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KMID : 1161519990030030311
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Animal Cells and Systems
1999 Volume.3 No. 3 p.311 ~ p.317
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Presence of pituitary specific transcription factor pit-1 in the rat brain: Intracerebroventricular administration of antisense pit-1 oligodeoxynucleotide decreases brain prolactin mRNA level
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Kim Tae-Woo
Kim Hyun-Ju
Lee Byung-Ju
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Abstract
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Prolactin (PRL) was reported to be locally synthesized in many brain areas including the hypothalamus, thalamus (TH) and hippocampus (HIP). In the pituitary lactotrophs, PRL synthesis is dependent upon a pituitary?specific transcription factor, Pit?1. In the present study, we attempted to identify Pit?1 or Pit?1?like protein in brain areas known as the synthetic sites of PRL. Reverse transcription?polymerase chain reaction (RT?PCR) and Northern blot analysis showed the same Pit?1 transcripts in brain areas such as the medial basal hypothalamus (MBH), preoptic area (POA), TH, and HIP with the Pit?1 transcripts in the anterior pituitary (AP). Electrophoretic mobility shift assay (EMSA) was run with nuclear protein extracts from brain tissues using a double strand oligomer probe containing a putative Pit?1 binding domain. Shifted bands were found in EMSA results with nuclear proteins from MBH, POA, TH and HIP. Specific binding of the Pit?1?like protein was further confirmed by competition with an unlabeled cold probe. Antisense Pit?1 oligodeoxynucleotide (Pit?1 ODN), which was designed to bind to the Pit?1 translation initiation site and block Pit?1 biosynthesis, was used to test Pit?1 dependent brain PRL transcription. Two nmol of Pit?1 ODN was introduced into the lateral ventricle of a 60?day old male rat brain. RNA blot hybridization and in situ hybridization indicated a decrease of PRL mRNA signals by the treatment of Pit?1 ODN. Taken together, the present study suggests that Pit?1 may play an important role in the transcriptional regulation of local PRL synthesis in the brain.
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KEYWORD
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Pit-1, Prolactin, Rat brain, Antisense Pit-1
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