Regulation of tyrosine hydroxylase levels and activity and Fos expression during opioid withdrawal in the hypothalamic PVN and medulla oblongata catecholaminergic cell groups innervating the PVN

Morphine withdrawal increases the hypothalamic-pituitary-adrenocortical (HPA) axis activity, which is dependent on an hyperactivity of noradrenergic pathways innervating the hypothalamic paraventricular nucleus (PVN). However, the possible adaptive changes that can occur in these pathways during morphine dependence are not known. We studied the alterations in tyrosine hydroxylase (TH; the rate-limiting enzyme in catecholamines biosynthesis) immunoreactivity levels and TH enzyme activity in the rat NTS-A2/VLM-A1 noradrenergic cell groups and in the PVN during morphine withdrawal. In the same paradigm, we measured Fos expression as a marker of neuronal activation. TH and Fos immunoreactivity was determined by quantitative Western blot analysis, combined with immunostaining for TH and Fos for immunohistochemical identification of active neurons during morphine withdrawal. Dependence on morphine was induced by a 7-day s.c. implantation of morphine pellets. Morphine withdrawal was precipitated on day 8 by an injection of naloxone (5 mg/kg s.c.). Morphine withdrawal induced the expression of Fos in the PVN and NTS/VLM, which indicates an activation of neurons in these nuclei. TH immunoreactivity in the NTS/VLM was increased 90 min after morphine withdrawal, whereas there was a decrease in TH levels in the PVN at the same time point. Following withdrawal, Fos immunoreactivity was present in most of the TH-positive neurons of the A2 and A1 neurons. TH activity was measured in the PVN, a projection area of noradrenergic neurons arising from NTS-A2/VLM-A1. Morphine withdrawal was associated with an increase in the enzyme activity at different time points after naloxone-precipitated morphine withdrawal. The present results suggest that an increase in TH protein levels and TH enzyme activity might contribute to the enhanced noradrenergic activity in the PVN in response to morphine withdrawal.