Novel opioid peptides derived from mitochondrial cytochrome b: cytochrophins.
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Animals↗
Biomedical subjects
C Gramsch
Publications and source records attributed to C Gramsch.
beta-Endorphin: surface binding and internalization in thymoma cells.
The opioid peptide beta-endorphin binds to specific nonopioid binding sites (Mr 72,000) that are present on the surface of thymoma cells. beta-Endorphin is then internalized, apparently via the Mr 72,000 species, and is subsequently found within intracellular, vesicular structures. This process is accompanied by the down-regulation of the Mr 72,000 binding sites. Our findings suggest that beta-endorphin may modulate cellular functions, such as T-lymphocyte proliferation, at intracellular rather than cell surface sites.
Animals↗
Multiple states of opioid receptors may modulate adenylate cyclase in intact neuroblastoma X glioma hybrid cells.
Opioid receptor binding and opioid-mediated inhibition of cAMP accumulation were studied simultaneously in intact NG108-15 cells. The dose-response curves for the biological response were suggestive of positive cooperativity and systematically occurred at lower ligand concentrations than those for the binding of [3H] [D-Ala2, D-Leu5]enkephalin (DADLE), which were instead shallow and suggestive of a site heterogeneity or of a cooperative phenomenon. Computer modeling of the binding isotherms revealed that the data are best described assuming two binding sites with different affinities for the agonist; the mean ratio between the DADLE concentrations yielding half-maximal occupancy of the high affinity site and half-maximal response was 1.5, but it was 36 when the fractional occupancy of the sum of the two sites was considered. On examining several opioids, no direct correlation was found between high affinity site and biological response; however, several agonists displayed different affinities for the two sites, while the antagonist naloxone and the partial agonist diprenorphine bound to them with identical affinities. Furthermore, naloxone exhibited a good agreement between half-maximal receptor occupancy and Ki in blocking the agonist response. Thus, the binding heterogeneity detectable in intact cells is agonist-specific, and suggests rather that the sites are states of an identical receptor population. When [3H]diprenorphine was used to label the opioid receptors, the competition curves for DADLE were consistent with the existence of an additional, very low affinity state undetectable by direct binding with labeled agonist and, again, not discriminated by naloxone. Multiple affinity states of the opioid receptor in intact cells may reflect its interaction with the effector system in the plasma membrane.
Adenylyl Cyclases↗
Differential effects of various opioid peptides on vasopressin and oxytocin release from the rat pituitary in vitro.
Dynorphin (1-17), and to a lesser extent, beta-endorphin and [Leu]enkephalin (10(-6) M each) decreased the spontaneous release of vasopressin (VP) from the rat neurointermediate pituitary in vitro, whereas the oxytocin (OT) release remained unchanged. Naloxone, however, did not significantly alter the spontaneous VP and OT release. Dynorphin (1-17) (10(-7) M) increased the electrically evoked release of VP and OT, while 10(-6) M had a significant, somewhat less pronounced stimulatory effect only on VP, but not on OT release. The opiate inactive fragment [des-Tyr1]dynorphin (1-17) did not change the evoked VP and OT release, indicating that the dynorphin effect was mediated by opiate receptors. beta-Endorphin (10(-6) M and 10(-7) M) did not alter the evoked VP and OT secretion. 10(-6) M [Leu]enkephalin induced a stimulation of the evoked OT, but not VP release; 10(-7) M [Leu]enkephalin had no effect, neither on VP nor on OT release. The opiate antagonist naloxone (10(-5) M) induced an increase in the evoked VP and, even more pronounced, OT release. In a concentration of 10(-6) M, however, naloxone only increased the evoked OT release. When naloxone and dynorphin (1-17) were concomitantly applied, their stimulatory effects on the evoked VP and OT release were additive. Similarly to the effects of naloxone, addition of a monoclonal antibody which binds to the common N-terminal sequence of all endogenous opioid peptides, resulted in a marked increase in the evoked secretion of VP and, to an even more pronounced degree, of OT.(ABSTRACT TRUNCATED AT 250 WORDS)
Animals↗
The effect of neuroleptic treatment and of high dosage diazepam therapy on beta-endorphin immunoreactivity in plasma of schizophrenic patients.
In 14 schizophrenic patients, treated with neuroleptic drugs, and in 7 patients, treated with high-dosage diazepam, beta-endorphin-like immunoreactivity in plasma has been measured by use of a highly sensitive and relatively specific radioimmunoassay. Neuroleptic treatment induced a significant increase of beta-endorphin-like immunoreactivity (beta-ELI). The pharmacological and clinical implications of this finding are discussed. High-dosage diazepam treatment induces a slight reduction of plasma beta-ELI, a finding which is attributed to antistress effect of diazepam.
Adolescent↗
Anti-idiotypic opioid receptor antibodies.
Anti-idiotypic antibodies were raised against the monoclonal antibody 3-E7. One of these anti-idiotypic antibodies interferes with the binding of 3H-diprenorphine at solubilized opioid receptors, and exhibits opioid agonistic activity on NG108CC15 hybrid cells.
Animals↗
Characterization and localization of immunoreactive dynorphin, alpha-neo-endorphin, Met-enkephalin and substance P in human spinal cord.
By use of specific antisera, the distributions of immunoreactive dynorphin (ir-DYN), alpha-neo-endorphin (ir-alpha-NEO), Met-enkephalin (ir-MET) and substance P (ir-SP) were evaluated in discrete regions of human spinal cord and spinal ganglia. The relative concentrations of immunoreactive peptides in particular regions were as follows: sacral greater than lumbar greater than cervical greater than thoracic. Concentrations of ir-DYN, ir-alpha-NEO and ir-SP were 2-10-fold, but of ir-MET 1-2-fold, higher in the dorsal as compared to the ventral parts of cervical, lumbar and sacral cord. The concentrations of all peptides (when examined in discrete areas of thoracic cord) were found to be highest in the substantia gelatinosa. All peptides were present in the gray matter but only ir-MET was found in white matter. Gel-permeation chromatography of dorsal sacral spinal cord extracts revealed two major ir-DYN peaks. The smaller molecular weight peak, eluted at the position of synthetic dynorphin1-17. ir-alpha-NEO and ir-SP comigrated exactly with their respective synthetic marker peptides. Substantial amounts of ir-SP and also, as confirmed by high pressure liquid chromatography, ir-MET, were found in the dorsal and ventral roots and spinal ganglia, and very low concentrations of ir-DYN or ir-alpha-NEO were also detected in these tissue. These results suggest that dynorphin and alpha-neo-endorphin, in addition to enkephalins, may be involved in transmission of somatosensory information in the human spinal cord.
Chromatography, Gel↗
The opioid peptide dynorphin, circadian rhythms, and starvation.
Dynorphin, an opioid peptide whose functions are unknown, is found in brain, pituitary, and peripheral organs. Specific radioimmunoassays were used to measure dynorphin in the hypothalamus and pituitary, during the day and at night, as a function of food and water deprivation. Immunoreactive dynorphin was increased in the hypothalamus and decreased in the pituitary at night. Water deprivation led to more than 50 percent reduction in daytime levels of pituitary dynorphin and concomitant increases in hypothalamic dynorphin.
Animals↗
Opioid peptide-like immunoreactivity localized in GABAErgic neurons of rat neostriatum and central amygdaloid nucleus.
Opioid peptide-like (OPL)-immunoreactivity and (the GABA-biosynthetic enzyme) glutamic acid decarboxylase-like (GAD)-immunoreactivity were localized in rat neostriatum and central amygdaloid nucleus (ACE) using a polyclonal sheep antiserum to rat brain GAD and a monoclonal mouse antibody to the N-terminus of beta-endorphin (3-E7) as primary antisera. PAP-immunohistochemistry revealed GAD-immunoreactivity in the majority of neurons in neostriatum and ACE. OPL-immunoreactivity was observed in numerous neurons in ACE, but only in few neostriatal nerve cells. In double immunofluorescence in the same section OPL- and GAD-immunoreactivity colocalized in few medium size cells in the neostriatum, but in numerous neurons in ACE. The existence of opioid peptide containing GABAergic neurons in ACE and neostriatum is demonstrated.
Amygdala↗
Uncoupling of receptors is essential for opiate-induced desensitization (tolerance) in neuroblastoma x glioma hybrid cells NG 108-15.
Neuroblastoma x glioma hybrid cells NG 108-15 were chronically treated with opioid agonists and tested for their development of "tolerance" as evaluated by the loss of inhibitory activity of opioids upon cAMP-accumulation. Desensitization was dose-dependent and non-competitive and highly selective, since the activity of other, non-opioid inhibitory compounds, such as noradrenaline and carbachol, was not altered. Moreover, naloxone-induced "withdrawal signs" (revealed in the lack of change in cAMP) were lacking in preparations completely desensitized ("tolerant") towards opiate effects. These results reject the assumption of a common biochemical mechanism underlying both opiate tolerance and dependence and rather support the significance of an uncoupling of receptors from subsequent effector systems upon chronic opiate action.
Animals↗
Activation of hypothalamic beta-endorphin pools by reward induced by highly palatable food.
Experiments were performed to find biochemical evidence of an activation of endogenous opiate peptides in the brain by incentive reward. A method used to estimate specific in vivo opiate binding in rats using the labelled opiate agonist, 3H-etorphine, indicated a considerable reduction in opiate binding exclusively in the hypothalamus of non-deprived animals given a highly palatable food to eat for 20 min. Radioimmunoassay of the hypothalamus of rats under similar conditions found a pronounced drop in the concentration of beta-endorphin, but not in dynorphin, in the hypothalamus, indicating a release and breakdown of beta-endorphin. Therefore, the reduction in opiate binding in the hypothalamus may at least be partially explained by an occupation of opiate receptors by beta-endorphin, causing a reduced availability of receptors to etorphine. A possible role of hypothalamic beta-endorphin in the facilitation of reward pathways in the brain is discussed.
Animals↗
Multiple endorphins in neuronal hybrid cell lines.
Neuroblastoma x glioma hybrid cells (NG 108CC15) and tumors derived thereof were examined for dynorphin- and alpha-neoendorphin-like material. The techniques employed for analyses of opiate-like material were the isolated mouse vas deferens bioassay and gel chromatography and high pressure liquid chromatography in combination with radioimmunoassays. Dynorphin- and alpha-neoendorphin-like material was detected in both the hybrid cells and the corresponding tumors. Immunoreactive dynorphin and alpha-neoendorphin was also in NCB 20 hybrid cells and in tumors thereof assayed. In all samples investigated, the amounts of alpha-neoendorphin-like material was higher than that of dynorphin-like material. The results revealed considerable variability in the amount of dynorphin- and alpha-neoendorphin-activity between particular samples, suggesting the need for studies into the responsible mechanisms.
Animals↗
Heroin addiction: beta-endorphin immunoreactivity in plasma increases during withdrawal.
Seven patients with heroin addiction were hospitalized and immediately withdrawn from opiates. Abstinence symptomatology was evaluated quantitatively by use of the Himmelsbach Score. Maximal intensity of withdrawal symptomatology was reached within 2 days. beta-Endorphin immunoreactivity in plasma was measured by use of a very sensitive radioimmunoassay with a low cross-reactivity (28%) against beta-LPH. A statistically significant increase of mean plasma beta-endorphin-like immunoreactivity during withdrawal could be demonstrated.
Adult↗
Monoclonal antibody to the message sequence Tyr-Gly-Gly-Phe of opioid peptides exhibits the specificity requirements of mammalian opioid receptors.
Six myeloma cell hybrids producing antibodies to human beta-endorphin were isolated from a single mouse spleen. The monoclonal antibodies displayed different binding patterns with the antigen. We report the characterization of one antibody which recognizes the tetrapeptide Tyr-Gly-Gly-Phe representing the message sequence found at the NH2 terminus of all naturally occurring mammalian opioid peptides. Competition experiments in radioimmunoassay and immunohistochemistry show that the antibody fails to bind the beta-endorphin precursor beta-lipotrophin, does not discriminate among opioid peptides that share the same message sequence but have different COOH-terminal extensions, and does not react with pharmacologically inactive derivatives of beta-endorphin. The antibody recognition of the message sequence of natural opioid peptides is sensitive to those molecular changes that affect their receptor binding competence.
Amino Acid Sequence↗
Binding characteristics of a monoclonal beta-endorphin antibody recognizing the N-terminus of opioid peptides.
The present paper describes the isolation and characterization of a clone of hybrid myelomas (3-E7) secreting a mouse monoclonal antibody to beta-endorphin. An examination of its specificity against a series of human beta-lipotropin fragments and other opioid peptides revealed that the N-terminus portion of beta-endorphin is the determinant. Complete or almost complete cross-reactivity was obtained to methionine- and leucine-enkephalin, beta-lipotropin 60-65, and BAM 22; partial cross-reactivity was seen to dynorphin1-13 and alpha-neo-endorphin, whereas beta-lipotropin, alpha-N-acetyl-beta-endorphin, Des-Tyr1-beta-endorphin, in addition to a series of synthetic enkephalin derivatives, completely lacked cross-reactivity. The use of the monoclonal antibody in radioimmunoassay (RIA) for beta-endorphin resulted in a lower sensitivity related to respective polyclonal antibodies. An increase of 100% in tracer binding could, however, be obtained by use of beta-endorphin iodinated with its N-terminal tyrosine protected by coupling to an antibody. A solid-phase RIA was developed involving the internally 3H-labeled monoclonal antibody, which resulted in a 10-fold increase in sensitivity as compared with the homogenous RIA. These data indicate that for the binding to this antibody a tyrosine residue in position 61 is essential, and it thus recognizes a site that is of functional significance for many naturally occurring opioid peptides.
Amino Acid Sequence↗
Regional levels of beta-lipotropin and beta-endorphin in the brain and hypophysis of victims of sudden infant death syndrome.
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Brain Chemistry↗
[Regional distribution of beta-endorphins in the brain of infants expected and not expected to die (preliminary results of an attempt to clarify the pathogenetic relationship between endorphins and sudden infant death)].
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Brain Chemistry↗
Elevation of blood levels of beta-endorphin-like immunoreactivity in patients with shock.
In ten shock patients studied, there was a significant positive correlation between beta-ELIR blood plasma levels and the ratio of pulse rate of radial artery to systolic blood pressure.
Blood Pressure↗