beta-funaltrexamine and Neuroblastoma

The potential interactions of natively expressed mu-opioid and opioid receptor-like (ORL1) receptors were studied by exposing intact BE(2)-C cells to agonists or antagonists for 1 h. Pretreatment with the mu-opioid receptor agonist, [D-Ala(2), N-Me-Phe(4),Gly(5)-ol]enkephalin (DAMGO), or the ORL1 receptor agonist, orphanin FQ/nociceptin desensitized both mu-opioid and ORL1 receptor responses. beta-Funaltrexamine (beta-FNA) pretreatment also blocked both mu-opioid and ORL1 receptor responses, but only mu-opioid receptor binding was reduced. Moreover, beta-FNA (1 microM) failed to inhibit specific ORL1 receptor binding.

Topics: Brain Neoplasms; Cyclic AMP; Enkephalin, Ala(2)-MePhe(4)-Gly(5)-; Humans; Morphine; Naltrexone; Narcotic Antagonists; Narcotics; Neuroblastoma; Nociceptin; Nociceptin Receptor; Opioid Peptides; Receptors, Opioid; Receptors, Opioid, mu; Tumor Cells, Cultured

The irreversible opioid receptor antagonists naloxonazine and beta-funaltrexamine have been used to determine whether multiple mu-opioid receptors exist on undifferentiated SH-SY5Y human neuroblastoma cells. Naloxonazine binds irreversibly to the mu 1-opioid receptor subtype and reversibly to the mu 2-opioid receptor subtype. On SH-SY5Y cells naloxonazine afforded a Ki of 3.4 +/- 0.7 nM, and was fully reversible, indicating the mu-opioid receptor population on SH-SY5Y cells was solely of the mu 2-opioid receptor subtype. The alkylating agent beta-funaltrexamine was maximally able to alkylate only 60% of the mu-opioid receptor sites on SH-SY5Y cells, labelled with [3H]diprenorphine or [3H][D-Ala2,MePhe4,Gly(ol)5]enkephalin (DAMGO). The reversible binding of naloxonazine and the insensitivity of a percentage of the mu-opioid receptor sites to alkylation by beta-funaltrexamine suggests that differences do exist in the mu 2-opioid receptor population on undifferentiated SH-SY5Y cells. This may indicate further heterogeneity or the inability of beta-funaltrexamine to alkylate all relevant nucleophilic groups in a single population of receptors.

Topics: Amino Acid Sequence; Animals; Brain Neoplasms; Cerebral Cortex; Diprenorphine; Enkephalin, Ala(2)-MePhe(4)-Gly(5)-; Enkephalins; Humans; Ligands; Molecular Sequence Data; Naloxone; Naltrexone; Neuroblastoma; Rats; Receptors, Opioid, mu; Somatostatin; Tumor Cells, Cultured

Topics: Analgesics; Cell Line; Enkephalin, Ala(2)-MePhe(4)-Gly(5)-; Enkephalins; Humans; Kinetics; Naloxone; Naltrexone; Narcotic Antagonists; Neuroblastoma; Receptors, Opioid, mu; Tumor Cells, Cultured

Human neuroblastoma SK-N-SH cells, which contain both mu- and delta-opioid receptors, were grown under conditions that provided a mu:delta ratio of 1.5:1. Both receptors were down-regulated after 72 hr of exposure to 100 nM etorphine. Selective down-regulation was demonstrated using selective opioid agonists; the mu agonist Tyr-D-Ala2-Gly-(Me)Phe4-Gly-ol down-regulated mu- but not delta-opioid receptors, whereas prolonged exposure to the selective delta agonist D-Pen2,D-Pen5-enkephalin resulted in delta- but not mu-opioid receptor down-regulation. Morphine, which binds mu- as well as delta-opioid receptors, down-regulated both receptor subtypes. NG108-15 cells, which contain delta receptors exclusively, were also tested. NG108-15 cells did not exhibit delta-opioid receptor down-regulation when exposed to morphine. The discrepancy between the effect of chronic morphine treatment on delta receptors in SK-N-SH cells and in NG108-15 cells raised the question of whether the coexistence of mu receptors in the former allowed morphine to down-regulate delta receptors. The role of mu-opioid receptors in morphine-induced delta receptor down-regulation was studied by using the irreversible mu antagonist beta-funaltrexamine. Pretreatment of SK-N-SH cells with beta-funaltrexamine prevented down-regulation of delta receptors in response to chronic exposure to morphine but did not affect down-regulation of delta receptors in response to D-Pen2,D-Pen5-enkephalin. The experimental data indicate that morphine-induced delta-opioid receptor down-regulation is dependent on the presence of functional mu receptors in the same cell.

Topics: 3,4-Dichloro-N-methyl-N-(2-(1-pyrrolidinyl)-cyclohexyl)-benzeneacetamide, (trans)-Isomer; Analgesics; Down-Regulation; Enkephalin, Ala(2)-MePhe(4)-Gly(5)-; Enkephalin, D-Penicillamine (2,5)-; Enkephalins; Humans; Morphine; Naltrexone; Neuroblastoma; Pyrrolidines; Receptors, Opioid, delta; Receptors, Opioid, mu; Tumor Cells, Cultured

The effects of beta-FNA, a highly selective and irreversible mu opioid receptor antagonist, in altering tumor response in A/Jax mice inoculated with S20Y cells were determined. Inoculation of neuroblastoma cells in control subjects resulted in 100% tumor incidence within 16 days, and mean and median survival times of 36 and 35 days, respectively, following tumor inoculation. Tumor incidence and survival times were comparable to controls for mice given chronic injections of 2 mg/kg and 10 mg/kg beta-FNA every 48 h beginning 2 days after tumor inoculation. Tumor growth was subnormal in the 10 mg/kg beta-FNA group. Both dosages of beta-FNA were found to block morphine-induced analgesia for 48 h. These results suggest that, in and by themselves, mu receptors selectively antagonized by beta-FNA do not play an important role in neuro-oncogenic events.

Topics: Animals; Male; Mice; Mice, Inbred A; Morphine; Naltrexone; Narcotic Antagonists; Neoplasm Transplantation; Neuroblastoma; Reaction Time