Morphine withdrawal regulates phosphorylation of cAMP response element binding protein (CREB) through PKC in the nucleus tractus solitarius-A2 catecholaminergic neurons.

The transcription factor cAMP response element binding protein (CREB) has been implicated in the actions of drugs of abuse in several brain areas. However, little is known about CREB regulation in the nucleus tractus solitarius (NTS)-A(2) catecholaminergic cell group, one of the key regions of the brain stress system. Morphine withdrawal modulates gene expression in the NTS through various second-messenger signal transduction systems including activation of extracellular signal-regulated kinases 1/2 (ERK(1/2)) and protein kinase C (PKC). In the current study we used immunoblotting and immunohistochemistry to investigate changes in CREB phosphorylation in the NTS and kinases that may mediate the morphine withdrawal-triggered activation of CREB and hypothalamo-pituitary-adrenocortical (HPA) axis (another stress system circuit) response after naloxone-induced morphine withdrawal. We found an increased phosphorylation of CREB (pCREB) selectively within tyrosine hydroxylase (TH) immunoreactive neurons in the NTS from morphine-withdrawn rats, which parallel elevated corticosterone levels. We also measured expression levels of TH and phosphorylated ERK(1/2) (pERK(1/2)), and found that both are up-regulated following morphine withdrawal. SL327, an inhibitor of ERK activation, at doses which reduced the hyperactive pERK(1/2) levels, did not attenuated the rise in pCREB and TH immunoreactivity or plasma corticosterone secretion during morphine withdrawal, indicating that ERK kinase/ERK pathway was not directly needed for either activation of CREB and TH expression in the NTS or HPA axis hyperactivity. In contrast, PKC inhibitor calphostin C reduced the withdrawal-triggered rise in pCREB, pERK(1/2), TH expression and corticosterone secretion. The results indicate that PKC mediates both CREB activation and HPA response by morphine withdrawal and might suggest that CREB activation in the NTS is related to TH expression associated with morphine withdrawal.