Calcif. Tissue Int. 33, 277-280 (1981) Calcified Tissue International 9 1981 by Springer-Verlag Growth Failure in Vitamin D-Deficient Rat Pups Robert Brommage* and William F. Neuman Department of Radiation Biologyand Biophysics, University of Rochester Medical Center, Rochester, New York 14642,USA Summary. Vitamin D-deficient rat pups were pro- duced by feeding normal impregnated rats a diet deficient in vitamin D after mating. The rat pups ap- peared normal at birth but stopped growing at 1 week of age. Despite this growth failure, these pups were normocalcemic. Analyses of calvaria from a similar group of dams given vitamin D3 showed that these dams mobilized skeletal calcium to meet the calcium requirements of their growing pups. Key words: Vitamin D deficiency -- Neonatal de- velopment -- Lactation -- Rat pups -- Calcium me- tabolism. Few studies have examined the role of vitamin Da and its metabolites in pregnancy, lactation, and neonatal development in the rat. Raisz [1] was un- successful in his attempts to breed vitamin D- deficient rats, whereas Toverud [2] concluded that vitamin D was not essential for lactation nor for maintaining a normal level of calcium in milk but reported that pup growth was slower after 7 days of age in vitamin D deficiency. Recently, both Gray et al. [3, 4] and Halloran and DeLuca [5] have shown that vitamin D is not necessary for reproduction in the rat. Moreover, although at weaning vitamin D- deficient pups had normal body weights and were both normocalcemic and normophosphatemic, Hal- loran and DeLuca [5] found that these pups had di- minished skeletal calcification. This investigation was undertaken to generate vi- tamin D-deficient rat pups and examine the effects * Present address: Department of Biochemistry, University of Wisconsin, Madison, Wisconsin 53706, USA. Send offprint requests to William F. Neuman at the above address. of vitamin D3 on the development of these pups when their dams were fed a calcium-deficient diet. Materials and Methods Twelve Sprague-Dawley sperm-positive female rats were pur- chased from Holtzman Co. (Madison, Wisconsin, U.S.A.) the day after mating and placed in individual cages in a room free from ultraviolet light. The vitamin D-deficient diets used in this study were purchased from Teklad Test Diets. Until parturition the rats were fed a diet containing 0.2% calcium, 0.4% phos- phorous (TD 76428), and after delivery the rats were given a cal- cium-free, 0.4% phosphorous diet (TD 76524). Vitamin D3 was purchased from ICN Pharmaceuticals, Inc. and dissolved in 95% ethanol before injection. Bone samples were weighed wet, dried overnight at 110~ re- weighed, and extracted overnight in 2N nitric acid. Plasma, serum, and bone extract calcium values were determined with a Calcette titrator. Plasma and serum phosphate, after deprotein- ization with 8% perchloric acid, were assayed by the method of Chen et al. [6]. All data are expressed as means -+ SEM. For statistical analy- ses, Student's t test was used. Results Of the 12 rats, 10 gave birth to apparently normal litters of pups and the litter sizes were evened out such that each dam had eight pups. Four of the eighty pups died within 2 days of birth, but all of the remaining pups survixied the full course of the ex- periment. On the 5th day after delivery, 500 units of vitamin Da were given by intraperitoneal injection to four of the dams. This dose of vitamin Da was chosen to provide an adequate but nontoxic supply of the vitamin throughout lactation. Figure 1 shows the normalized body weight gains of these pups (defined as the mean body weight of the pups on a given day divided by the same litter's mean body weight at 1 day of age). After 1 week the 0171-967X/81/0033-0277 $01.00