Treatment with estrogenic compounds or vitamin D metabolites and analogs increased the specific activity of creatine kinase (CK) in different organs from intact or ovariectomized female rats. We now assessed the hormonal responsiveness of para-uterine fat from female rats. Injection or feeding of immature female rats with different estrogens resulted in increased CK. Injection with anti-estrogens resulted in tamoxifen stimulated CK, whereas all others were effective only in inhibition of E₂-stimulated CK. E₂ injection at different days of the menstrual cycle resulted in highest increased CK in pro-estrous, lower response in di-estrous and met-estrous and no response at estrous. E₂ induced-CK was age-dependent and increased at different time after ovariectomy. CK induction by E₂ was abolished by inhibitors of both protein and RNA synthesis. Induction of diabetes led to decreased basal CK in para-uterine fat, with no hormonal response. Fat cells responded to the vitamin D metabolites 1,25(OH)₂D₃ (0.5 ng) or 24,25(OH)₂D₃ (5 ng) and to the non-hypercalcemic analogs of 1,25(OH)₂D₃ i.e. hexa-fluoro vitamin D₃ (FL), the side chain modified derivatives: EB 1089 (EB), CB 1093 (CB) and MC 1288 (MC), or the less-calcemic analogs: JK1624F₂-2 (JKF) and QW1624F₂-2 (QW). But these analogs unlike other organs and similar to the uterus, did not up-regulate the response to E₂. In conclusion, rat female derived fat tissues responded to estrogenic compounds and to vitamin D metabolites and analogs, by increased CK and should be considered as a target for both the beneficial and the hazardous effects of hormonal treatments.