The regulation of prodynorphin gene expression in cultured spinal cord cells: involvement of second messengers

Neuropeptides
T S HaH W Suh

Abstract

The regulation of prodynorphin (proDYN) mRNA levels by cAMP and protein kinase C (PKC) pathways was studied in cultured rat spinal cord cells. Spinal cord cells were cultured from 14 day (E 14) embryos of Sprague-Dawley rats. After 7 days in vitro, the spinal cord cells were incubated with either forskolin (5 microM) or phorbol-13-myristate acetate (PMA; 2.5 microM) for 1, 3, 6, 9, 12, or 24 h and the total RNA was isolated for Northern blot analyses. The proDYN mRNA level began to increase 1 h, then reached and remained at a peak 3-6 h after stimulation by forskolin or PMA. proDYN mRNA levels in forskolin treated cells decreased slightly from their peak after 9 h of treatment, whereas the level of proDYN mRNA returned to the basal level in PMA-treated cells. Pretreatment of cells with cycloheximide (a protein synthesis inhibitor; 10 microM) did not affect the forskolin- or PMA-induced increase in proDYN mRNA, but pretreatment with nimodipine (a L-type Ca2+ channel blocker; 2 microM), omega-conotoxin (a N-type Ca2+ channel blocker; 1 microM), or KN-62 (a Ca2+/calmodulin-dependent protein kinase II inhibitor; 5 microM) inhibited induction of proDYN mRNA both by forskolin and PMA. Additionally, dexamethasone did not affect the ex...Continue Reading

References

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