ER calcium discharge stimulates GDNF gene expression through MAPK-dependent and -independent pathways in rat C6 glioblastoma cells

Glial cell line-derived neurotrophic factor (GDNF), a neurotrophic and differentiation factor, is expressed under several pathophysiological conditions but its regulatory signals have not yet been clarified. Here, we found that endoplasmic reticulum (ER) Ca2+ discharge by thapsigargin induced GDNF mRNA as well as COX2 and GRP78 expression in rat C6 glioblastoma cells. GDNF mRNA was immediately induced and peaked at 2 h by thapsigargin, and the alternative transcript consisting of exon 3 and exon 4 appeared to be most inducible. In spite of intracellular Ca2+ perturbation, Ca2+-dependent PKC was not responsible for this induction. Instead, a PKCδ-specific inhibitor, rottlerin, suppressed the thapsigargin-induced GDNF mRNA expression. On the other hand, thapsigargin transiently enhanced phosphorylation status of mitogen-activated protein kinase (MAPK) pathway, including extracellular signal-regulated kinase (Erk), p38 MAPK and c-JUN amino-terminal kinase1 (JNK1) simultaneously; whereas specific inhibitors against MEK1 and JNK only reduced the thapsigargin-induced GDNF mRNA expression. In addition, a pan-PKC inhibitor (Ro-31-8220) attenuated the thapsigargin-enhanced phosphorylation levels of Erk1/2 and JNK1, whereas rottlerin did not. Thus, the present study demonstrated that the thapsigargin-stimulated ER Ca2+ discharge up-regulated GDNF gene expression through both MAPK-dependent and -independent pathways in C6 glioblastoma cells.