimited.In this study a comparison was made of enzyme activities in tissues of normal and vitamin D-deficient rats. The tissues, other than bone, where lack of vitamin D is considered to have an effect, are those associated with calcium and phosphorus absorption and excretion and those associated with the control of calcium and phosphorus metabolism. The intestine and its associated glands, and the kidney and the parathyroid gland have therefore been selected for investigation. EXPERIMENTAL AnimalsThe rats were of the Lister Institute black and white stock maintained at Queen Elizabeth College. They were weaned at 23 days and taken for experiment at 28-30 days at weights ranging from 57 to 80 g. The histological procedure required pairs of animals for comparison, and limitation of time permitted only two or four animals to be dealt with in a week. Pairs of male rats were therefore taken for experiment at intervals, caged separately and fed on Steenbock's rachitogenic diet 2965 (Steenbock & Black, 1925) with or without a dose of vitamin D. Vitamin D was given as a solution of Adexolin (a concentrate of vitamins A and D ; Glaxo Research Ltd) in arachis oil in a single weekly dose of 10 i.u. vitamin D. The animals without vitamin D received arachis oil to compensate for any effect of the inclusion of the small amount of oil in the diet. The diet was offered as a dry powder and was made once or twice weekly. It consisted of freshly ground whole yellow maize complete with the germ 76%, wheat gluten 20%, calcium carbonate 3 yo, and sodium chloride I %, with a Ca:P ratio of 4:'.At the beginning of the experiment 'line tests' (Bills, Honeywell, Wirick & Nussmeier, 1931) were made on the right radius and ulna of every rat on the day it was killed. The left radius and ulna were put into the deep freeze and examined all at the same time at the end of the experiment. The femur, tibia and fibula from all rats were kept in the deep freeze until the end of the experiment so that all bone ash determinations could be done at the same time. Bone ash was measured by the method of 10-2

I . Rats were given a rachitogenic diet with or without vitamin D for I , 2, 3 or 4 weeks, and vitamin D was administered to a group of rats which had been deprived of vitamin D for 3 weeks. 2. Changes in enzyme activities of some tissues were followed histochemically. In general, changes in parathyroid preceded those in gut and kidney. Leucine aminopeptidase activity rose in the parathyroid, kidney and jejunum of the vitamin D-deficient rat as did the ribonucleic acid content of the parathyroid and the dehydrogenase activity of the gut. In contrast, the phosphatase activity of the gut and kidney fell in the vitamin D-deficient rat. All these changes were reversed during the I-week recovery period. 3. A slower response of bones to both vitamin D deprivation and vitamin D dosage after deprivation was indicated by the changes in the percentages of ash in the bones and the changes in number and distribution of osteoclasts.In a previous study (Cheesman, Copping & Prebble, 1964) a comparison was made of enzyme activities in the tissues of normal and vitamin D-deficient rats after 4, 5 and 7 weeks of deprivation. Among the differences observed was a consistent rise in leucine aminopeptidase activity in the parathyroids of the deficient rat. Since the parathyroid is so intimately concerned with the distribution of calcium and phosphorus, changes in other tissues could be secondary to those of the parathyroid rather than primary effects of the lack of vitamin D.In the present study some rats were deprived of vitamin D for shorter periods and some deficient rats given a curative dose of vitamin D, to determine whether enzyme activities of other tissues altered before those of the parathyroid during either depletion or recovery. EXPERIMENTAL Animals and procedurePairs or trios of male rats, of the Lister Institute black and white stock maintained at Queen Elizabeth College, were placed on a rachitogenic diet with or without vitamin D (Cheesman et al. 1964) at 28 days of age and maintained on the diet for I , 2 or 3 weeks in experimental series C 35 and for 4 weeks in series C 38. In series C 35 one animal of each pair received vitamin D for the whole experimental period and the other received none. In series C 38 one animal of each trio received vitamin D for the whole experimental period, one animal received vitamin D for the last week of the period only and the third received none.In series C 35 one normal and one deficient animal were killed on each occasion https://www.cambridge.org/core/terms. https://doi