Blood samples from 12 Shorthorn heifers, fed a grass silage ration and housed indoors or outdoors, were assayed for 25-hydroxyvitamin D during 9 mo. There was more in plasma during the summer from the outdoor group, reflecting a greater exposure to ultraviolet light and dermal synthesis of vitamin D, the precursor of 25-hydroxyvitamin D. In another experiment, assays were in four groups of six heifers each; 1) control, 2) single oral dose of 1,000,000 IU vitamin D3, 3) injected intramuscularly with 1,000,000 IU of vitamin D3, and 4) free access to a mineral mixture containing 32,000 IU vitamin D3/kg. In all groups, 25-hydroxyvitamin D was higher in plasma in summer than in winter. All heifers given vitamin D had more 25-hydroxyvitamin D in plasma during winter than controls. Animals injected with vitamin D had more 25-hydroxyvitamin D in plasma during part of the winter than those on the other treatments with vitamin D.