Effect of opioid antagonism on β-endorphin processing and proopiomelanocortin-peptide release in the hypothalamus

Abstract

Previous studies have shown that chronic opioid receptor blockade has significant effects on POMC gene expression and peptide levels in the hypothalamus. We have now examined the effects of the opioid antagonist naltrexone on β-EP processing in the hypothalamus and on the release of 2 POMC-derived peptides, β-EP andγ 3-MSH, from the perifused hypothalamus in vitro. The β-EP immunoactivity in the medial basal hypothalamus (MBH) of 7 rats infused for 1 week with naltrexone by osmotic minipump, was individually analyzed by HPLC and compared to 7 control rats. The mean ratio of β-EP 1–31 compared to β-EP 1–27 plus β-EP 1–26 was 2.34 ± 0.41 in the naltrexone treated rats, significantly higher than the ratio of 1.26 ± 0.09 in the control rats ( P < 0.02). Thus in the setting of chronic opioid antagonism although β-EP content decreases, there is relatively more β-EP 1–31, the biologically active opioid form of the peptide, compared to the C-terminally cleaved forms of β-EP which have reduced biological activity. To study the effects of naltrexone on β-EP andγ 3-MSH release, hypothalami were perfused in vitro with 10 −6M naltrexone. Basal release ofγ 3-MSH was significantly higher from the naltrexone treated brains compared to the controls (221 ± 20pg/60min vs.161 ± 6.7pg/60 min) ( P < 0.01); KCl stimulatedγ 3-MSH was also significantly higher in the naltrexone group (951 ± 94 vs.543 ± 85pg/60 min) ( P < 0.005). Basal release of β-EP was136 ± 45pg/60 min in the naltrexone treated brains compared to 93 ± 15pg in the controls, but this difference was not significant; KCl stimulated release of β-EP, however was significantly higher in the naltrexone group (558 ± 103 vs. 275 ± 49pg/60 min ( P < 0.02). To study the acute and chronic effects of naltrexone in vivo on β-EP andγ 3-MSH release, rats were either injected with naltrexone and sacrificed 40–60 min later or were infused with naltrexone for 7 days. Baselineγ 3-MSH release was significantly higher in rats treated with naltrexone 40–60 min prior to the perifusion ( P < 0.01). Baseline β-EP release was below the limit of assay detection. No differences were noted in the responses ofγ 3-MSH or β-EP to KCl in either group. In contrast after chronic treatment with naltrexone for 1 week, baseline peptide release was not different from the control animals despite a more than 50% fall in peptide content. Theγ 3-MSH and β-EP responses to KCl stimulation, however, were significantly less in the naltrexone treated animals. Thus there is an increase in POMC peptide release acutely after treatment with naltrexone in vitro and in vivo. After 1 week of naltrexone, baseline POMC peptide release continues unchanged despite the fall in peptide content, however, the response to KCl is blunted possibly reflecting the decrease in peptide content after chronic stimulation with naltrexone. We conclude that naltrexone has significant effects on POMC peptide release and on β-EP processing in the hypothalamus. These results further demonstrate that the brain POMC system can respond to opioid blockade at several levels and are consistent with inhibitory feedback mechanisms for the autoregulation of the POMC system by endogenous β-EP.