Search PubMed⌕ Search

Biomedical subjects

H W Kosterlitz

Publications and source records attributed to H W Kosterlitz.

At least 37 records · Page 2Linked to original sources
PubMed1985

The Wellcome Foundation lecture, 1982. Opioid peptides and their receptors.

The remarkable feature of the opioid system is the complexity of its ligands and their interactions with the mu-, delta- and kappa-binding sites. The three endogenous opioid precursors give rise to more than ten opioid fragments. The fragments of pro-opiocortin and pro-enkephalin have affinities mainly to the mu- and delta-binding sites and those of pro-dynorphin have a preference for the kappa-binding site. It is important to realize that some of the larger fragments may have pharmacological actions that are of a non-opioid character. As the endogenous opioid peptides bind to more than one of the types of binding sites, it was necessary to obtain synthetic compounds that bind almost exclusively at one site. There are now agonists for which this aim has been achieved but we still require antagonists that are exclusively selective for only one opioid site. The results obtained with opioid peptides or non-peptides having such qualities would be the physiological basis for a correlation of the binding at mu-, delta- and kappa-receptors with their pharmacological effects. Furthermore, since almost all endogenous opioid ligands are degraded by peptidases, it is necessary to synthesize non-toxic inhibitors of those peptidases that play a role in opioid transmission. Related to this problem is the need to develop methods for the study of the release of various endogenous opioid peptides under physiological conditions.

Animals↗
PubMed1985

Species differences in the concentrations and distributions of opioid binding sites.

Binding at the mu, delta- and kappa-types of opioid binding sites was compared in homogenates from the brains of guinea-pig, rabbit, rat and two mouse strains, under conditions of selective labelling. Species differences were shown by two observations. Firstly, analysis of saturation curves in homogenates of brain from which the cerebellum had been removed showed that in guinea-pig brain the opioid binding sites consist of 24% mu-sites, 32% delta-sites and 44% kappa-sites. In contrast, in rabbit brain the corresponding values are 43% mu-sites, 19% delta-sites and 37% kappa-sites and in rat brain, 46% mu-sites, 42% delta-sites and 12% kappa-sites. In the brains of DBA/2 mice the opioid binding sites are comprised of 51% mu-sites, 29% delta-sites and 20% kappa-sites and in C57BL/10 mice, of 44% mu-sites, 35% delta-sites and 21% kappa-sites; these strain differences are due to significant differences in the concentrations of the mu-sites. Secondly, species differences were found when the binding of single concentrations of tritiated ligands (1 X KD value in whole brain) was determined at mu-, delta- and kappa-sites in six brain regions from guinea-pig, rat or rabbit.

Animals↗
PubMed1985

Types of opioid receptors: relation to antinociception.

The endogenous opioid peptides are derived from three large precursors. Pro-opiocortin and proenkephalin yield [Met]enkephalin, carboxy-extended [Met]enkephalins and [Leu]enkephalin. The fragments of prodynorphin are all carboxy-extended [Leu]enkephalins. Three approaches are of importance for an analysis of the physiological functions of the different endogenous opioid peptides. First, since these peptides interact with more than one of the mu-, delta- and kappa-binding sites and thus with their receptors, it is necessary to synthesize peptides or non-peptides, which bind to only one of the sites. As far as narcotic analgesics are concerned, morphine fulfils these conditions since it interacts almost exclusively with the mu-receptor. Secondly, antagonists are required that are selective for only one of the opioid receptors, even when used in high concentrations. Finally, it is important to find circumscribed areas in the nervous system that possess only one type of opioid receptor. It is now known that in the rabbit cerebellum the opioid receptors are almost exclusively of the mu-type whereas in the guinea-pig cerebellum they are almost exclusively of the kappa-type.

Animals↗
PubMed1985

The opioid receptors in the hamster vas deferens are of the delta-type.

The motor responses of the isolated vas deferens of the hamster were unaffected by opioid-receptor agonists which are selective for the mu- or kappa-receptor, while agonists which show degrees of selectivity for the delta-opioid receptor caused dose-related inhibition of the stimulation-evoked contractions. The agonist action of the enkephalins and their congeners was only apparent when various inhibitors of tissue peptidases were present. Responses to opioid agonists were antagonised in a competitive manner by naloxone and by the selective delta-receptor antagonist, ICI 174864. It is noteworthy that the benzomorphans bremazocine, ethylketocyclazocine and Mr 2034, which are agonists at kappa-receptors in other tissues, are antagonists at the delta-receptor of the hamster vas deferens. Thus, the vas deferens of the hamster contains opioid receptors only of the delta-type and may therefore provide a simple and specific test for the assay of activity at the delta-opioid receptor.

Animals↗
PubMed1985

Proenkephalin- and prodynorphin- derived opioid peptides in guinea-pig heart.

Consecutive high performance liquid chromatography (HPLC) fractionation and the mouse vas deferens assay were used to characterize opioid peptides in the guinea-pig heart. Atria were found to contain at least nine different opioid peptides derived from proenkephalin and prodynorphin. In ventricles at least seven different molecular species which may be derived only from prodynorphin were present. The total opioid activity in atria averaged 22 pmol and in ventricles 11 pmol [Met]enkephalin equivalents per g wet wt. The high content of [Leu]enkephalin in relation to [Met]enkephalin indicates the possibility of a dynorphinergic pathway of cardiac [Leu]enkephalin. Multiple cardiac opioid ligands and receptors, including kappa, may be functionally important in the peripheral control of cardiac performance and coronary circulation.

Animals↗
PubMed1985

[3H]-Dynorphin A (1-8): degradation profile and binding characteristics in brain homogenates.

[3H]-Dynorphin A (1-8) was degraded in brain homogenates at 25 degrees and even at 0 degree C. The peptidase inhibitors bestatin and captopril almost completely protected[3H]-dynorphin A (1-8) from degradation at 0 degree C but had only little effect on binding at this temperature. At 25 degrees C, the binding of [3H]-dynorphin A (1-8) was markedly improved by addition of bestatin, captopril and L-leucyl-L-arginine, which afforded some, but not complete protection from degradation. The results of saturation binding assays at 25 degrees C in the presence of the peptidase inhibitors were variable. However, it was found from saturation binding assays at 0 degree C that the maximum binding capacity for [3H]-dynorphin A (1-8) at the kappa-site is similar to that of [3H]-(-)-bremazocine and [3H]-dynorphin A (1-9).

Animals↗
PubMed1985

Differential postnatal development of mu-, delta-and chi-opioid binding sites in mouse brain.

By selective labelling techniques together with analysis of saturation curves it is shown that the concentrations of the mu-, delta- and chi-binding sites increase during postnatal development particularly in the first few weeks after birth. There are little or no changes in affinity. The rate of development is different for each type of site. The most striking finding is that the development of mu- and chi-sites precedes that of delta-sites.

Age Factors↗
PubMed1985

10-Ketonaltrexone and 10-ketooxymorphone.

Ethylketocyclazocine (1) has greater kappa/mu selectivity than cyclazocine in brain binding assays. 10-Ketonaltrexone (11) and 10-ketooxymorphone (10) were prepared from naltrexone 3-methyl ether and oxycodone, respectively. Bioassays in the myenteric plexus longitudinal muscle preparation of the guinea pig ileum and in the mouse vas deferens, in addition to brain binding assays, demonstrated that 10 and 11 were far less potent than naltrexone (2) and oxymorphone (3) at mu sites and also had little affinity for kappa and delta sites. It is concluded that introduction of the 10-keto group in naltrexone and oxymorphone diminished opioid effects at all binding sites.

Animals↗
PubMed1985

Kappa-binding and degradation of [3H]dynorphin A (1-8) and [3H]dynorphin A (1-9) in suspensions of guinea pig brain membranes.

Following incubation of [3H]dynorphin A (1-8) and [3H]dynorphin A (1-9) with suspensions of guinea pig brain membranes, analysis of the supernatants by HPLC has shown that both peptides are degraded at 25 degrees C and at 0 degrees C. Bestatin and captopril reduce degradation at 0 degrees C but for a similar degree of protection at 25 degrees C arginine-containing dipeptides are also required. The effects of these peptidase inhibitors on the degradation profiles indicate that [3H]dynorphin A (1-8) has three main sites of cleavage: the Tyr1-Gly2, Arg6-Arg7, and Leu5-Arg6 bonds. With [3H]dynorphin A (1-9) as substrate the Arg7-Ile8 and Ile8-Arg9 bonds are also liable to cleavage. In binding assays, in contrast to the effects of peptidase inhibitors on the degradation of unbound [3H]dynorphin A (1-8) and [3H]dynorphin A (1-9), bestatin and captopril have little effect on the binding characteristics of the tritiated dynorphin A fragments at the kappa-site at 0 degrees C. However, at 25 degrees C binding is low in the absence of peptidase inhibitors. When binding at mu- and delta-sites is prevented, the maximal binding capacities of [3H]dynorphin A (1-8), [3H]dynorphin A (1-9), and [3H](-)-bremazocine at the kappa-site are similar; [3H]dynorphin A (1-9) has 5-10 times higher affinity for the kappa-site than [3H]dynorphin A (1-8). Comparison of the effects of peptidase inhibitors on unbound dynorphin A fragments with their effects in binding assays suggests that the bound peptides are protected from the action of peptidases.

Animals↗
PubMed1985

The use of [3H]-[D-Pen2,D-Pen5]enkephalin as a highly selective ligand for the delta-binding site.

The characteristics of the binding of [3H]-[D-Pen2,D-Pen5]enkephalin were determined in homogenates of guinea-pig and rat brain. In the guinea-pig, the maximum binding capacity for [3H]-[D-Pen2,D-Pen5]enkephalin was 4.19 pmol g-1 and the KD 1.61 nM. In the rat, the corresponding values were 2.47 pmol g-1 and 5.42 nM. In both species, the maximum binding capacity and the affinity were not altered when mu-binding was suppressed with [D-Ala2,MePhe4,Gly-ol5]enkephalin. The mu-agonists, [D-Ala2,MePhe4,Gly-ol5]enkephalin and morphine, displaced a small portion of the binding of [3H]-[D-Pen2,D-Pen5]enkephalin with high affinities.

Animals↗
PubMed1985

Tritiated etorphine and naloxone binding to opioid receptors in caudate nucleus in schizophrenia.

Opioid receptor binding sites were assessed in membrane preparations of caudate nucleus from post-mortem brains of controls and of patients with schizophrenia. There was no difference between the two groups in the total specific binding of 3H-etorphine or in its 'mu' and ('delta + kappa') components. Similarly, the binding of 3H-naloxone did not differ between patients and controls. It is concluded that a previous report of reduced opioid receptors in caudate of schizophrenics is unlikely to prove a consistent finding and that the results of the present study offer no support to the claim that there is a general disturbance in opiate mechanisms in schizophrenia.

Aged↗
PubMed1984

Hamster vas deferens contains delta-opioid receptors.

Electrical field stimulation of the hamster, isolated vas deferens produces regular contractions that are unaffected by opioid-receptor agonists which are selective for the mu- or kappa-receptor types. However, agonists which show a selectivity for the delta-opioid receptor produce dose-related inhibitions of the stimulation-evoked contractions. Response of delta-opioid receptor agonists are antagonized in a competitive fashion by the selective delta-receptor antagonist ICI 174864, and also by naloxone. The "kappa-opioid-agonist" benzomorphans, bremazocine and ethylketocyclazocine are antagonists in the hamster vas deferens; bremazocine is particularly potent in this regard. In conclusion, the hamster, isolated vas deferens may contain only delta-opioid receptors and provides a simple and specific test for the assay of activity at the delta-opioid receptor.

Animals↗
PubMed1984

Acute and persistent effects of beta-funaltrexamine on the binding and agonist potencies of opioids in the myenteric plexus of the guinea-pig ileum.

In binding assays with homogenates of the myenteric plexus-longitudinal muscle of the guinea-pig, beta-funaltrexamine is more potent at displacing mu-binding than kappa-binding. Incubation of homogenates with beta-funaltrexamine (100 or 1000 nM) for 30 min at 37 degrees C followed by repeated washing with drug-free Tris buffer does not alter the binding of either the selective mu-ligand [3H]-[D-Ala2, MePhe4,Gly-ol5]enkephalin or of [3H]-(-)-bremazocine made selective for the kappa-binding site by the addition of unlabelled mu- and delta-ligands. This observation is surprising since, after treatment with beta-funaltrexamine (100 nM), the IC50 values for the inhibition of the contraction of the myenteric plexus-longitudinal muscle by the mu-ligand [D-Ala2,MePhe4, Gly-ol5]enkephalin are increased 12-fold whereas the IC50 values obtained with the selective kappa-ligand U-50, 488H remain unaltered. It is proposed that the irreversible blockade produced by beta-funaltrexamine is not due to inhibition at the mu-site per se but to an interference with the link between the binding and the effector response.

Animals↗
PubMed1984

Effects of D-amino acid substitution in dynorphin A (1-9) on binding at the mu-, delta- and kappa-sites.

Incorporation of D-Ala into position 2 of dynorphin A (1-9) (Tyr-Gly-Gly-Phe-Leu-Arg-Arg-Ile-Arg) causes a loss of kappa-selectivity. Similarly, replacement of L-Arg by D-Arg also diminished kappa-selectivity. The combination of D-Ala in position 2 and D-Arg in positions 6, 7 and 9 further reduces kappa-binding. In experiments at 25 degrees C, D-Ala substitution leads to a stable peptide whereas D-Arg substitutions do not stabilize the peptides.

Animals↗
PubMed1984

Opioid binding sites of the kappa-type in guinea-pig cerebellum.

(-)-[3H]Bremazocine interacts almost equally well with the mu-, delta- and kappa-types of opioid binding sites. In homogenates of guinea-pig cerebellum, it was bound with high affinity (KD = 0.046 nM) and the maximum binding capacity was 7.04 pmol/g wet wt of tissue. When the mu- and delta-binding of (-)-[3H]bremazocine was prevented with unlabelled ligands, a KD of 0.034 nM and a capacity of 5.94 pmol/g tissue was found for the kappa-binding site, which therefore comprised 84% of the opioid binding sites in the cerebellum. Autoradiographic analysis showed that the binding of (-)-[3H]bremazocine was relatively low in lobules IX and X and that it was predominantly located in the molecular and to a lesser extent in the granular layers. The addition of unlabelled mu- and delta-ligands did not alter the distribution. Thus, the guinea-pig cerebellum contains opioid binding sites of which almost all are of the kappa-type and is therefore an ideal tissue for the isolation of kappa-receptors and for the investigation of their biochemical and pharmacological properties.

Animals↗
PubMed1984

Selectivities of opioid peptide analogues as agonists and antagonists at the delta-receptor.

The endogenous opioid ligands interact with more than one of the mu-, delta- and kappa-binding sites. By the use of binding assays and bioassays, enkephalin analogues have been assessed for their selectivity for binding at the delta-binding site and for their agonist and antagonist activities at the delta-receptor. The electrically-induced contractions of myenteric plexus-longitudinal muscle preparations of the guinea-pig ileum were inhibited by mu- and kappa-receptor ligands. Inhibitions were seen with mu-, delta- and kappa-receptor ligands in the mouse vas deferens, mainly with mu-receptor ligands in the rat vas deferens and only with kappa-receptor ligands in the rabbit vas deferens. From observations on a considerable number of [Leu5] enkephalin analogues, it has been concluded that [D-Pen2, D-Pen5] enkephalin and [D-Pen2, L-Pen5] enkephalin are the most selective delta-agonists and that N,N-diallyl-Tyr-Aib-Aib-Phe-Leu-OH is the most selective antagonist (Aib = alpha-aminoisobutyric acid). The binding of these peptides at the delta-site is 99% of the total binding. As to potency, the agonists are superior to the antagonists.

Animals↗
PubMed1984

Radioligands for probing opioid receptors.

The three endogenous opioid precursors of almost 30000 Da are pro-opiocortin, proenkephalin and prodynorphin. Pro-opiocortin contains beta-endorphin, melanotropins and ACTH. Proenkephalin yields one [Leu5]enkephalin, three [Met5]enkephalins, one [Met5] enkephalyl-Arg-Arg-Val-NH2 (metorphamide or adrenorphin), one [Met5]enkephalyl-Arg-Gly-Leu and one [Met5]enkephalyl-Arg-Phe. [Leu5]enkephalin is common to all fragments of prodynorphin; its carboxyl extension by Arg-Lys leads to alpha- and beta-neo-endorphin and its carboxyl extension by Arg-Arg gives two dynorphins A and B of 17 and 13 amino acids, respectively. Another endogenous peptide is dynorphin A (1-8). The three main opioid binding sites are mu, delta and kappa. Their analysis has been facilitated by the synthesis of analogues of peptides and non-peptide compounds, which have selective agonist or antagonist action at only one site. The various physiological roles of the three types of the opiate receptor have so far not been sufficiently investigated.

3,4-Dichloro-N-methyl-N-(2-(1-pyrrolidinyl)-cycloh↗