In situ hybridization for creatine kinase-B messenger RNA in rat uterus and brain

Creatine kinase-B (CKB) is present in both uterus and brain, and in uterus its synthesis (protein and mRNA) is regulated by estrogen. In the present study we have used in situ hybridization to detect CKB mRNA in uterus and brain, and to determine whether there is cell type specific induction of CKB by estrogen in these tissues. Tissue was taken from ovariectomized (ovx) rats that had been injected with either estrogen (17 beta-estradiol-3-benzoate and/or 17 beta-estradiol) or vehicle alone, 2, 8, 24 and 72 h previously. The brains and uteri were removed, frozen, cryostat-sectioned, and processed for in situ hybridization histochemistry. The uterine and brain sections were incubated with a tritiated DNA probe complementary to a 3' fragment of CKB mRNA, or a control sense probe to the same 3' fragment. In uterine smooth muscle cells, a 2.5- and 3.5-fold induction of CKB mRNA was observed 2 and 24 h after estrogen administration, respectively, and levels approached ovx controls at 72 h. A smaller induction (1.9-fold, 2 h) was observed in uterine epithelium, with little induction of CKB mRNA in stroma. In the brain CKB mRNA was detected in neurons, but not in clearly identified glia, and only occasionally in ependymal cells. In brain regions containing estrogen receptors there was no evidence of a significant estrogen effect on CKB mRNA levels. Some brain regions had higher neuronal expression than others (e.g. medial septum vs. preoptic area), but expression was widespread and not limited to neuroendocrine sites.