beta-endorphin and Neuroblastoma

Cross-linking experiments using the (125)I-beta-endorphin revealed the presence of several receptor-related species in cell lines expressing endogenous opioid receptors, including a small molecular mass protein (approximately 22 kDa). Previous reports have suggested that this 22-kDa (125)I-beta-endorphin cross-linked protein could be the degradative product from a higher molecular mass species, i.e., a fragment of the receptor. To determine if this protein is indeed a degraded receptor fragment, (125)I-beta-endorphin was cross-linked to the (His)(6) epitope-tagged mu-opioid receptor (His-mu) stably expressed in the murine neuroblastoma Neuro(2A) cells. Similar to earlier reports with cell lines expressing endogenous receptors, two major bands of 72- and 25-kDa proteins were specifically cross-linked. Initial cross-linking experiments indicated the absolute requirement of the high-affinity (125)I-beta-endorphin binding to the mu-opioid receptor prior to the appearance of the low molecular weight species, suggesting that the 22-kDa protein could be a degraded fragment of the receptor. However, variations in the ratios of these protein bands being cross-linked by several homo- or heterobifunctional cross-linking agents were observed. Although neither the carboxyl terminus mu-opioid receptor-specific antibodies nor the antibodies against the epitope at the amino terminus of the receptor could recognize the 22-kDa protein, this (125)I-beta-endorphin cross-linked species could be coimmunoprecipitated with the receptor antibodies or could be isolated with a nickel resin affinity chromatography. The direct physical association of the 22-kDa protein with the receptor was demonstrated also by the observation that the 22-kDa protein could not bind to the nickel resin alone, but that its binding to the nickel resin was restored in the presence of the His-mu. Taken together, these results suggest that the 22-kDa protein cross-linked by (125)I-beta-endorphin is not a degradative product, but a protein located within the proximity of the mu-opioid receptor, and that it is tightly associated with the receptor.

Topics: Animals; beta-Endorphin; Cross-Linking Reagents; Enkephalin, Ala(2)-MePhe(4)-Gly(5)-; Histidine; Iodine Radioisotopes; Mice; Molecular Weight; Morphine; Neuroblastoma; Proteins; Receptors, Opioid, mu; Somatostatin; Tumor Cells, Cultured

The purpose of this study was to assess whether xenogeneic tumor cells secreting beta-endorphin and immunologically isolated in polymer capsules could survive and continue to reduce pain when transplanted into the spinal cerebro-spinal fluid (CSF) space of rats. Also, a silicone container for polymer capsules was designed for the clinical application of this method of cell therapy. The mouse tumor cell lines, proopiomelanocortin gene transfected Neuro2A which secrete beta-endorphin, were enclosed in polymer capsules at a density of 5 x 10(6)/mL, and transplanted into the spinal CSF space from the occipito-atlantal junction of male Sprague-Dawley rats. Three analgesiometric tests--the tail pinch test, the hot plate test, and electrical stimulation test--showed that the rats with encapsulated Neuro2A (n = 6) were significantly less sensitive to pain after transplantation than control animals (n = 8). The analgesia induced by the encapsulated cells secreting beta-endorphin was attenuated by the opiate antagonist naloxone. Morphological study revealed that the encapsulated cells survived for 1 mo after transplantation into the CSF space. An in vitro experiment on cultured capsules (3 cm long) with a silicone container (Kaneka Medics Co) showed that the encapsulated Neuro2A (5 x 10(6) mL) could secrete peptides for 1 mo. The results of this study indicate that immunologically isolated xenogeneic tumor cells can secrete opiate in the CSF space, and that a silicone container may help the application of this method to the treatment of cancer pain.

Topics: Adrenocorticotropic Hormone; Analgesia; Analysis of Variance; Animals; beta-Endorphin; Capsules; Cell Line; Cell Transplantation; Electric Stimulation; Hot Temperature; Male; Mice; Neuroblastoma; Pain Management; Physical Stimulation; Rats; Rats, Sprague-Dawley; Recombinant Proteins; Transfection; Transplantation, Heterologous; Tumor Cells, Cultured

The purpose of this study was to assess whether xenogeneic tumor cells immunologically isolated in polymer capsules could survive and continue to reduce pain when transplanted into the cerebrospinal fluid (CSF) of rats. The mouse tumor cell lines AtT-20 and gene-transfected Neuro2A, which secrete beta-endorphin, were enclosed in polymer capsules at a density of 5 x 10(6) cells/ml and transplanted into the spinal CSF space of the occipitoatlantal junction in male Sprague-Dawley rats. The analgesiometric tests (tail pinch, hot plate, and electrical stimulation) showed that the five rats with encapsulated AtT-20 or Neuro2A (eight rats) were significantly less sensitive to pain after transplantation than the eight control animals (analysis of variance; p < 0.05). The analgesia induced by encapsulated cells secreting beta-endorphin could be attenuated by the opiate antagonist naloxone, which suggested the involvement of opiate in mediating this response. Morphological study revealed that the cells in polymer capsules survived 1 month after transplantation in the CSF space. In vitro experiments with cultured capsules showed that both encapsulated AtT-20 and Neuro2A secrete peptide for 1 month. The results of this study suggest that immunologically isolated xenogeneic tumor cells can secrete opiate in the CSF space, and this method may be applied to the treatment of cancer pain.

Topics: Analgesia; Animals; beta-Endorphin; Capsules; Cell Aggregation; Male; Mice; Neoplasm Transplantation; Neuroblastoma; Pain; Pain Measurement; Pituitary Neoplasms; Polymers; Prostheses and Implants; Rats; Rats, Sprague-Dawley; Subarachnoid Space; Tumor Cells, Cultured

Neuroblastoma cell lines have been reported to contain two proopiomelanocortin (POMC) mRNA transcripts. We have now shown by immunocytochemistry and radioimmunoassay (RIA) that a number of neuroectodermally derived cell lines contain immunoreactive beta-endorphin although cell concentrations were not characteristic of any tumour type. To explore further the functional significance of beta-endorphin expression, we analysed neuroblastoma cell lines having intermediate (I), substrate adherent (S) and neuronal (N) phenotypes. No differences in cell beta-endorphin content were detected. However, the expression of POMC mRNA and of immunoreactive beta-endorphin was reduced within a few hours of treatment of these cell lines with retinoic acid. Culture of the cell lines in the presence of beta-endorphin resulted in small but significant increases in growth. Although the POMC gene is in the same chromosomal segment as N-myc, which is normally amplified in neuroblastoma, no corresponding amplification of POMC could be demonstrated. The data suggest that POMC gene products may contribute to the autocrine/paracrine growth of neuroectodermal tumours.

Topics: beta-Endorphin; Cell Line; Clone Cells; Ectoderm; Gene Expression; Glioblastoma; Humans; Immunohistochemistry; Melanoma; Neuroblastoma; Neuroectodermal Tumors, Primitive, Peripheral; Pro-Opiomelanocortin; Radioimmunoassay; RNA, Messenger; Tretinoin; Tumor Cells, Cultured

Human beta-endorphin 1-31 (beta-END) stimulated low-Km GTPase activity in a concentration-dependent and saturable manner in membranes prepared from the delta opioid receptor-containing hybrid cell line NG108-15 and from the mu opioid receptor-enriched human neuroblastoma cell line SK-N-SH. Naloxone and the delta-selective antagonist, ICI 174,864, blocked the stimulation of the GTPase activity produced by beta-END in NG108-15 cell membranes, whereas only naloxone inhibited the beta-END-induced stimulation in SK-N-SH cell membranes, suggesting that beta-END was acting through both mu and delta opioid receptors. Treatment of the cells with Bordetella pertussis toxin before the preparation of membranes blocked the stimulation of low-Km GTPase by beta-END in both cell lines. Activation of NG108-15 and SK-N-SH low-Km GTPase by beta-END was sodium-dependent, and lithium and potassium were poor promoters of this activation. These results demonstrate that beta-END stimulates the interaction of both mu and delta opioid receptors with B. pertussis toxin-sensitive G-proteins in SK-N-SH and NG108-15 cell membranes, respectively.

Topics: beta-Endorphin; Cell Membrane; GTP Phosphohydrolases; GTP-Binding Proteins; Humans; Narcotic Antagonists; Neuroblastoma; Peptide Fragments; Receptors, Opioid, delta; Receptors, Opioid, mu; Tumor Cells, Cultured

The mouse neuroblastoma x rat glioma hybrid NG108-15 was previously shown to express delta opioid receptors. Because neuroblastoma cells display different phenotypes and cloned cell lines are heterogenous, we studied the characteristics and distribution of human 125I-beta-endorphin (125I-beta E) binding sites in cultures of NG108-15 cells with the use of micro-autoradiography and light microscopy. 125I-beta E labeled delta sites in NG108-15 in the presence of the non-opioid blocking peptide, beta-endorphin (6-31) (beta E (6-31)). Silver grains resulting from 125I-beta E binding to the opioid sites occurred in diffuse patches over several cells, with preferential location in dense cell patches. Pretreatment of NG108-15 with the delta agonist DADLE, previously shown to decrease beta E binding to delta sites on intact cells, also reduced silver grain density; however, some cells located in dense cell clusters were resistant to substantial agonist induced loss of labeling. These results suggest that delta opioid binding has a heterogenous cellular distribution in NG108.

Topics: Animals; Autoradiography; beta-Endorphin; Binding Sites; Binding, Competitive; Glioma; Mice; Neuroblastoma; Rats; Receptors, Opioid, delta; Tumor Cells, Cultured

The affinity cross-linking of the delta-opioid receptor in neuroblastoma x glioma NG108-15 cells was undertaken using (3-[125I]iodotyrosyl27)human-beta-endorphin ([125I]beta-endorphin) and disuccinimidyl suberate (DSS) or bis(sulfosuccinimidyl) suberate (BS3) in order to estimate molecular size. Following sodium dodecyl sulfate-polyacrylamide gel electrophoresis (SDS-PAGE) analysis, two radioactive bands were observed. Labeling of a major band of 29 kDa diminished in the presence of unlabeled selective delta-opioid agonist, [D-Pen2,D-Pen5]enkephalin (DPDPE), in a concentration-dependent manner, while labeling of a minor band of 58 kDa was hardly affected. The labeling intensity of the 29 kDa band decreased by addition of guanosine 5'-(3-o-thio)triphosphate (GTP gamma S) or by pretreatment of cells with pertussis toxin. These results, taking the molecular weight of covalently bound beta-endorphin (3.6 kDa) into consideration, suggest that the delta-opioid receptor in NG108-15 cell membrane is a 25 kDa protein which is coupled to pertussis toxin-sensitive guanosine triphosphate-binding proteins (G-proteins).

Topics: beta-Endorphin; Cell Membrane; Chemical Fractionation; Electrophoresis, Polyacrylamide Gel; Glioma; GTP-Binding Proteins; Humans; Molecular Weight; Neuroblastoma; Pertussis Toxin; Receptors, Opioid; Succinimides; Tumor Cells, Cultured; Virulence Factors, Bordetella

We have measured mu and delta opioid receptor sites on intact SK-N-SH and NG108-15 neuroblastoma cells, respectively, in culture. Use of 125I-beta-endorphin (beta E) as a tracer, together with beta E(6-31) to block high-affinity non-opioid binding in both cell lines, permitted the measurement of cell surface mu and delta opioid receptor sites. Labeling was at delta sites in NG108-15 cells and predominantly at mu sites in SK-N-SH cells. Pretreatment with the mu and delta agonist, DADLE, caused a rapid loss of cell surface delta receptor sites in NG108-15 cells, but failed to reduce significantly mu receptor density in SK-N-SH cells.

Topics: Analgesics; Animals; beta-Endorphin; Binding Sites; Enkephalin, Ala(2)-MePhe(4)-Gly(5)-; Enkephalin, D-Penicillamine (2,5)-; Enkephalin, Leucine-2-Alanine; Enkephalins; Humans; Mice; Neuroblastoma; Receptors, Opioid; Receptors, Opioid, delta; Receptors, Opioid, mu; Tumor Cells, Cultured

Topics: 3-Iodobenzylguanidine; Antineoplastic Agents; beta-Endorphin; Child; Child, Preschool; Enkephalin, Methionine; Humans; Iodine Radioisotopes; Iodobenzenes; Neuroblastoma; Pain; Time Factors

Regulation of [125I]beta h-endorphin binding by guanine nucleotides was investigated in membrane preparations from two opioid receptor-containing cell lines: NG108-15, which contains only delta opioid receptors, and SK-N-SH, which contains predominantly mu opioid receptors. In contrast to the binding of the delta-selective agonist [3H][D-penicillamine2,D-penicillamine5]enkephalin to NG108-15 cell membranes, and of the mu-selective agonist [3H][D-Ala2,MePhe4,Gly-ol5]enkephalin to SK-N-SH cell membranes, [125I]beta h-endorphin binding to NG108-15 and SK-N-SH cell membranes was not altered by guanosine triphosphate (GTP) or guanylyl-5'-imidodiphosphate (Gpp(NH)p) in the absence of cations. However, in the presence of NaCl, [125I]beta h-endorphin binding to both cell lines was inhibited by GTP and Gpp(NH)p in a concentration-dependent manner. In SK-N-SH cell membranes, the ability of sodium to promote regulation of [125I]beta h-endorphin binding by GTP was mimicked by the monovalent cations lithium and potassium, but not by the divalent cations magnesium, calcium, or manganese. In NG108-15 cell membranes, only sodium was effective in promoting inhibition of [125I]beta h-endorphin binding by GTP. The effect of GTP or Gpp(NH)p in the presence of sodium was also observed with guanosine diphosphate, but not guanosine monophosphate or any of the non-guanine nucleotides tested. These results indicate that the presence of monovalent cations is required for regulation of [125I]beta h-endorphin binding by guanine nucleotides, and that the specificity of this cation requirement differs between the mu and delta receptor-containing cell lines.

Topics: beta-Endorphin; Cations; Cell Membrane; Enkephalin, Ala(2)-MePhe(4)-Gly(5)-; Enkephalin, D-Penicillamine (2,5)-; Enkephalin, Leucine; Enkephalins; Glioma; Guanine Nucleotides; Guanosine Diphosphate; Guanosine Triphosphate; Guanylyl Imidodiphosphate; Humans; Hybrid Cells; Neuroblastoma; Oligopeptides; Receptors, Opioid; Receptors, Opioid, delta; Receptors, Opioid, mu; Sodium Chloride; Tumor Cells, Cultured

The reaction of human beta-endorphin and biotinyl N-hydroxysuccinimide with or without spacer arm, afforded a series of products that were separated by high performance liquid chromatography (HPLC). Liquid secondary ion mass spectrometry of the biotinylated products and their tryptic digests produced abundant protonated molecular ions (MH+), which specified the number and location of biotinylation. Between 1 and 4 biotinyl residues were incorporated per human beta-endorphin molecule, at Lys-9, -19, -24, -28, and -29, but not at the amino-terminal Tyr-1. Three HPLC fractions were isolated for receptor binding studies with monobiotinylation of Lys-9 (B1 beta and B1X beta; X = C6 spacer arm), Lys-19 (B1 gamma), and a mixture of Lys-24, Lys-28, and Lys-29 derivatives (B1 alpha, BX1 alpha). All derivatives displayed tight binding to avidin, and no dissociation from avidin was detectable over several hours at 0 degrees C for the derivatives (BX1 alpha) tested. IC50 values for binding to mu and delta opioid receptor sites were 3-8 times higher for monobiotinylated derivatives than for the parent human beta-endorphin (IC50,mu = 1.5 nM, IC50,delta = 1.3 nM). Association with avidin decreased opioid receptor affinities for the C6 spacer derivative biotinylated at position Lys-9, which is close to the (1-5) enkephalin receptor region. In contrast, avidin did not affect or even increased apparent affinities to mu and delta sites for derivatives biotinylated at the alpha-helical part of the molecule (Lys-19, -24, -28, and -29). Thus, when bound to avidin, the biotinylated human beta-endorphin derivatives with spacer arm (BX1 alpha), substituted near the carboxyl terminal (Lys-24, -28, and -29), displayed mu binding affinities equal to and delta binding affinities only four times lower than underivatized human beta-endorphin. Biotinylated human beta-endorphins also bound to low affinity nonopioid binding sites on NG-108-15 cells; however, affinities to these sites were considerably reduced when derivatives were bound to avidin. The ability of biotinylated human beta-endorphin to cross-link the mu and delta opioid receptors to avidin allows application of the biotin-avidin system as a molecular probe of the opioid receptor.

Topics: Animals; beta-Endorphin; Binding, Competitive; Biotin; Cell Line; Glioma; Humans; Hybrid Cells; Kinetics; Neuroblastoma; Receptors, Opioid; Structure-Activity Relationship

The properties of [125I]beta h-endorphin-binding sites from rat brain membranes and membranes from the NG108-15 cell line were compared using a monoclonal antibody directed against the opioid receptor and opioid peptides as probes. The binding of [125I]beta h-endorphin to both rat brain and NG108-15 membranes yielded linear Scatchard plots with Kd values of 1.2 nM and 1.5 nM, respectively, and Bmax values of 865 fmol/mg rat brain membrane protein and 1077 fmol/mg NG108-15 membrane protein. A monoclonal antibody, OR-689.2.4, capable of inhibiting mu and delta binding but not kappa binding to rat brain membranes, noncompetitively inhibited the binding of 1 nM [125I]beta h-endorphin to rat brain and NG108-15 membranes with an IC50 value of 405 nM for rat brain membranes and 543 nM for NG108-15 membranes. The monoclonal antibody also inhibited the binding of 3 nM [3H] [D-penicillamine2, D-penicillamine5] enkephalin to NG108-15 membranes with an IC50 value of 370 nM. In addition to blocking the binding of [125I]beta h-endorphin to brain membranes, the antibody also displaced [125I]beta h-endorphin from membranes. Site-specific opioid peptides had large variations in their IC50 values depending on whether they were inhibiting [125I]beta h-endorphin binding to rat brain or the NG108-15 membranes. When the peptides were tested with the monoclonal antibody for their combined ability to inhibit [125I]beta h-endorphin binding to both membrane preparations, the peptides and antibody blocked binding as though they were acting at allosterically coupled sites, not two totally independent sites. These studies suggest that mu-, delta-, and beta-endorphin-binding sites share some sequence homology with the 35,000-dalton protein that the antibody is directed against.

Topics: Animals; Antibodies, Monoclonal; beta-Endorphin; Brain; Enkephalin, Ala(2)-MePhe(4)-Gly(5)-; Enkephalin, D-Penicillamine (2,5)-; Enkephalin, Leucine; Enkephalin, Leucine-2-Alanine; Enkephalins; Glioma; Immunoglobulin M; In Vitro Techniques; Iodine Radioisotopes; Male; Molecular Weight; Neuroblastoma; Rats; Rats, Inbred Strains; Receptors, Opioid; Receptors, Opioid, delta; Receptors, Opioid, mu; Tumor Cells, Cultured

Naltrexone, an opioid antagonist, had an inhibitory effect on the growth of murine S20Y neuroblastoma in BALB/c nude mice. Daily injections of 0.1 mg naltrexone/kg, which invoked a receptor blockade for 6-8 hours/day, resulted in 31-92% delay in latency time prior to tumor expression and a 27-49% increase in mean survival time; the magnitude of antitumor response was governed by tumor burden. Inoculation of neuroblastoma (10(6)-2.5 X 10(4) cells) resulted in measurable tumors in 10-13 days and mean survival times of 30-34 days. Immunoreactive beta-endorphin was detected in tumor tissue (39.7 pg/mg protein). Receptor binding assays revealed specific saturable binding of ligands related to delta- and kappa-binding sites, but not for the mu-binding site. These results demonstrate that opioid antagonist modulation of neuro-oncogenesis is not dependent on the integrity of T-cell-mediated immunity and suggest the feasibility of utilizing the nude mouse model in exploring the role of endogenous opioids in human cancers.

Topics: Animals; beta-Endorphin; Disease Models, Animal; Endorphins; Graft Survival; Immunity, Cellular; Male; Mice; Mice, Inbred BALB C; Mice, Nude; Naltrexone; Neoplasm Transplantation; Neuroblastoma; Receptors, Opioid; Receptors, Opioid, delta; Receptors, Opioid, kappa; Receptors, Opioid, mu; T-Lymphocytes; Time Factors

The putative delta-opiate receptor complex has been identified by a new approach which employed an antibody that is directed against a peptide which binds gamma-endorphin and is specified by RNA that is complementary to that of gamma-endorphin mRNA. This antibody competes with beta-endorphin and naloxone for binding sites on the surface of neuroblastoma X glioma NG108-15 hybrid cells. The opiate receptor complex has an apparent molecular weight of 210,000 and is composed of four noncovalently associated subunits with apparent molecular weights of 68,000, 58,000, 45,000 and 30,000.

Topics: Antibodies; beta-Endorphin; Binding, Competitive; Cell Line; Endorphins; gamma-Endorphin; Glioma; Hybrid Cells; Immunologic Techniques; Molecular Weight; Naloxone; Neuroblastoma; Peptides; Receptors, Opioid; Receptors, Opioid, delta; RNA, Messenger

A photoaffinity reagent 2-nitro-4-azidophenylsulfenyl chloride (2,4-NAPS-Cl) and synthetic analogs of human beta-endorphin (beta h-EP) were employed to demonstrate the presence of receptor sites specific for beta h-EP but of non-opioid character in a human neuroblastoma cell line (IMR-32). The radioactive photoaffinity probe was carried out using [125I-Tyr1,2,4-NAPS-Trp27]-beta h-EP and IMR-32 cell membranes. After solubilization with sodium dodecyl sulfate (SDS) and SDS polyacrylamide gel electrophoresis, a single labelled protein band was identified with a molecular weight of 72,000. Labelling was blocked by beta h-EP or beta h-EP-(6-31) but remained in the presence of beta h-EP-(1-27). The specificity of this band is thus identical to that of the non-opioid site previously characterized. Various nonionic or zwitterionic detergents did not extract the labelled non-opioid site.

Topics: Affinity Labels; Azides; beta-Endorphin; Binding Sites; Cell Line; Endorphins; Humans; Molecular Weight; Neuroblastoma; Receptors, Opioid

Acid extracts from mouse Neuroblastoma x rat Glioma hybrid cells have been purified by means of Sep-Pak C-18 and fractionated by high performance liquid chromatography. Each fraction has been submitted to a sensitive beta-endorphin radioimmunoassay and an immunoreactivity peak at camel beta-endorphin retention time was found.

Topics: Animals; beta-Endorphin; Cell Line; Chromatography, High Pressure Liquid; Endorphins; Glioma; Hybrid Cells; Mice; Neuroblastoma; Radioimmunoassay; Rats; Spectrophotometry, Ultraviolet

Inhibition of binding of beta h-endorphin or Leu-enkephalin by beta h-endorphin analogs of various chain lengths in memmbrane preparations of the neuroblastoma x glioma NG108-15 cells has been investigated. The removal of even a single residue from the C-terminus results in the inability of the resulting peptide to completely displace beta h-endorphin. In addition, the proportion of nondisplaceable binding increases with decreasing chain length.

Topics: Animals; beta-Endorphin; Cell Membrane; Endorphins; Enkephalin, Leucine; Glioma; In Vitro Techniques; Kinetics; Neuroblastoma; Rats; Receptors, Opioid; Structure-Activity Relationship

This communication reports for the first time specific binding sites in human neuroblastoma cells for human beta-endorphin (beta h-EP). Three cell lines (IMR-32, NMB and Kelly) were investigated and found to bind tritiated beta h-EP with an apparent dissociation constant of 2.2-4.2 nM. Further characterization with camel beta-EP and synthetic analogs indicated that the binding is most likely mediated by the COOH-terminal segments. beta h-EP-(6-31) had significant potency (15-75%) and beta h-EP-(1-27) was without displacing activity. The camel beta-EP has below 1% of the human beta-EP activity.

Topics: beta-Endorphin; Binding, Competitive; Cell Line; Computers; Endorphins; Humans; Neuroblastoma; Peptide Fragments; Receptors, Opioid

Human beta-endorphin (beta h-EP) binding on neuroblastoma X glioma hybrid NG108-15 cells using tritiated human beta endorphin (3H-beta h-EP) as a primary ligand was found to have a component which was not displacable with [D-Ser2 )-Leu-enkephalin-Thr6 (DSLET). The beta h-EP binding on these cells after saturation of the delta opiate sites with 200 nM DSLET was further characterized with synthetic beta h-EP analogs. The nonopioid binding site appears to recognize beta h-EP-(6-31), beta h-EP-(21-31) and beta h-EP-(28-31). Under these conditions, these COOH-terminal segments fully displace the tritiated beta h-EP. However, beta h-EP-(1-27) does not further displace 3H-beta h-EP in the presence of DSLET. The fact that a combination of DSLET and beta h-EP-(6-31) results in a full displacement of 3H-beta h-EP provides direct evidence for the existence of two binding sites for beta h-EP in NG108-15 cells, one recognizing the NH2-terminal enkephalin sequence and the other the non-opioid COOH-terminal segment.

Topics: beta-Endorphin; Binding, Competitive; Cell Line; Endorphins; Glioma; Humans; Hybrid Cells; Neuroblastoma; Neuroglia; Neurons; Receptors, Opioid; Tritium

The potency of a series of synthetic analogs of beta-endorphin in inhibiting binding of [3H2-Tyr27]-beta h-endorphin to either rat brain membranes or neuroblastoma x glioma hybrid cells (NG108-15) has been determined and compared with the previously determined analgesic potency. There is a very good correlation between inhibitory potency in membranes and cells, but the correlation between analgesic potency and inhibitory potency in either membranes or cells is not as good.

Topics: Animals; beta-Endorphin; Brain; Cell Membrane; Endorphins; Glioma; Hybrid Cells; Kinetics; Mice; Neuroblastoma; Rats; Receptors, Opioid; Structure-Activity Relationship

Topics: Amino Acid Sequence; Animals; beta-Endorphin; Cell Line; Endorphins; Female; Glioma; Humans; Hybrid Cells; Male; Mice; Neuroblastoma; Pregnancy; Rats; Receptors, Opioid; Tissue Distribution

Topics: Amino Acid Sequence; Animals; Behavior, Animal; beta-Endorphin; Cell Line; Endorphins; Glioma; Humans; Hybrid Cells; Mice; Neuroblastoma; Rats; Receptors, Opioid; Species Specificity

The mouse neuroblastoma N18TG2 has specific, saturable binding sites for human beta-endorphin (beta h-EP). The affinity and number of sites are 1.1 nM and 280,000 per cell, respectively, beta h-EP binding is not inhibited by [Leu]enkephalin or morphine at concentrations up to 0.1 mM; beta h-EP-(6--31) is a potent inhibitor of binding, and camel beta-EP is much less potent. The data suggest the importance of the nonenkephalin segment of the beta h-EP molecule for interaction with the binding site in N18TG2 cells.

Topics: Animals; beta-Endorphin; Binding, Competitive; Cell Line; Endorphins; Humans; Kinetics; Mice; Neoplasms, Experimental; Neuroblastoma; Receptors, Opioid