RESUMO
AIM: To investigate the inhibitory effect s of a synthetic CRE-transcription factor decoy oligodeoxynucleotide (CRE-decoy ODN) on the upregulation of the expression of cholecystokinin (CCK) and fosB mRN A induced by chronic morphine administration in SK-N-SH cells. METHODS: The CRE cis-element, TGACGTCA, was palindromic, a sy nthetic single-stranded phosphorothioate oligodeoxynucleotide composed of the CR E sequence self-hybridizes to form a duplex/hairpin. The CRE-palindromic decoy a nd control oligodeoxynucleotides were added to the medium (1 h before exposure t o morphine) at 150 nmol/L in the presence of cationic lipid DOTAP. After the cel ls were treated with 100 ?mol/L morphine for 48 h, 10 ?mol/L naloxone was use d for 15 min. The effects of CRE-decoy ODN on the DNA-binding activity of CREB, the expression of CCK and fosB mRNA were detected by electrophoresis mobi lity shift assay (EMSA) and RT-PCR, respectively. The stability of cell-incorpo rated [ 32P]-labeled CRE-decoy ODN was extracted with phenol:chloroform a nd then subjected to 20% nondenaturing polyacrylamide gel electrophoresis and au toradiography. RESULTS: Chronic morphine administration and acute naloxone-prec ipitated withdrawal significantly activated the DNA-binding activity of CREB and the expression of CCK and fosB mRNA in SK-N-SH cells. The CRE-decoy ODN pen etrated into the cells, specifically downregulated these indexes. CONCLUSIONS: CRE-decoy ODN can significantly downregulates the e xpre ssion of CCK and fosB mRNA through specifically suppressing the DNA-binding activity of CREB activated by chronic morphine administration in SK-N-SH cells.