Search PubMed⌕ Search

Biomedical subjects

H H Loh

Publications and source records attributed to H H Loh.

At least 163 records · Page 9Linked to original sources
PubMed1985

Specific opioid-amphetamine interactions in the caudate putamen.

Bilateral microinjection of morphine (0.003-3 micrograms/side) into the caudate putamen enhances the behavior induced by the IP injection of 1 mg/kg d-amphetamine phosphate in a dose-related manner. The duration of activity was prolonged and ambulation was changed to d-amphetamine stereotypy, a behavior normally associated with higher doses of d-amphetamine. The opioid activity was stereospecific in that levorphanol was active, whereas dextrorphan was not. The enhancement of d-amphetamine-induced behavior by the opioids was blocked by naloxone. D-ala2-met-Enkephalin also enhanced the amphetamine-induced behavior. This enhancement appears to be specific to the caudate putamen because the oral stereotypy observed appears to be a unique action of amphetamine in this region of the brain.

Amphetamine↗
PubMed1985

Involvement of both inhibitory and stimulatory guanine nucleotide binding proteins in the expression of chronic opiate regulation of adenylate cyclase activity in NG108-15 cells.

Chronic etorphine treatment of neuroblastoma X glioma NG108-15 cells results in both an increase in adenylate cyclase activity (upon addition of the opiate antagonist naloxone) as well as an homologous desensitization of the opiate receptor. The continued ability of opiate agonists to regulate adenylate cyclase activity following opiate receptor desensitization can be understood by proposing that the catalytic subunit of adenylate cyclase in NG108-15 cells is under tonic regulation by both guanine nucleotide regulatory (Ni) and stimulatory (NS) components. Inactivation of Ni by pertussis toxin (PT) treatment resulted in elevated adenylate cyclase activities comparable to those observed in control cells following chronic opiate treatment. This increased enzymatic activity could not be further induced by PT treatment of cells exposed to opiate previously. In addition, procedures that prevented receptor-mediated activation of NS, i.e., treatment with NaF or desensitization of the stimulatory receptors (prostaglandin E1, adenosine) eliminated the increase in adenylate cyclase activity induced by naloxone following chronic opiate exposure. Hence, the increase in enzymatic activity observed following chronic opiate treatment may be due to a loss in tonic inhibitory regulation of adenylate cyclase mediated through Ni resulting in the unimpeded expression of NS activity. This tonic inhibition of adenylate cyclase activity is one of the multiple mechanisms by which Ni regulates adenylate cyclase in this cell line.

Adenosine↗
PubMed1984

Dynorphin A: inhibitory effect on other opiate ligand bindings in the mouse brain.

The opioid peptide dynorphin A antagonizes morphine-induced analgesia in vivo and inhibits opiate binding in vitro, although most of it is rapidly degraded under both conditions. The inhibitory effect was present even in tissue treated in vivo with dynorphin A and assayed in vitro without it. Shorter fragments of this peptide lacked these effects, indicating that the apparent potency did not result from a metabolite. Na+ ion specifically reversed both agonist and antagonist binding from in vitro inhibition by dynorphin A. These results are discussed in terms of current opioid receptor theories.

Animals↗
PubMed1984

Dynorphin1-13: interaction with other opiate ligand bindings in vitro.

Dynorphin1-13 is a potent inhibitor of electrically-induced contractions in the guinea pig ileum, where it has the properties of kappa-(ethylketocyclazocine) type opioids. In the brain, however, it has no analgesic potency, yet inhibits that induced by morphine. To gain further insight into its mechanism of action in the latter system, we tested its ability to compete for the binding of several opiates to brain membranes in vitro. Dynorphin1-13 inhibited the binding of all ligands examined, including dihydromorphine, D-Ala2-D-Leu5-enkephalin, ethylketacyclazocine (EKC) and naloxone. In all cases, it reduced the number of high affinity sites and, in the case of EKC, it also increased the Kd. We conclude that the mechanism of dynorphin inhibition is not simple rapidly reversible competition and is certainly not identical with respect to all opiate ligands.

Animals↗
PubMed1984

Down-regulation of opiate receptor in neuroblastoma x glioma NG108-15 hybrid cells. Chloroquine promotes accumulation of tritiated enkephalin in the lysosomes.

Opiate receptor down-regulation in neuroblastoma X glioma NG108-15 hybrid cells possibly involved the internalization of ligand-receptor complexes during chronic treatment. However, receptor internalization was not supported by the observed decrease in [3H] enkephalin(D-Ala2,D-Leu5) ( [3H]DADLE) associated with the hybrid cells during prolonged incubation with 10 nM [3H]DADLE at 37 degrees C. This decrease in [3H]DADLE bound was determined to be due to degradation of the ligand-receptor complexes, for a time-dependent increase in [3H]DADLE bound was observed when the incubations were carried out in the presence of 0.1 mM chloroquine. The increase did not exceed the amount of down-regulated receptor, could be blocked by naloxone, and was not observed at 24 degrees C. The [3H]DADLE bound in the presence of chloroquine was not sensitive to trypsin or to 20 microM diprenorphine. The accumulated [3H]DADLE was demonstrated to be intracellularly located by the fractionation of the homogenates in self-generating Percoll gradients. In the presence of chloroquine, a time-dependent translocation of [3H]DADLE from the plasma membrane-enriched fractions to the lysosome-enriched fractions was observed. The translocation was not observed at 24 degrees C in the presence of chloroquine or at 37 degrees C in the absence of chloroquine. The [3H]DADLE in the lysosome-enriched fractions was not sensitive to trypsin and remained bound in the presence of chloroquine. With the removal of chloroquine, an increase in the release of [3H]DADLE into the medium was observed. Sephadex G-50 column chromatography of the sodium deoxycholate extracts of the lysosome-enriched fractions suggested that the [3H]DADLE was bound to macromolecules intracellularly. Thus, chronic [3H]DADLE treatment of the hybrid cells resulted in an internalization of ligand-receptor complexes which were degraded in the lysosomes. Subsequently, the [3H]DADLE was regurgitated by the hybrid cells.

Animals↗
PubMed1984

Dynorphin inhibition of the neurotensin contractile activity on the myenteric plexus.

Nanomolar concentrations of neurotensin caused a dose-dependent contraction of the longitudinal muscle layer of the guinea-pig ileum. The contractile activity of neurotensin was partially blocked by tetrodotoxin or atropine, indicating that a component of the neurotensin-mediated contraction is indirect in nature and likely involves the release of endogenous acetylcholine from nervous terminals in the myenteric plexus. Dynorphin and related peptide fragments also blocked in part the neurotensin contraction; the potency of this opioid peptide was about the same as that of atropine. Other peptides and alkaloids tested for ability to block the neurotensin contractures included the enkephalins, beta-endorphin, normorphine and the ketocyclazocines; all these opioids inhibited in a dose-dependent fashion the neuronal component of the excitatory effect of neurotensin. The potency of these compounds to reduce the contractions of neurotensin showed good correlation with the potency of these agents to depress by 50% the electrically evoked neuromuscular twitches in the same tissue (r = 0.99); in these tests dynorphin was found to be the most potent of the endogenous opioid-like peptides. The dynorphin blockade was selective to the excitatory effect of neurotensin because the opioid peptide did not antagonize the contractile action of acetylcholine, histamine, substance P, angiotensin II, bradykinin, Ba++ or K+ ions. In addition, somatostatin, vasointestinal peptide, gastrin or adenosine did not modify the potency of neurotensin whereas thyrotropin releasing hormone and epinephrine caused a modest doubling of the neurotensin EC50. The inhibitory action of dynorphin was reduced in the presence of naloxone, suggesting that the interaction involved opiate receptors. Morphine tolerance was not extended to the inhibitory action of dynorphin as evidenced by the finding that the potency of dynorphin-(1-13) to block the neurotensin responses was increased after chronic morphine exposure. In contrast, the potency of dynorphin-(1-13) was significantly reduced in tissues rendered tolerant to the action of ketocyclazocine or ethylketocyclazocine, suggesting that the action of dynorphin could be partially mediated via occupation of K-opiate receptors. Thus, a cholinergic-neuronal component activated by neurotensin on the myenteric plexus appears to be under the inhibitory influence of opiate receptors, suggesting that dynorphin may play a role in the modulation of cholinergic synapses on the enteric nervous system.

Animals↗
PubMed1983

Effect of dynorphin-(1-13) and related peptides on respiratory rate and morphine-induced respiratory rate depression.

Previous studies from our laboratory have shown that the opioid peptide dynorphin-(1-13), although not analgesic when given by itself, can inhibit morphine-induced analgesia in naive mice and potentiate it in morphine tolerant mice. In the present study, we examined the effect of dynorphin-(1-13) with two other dynorphin-like peptides, alpha-neoendorphin and dynorphin-(1-10) amide, on respiration. Our results show that none of the peptides studied had any significant activity on the respiratory rate in mice when given alone. However, in the presence of morphine, dynorphin-(1-13) antagonized the morphine-induced respiratory rate depression in morphine-tolerant animals; alpha-neoendorphin enhanced the morphine-induced respiratory rate depression in naive but had no effect in morphine-tolerant animals and dynorphin-(1-10) amide had no modulatory effect on the morphine-induced respiratory rate depression in either group of animals.

Animals↗
PubMed1983

Dynorphin-(1-13): the effect of in vivo treatment on opiate bindings in vitro.

Mice were injected intracerebroventricularly with different doses of dynorphin-(1-13) in vivo and decapitated after different intervals of time; crude P2 fractions were then prepared and used for in vitro binding with [3H]dihydromorphine (DHM) and [3H][D-Ala2,D-Leu5]enkephalin. Dynorphin pretreatment was found to decrease DHM binding in vitro in a both time-dependent and dose-dependent manner. Its maximal effect lasted from 30 min to 3 h and recovery took place in 6-12 h. Analysis of the Scatchard plots showed that dynorphin decreased the number of high affinity-binding site for DHM, while KD of this site was essentially unaltered. Neither Bmax nor KD of low affinity site were much affected. A similar decrease in high affinity Bmax for DADLE was also obtained; however, the recovery was even more rapid. Extra washes of tissue did not significantly increase the binding, but preincubation of tissue in the presence of morphine reversed the dynorphin effect and increased DHM binding significantly to almost control levels. The probable mechanism of dynorphin in decreasing opiate receptor binding is discussed.

Animals↗
PubMed1983

Endogenous opioid peptides: comparative evaluation of their receptor affinities in the mouse brain.

The affinities of certain endogenous opioid peptides and related sequences for mu, delta and k opiate receptors have been determined in membrane preparations from mouse brain. It was found that the KIs for the delta receptor changed very little when the sequence of the enkephalins was enlarged; on the contrary, the mu and especially the k activity were highly dependent not only on the specificity of the sequence but also on the length of the peptide. Most of the peptides have similar affinities for more than one receptor type. The enkephalins are the most selective for the delta receptor. beta-Endorphin, BAM-12P, BAM-22P, peptide E and dynorphin A display the best potency at the mu site. Dynorphin A (1-8), dynorphin A and dynorphin B are the most selective for the k site.

Animals↗
PubMed1983

Modulation of adenylate cyclase activity by a cytosolic factor following chronic opiate exposure in neuroblastoma x glioma NG108-15 hybrid cells.

A soluble cytosolic factor from neuroblastoma x glioma NG108-15 hybrid cells stimulates adenylate cyclase activity in isolated membrane preparations. This cytosolic component is heat stable, pronase insensitive, has a molecular weight less than 350 daltons and an absorbance peak at 260 nm. The stimulation is immediate, independent of Ca++ and exhibits a sigmoidal concentration dependency curve. The cytosolic factor stimulated adenylate cyclase activity in etorphine treated cells (100 nM etorphine, 16 hrs) to a greater extent than in control cells. In addition, cytosolic factor derived from etorphine treated cells, as compared to control cells, displayed an increased capacity to stimulate adenylate cyclase. It is suggested that the observed cytosolic factor may be adenosine and that cells chronically treated with an opiate exhibit an increase in both concentration and sensitivity to this agent.

Adenylyl Cyclases↗
PubMed1983

In vivo incorporation of a cholesterol-like fluorescent probe into rat brain membranes.

The incorporation of the fluorescent sterol analog, cholesta-5,7,9-trien-3 beta-ol, into rat brain P2 fractions and its feasibility to detect membrane perturbations by polarization analysis were evaluated. This investigation involved the administration (i.c.v.) of the fluorescent compound in sesame oil with subsequent determination of optimal dose and the half-life of this compound within the membrane fraction. The dose giving maximum polarization was found to be 0.65 mumole, and pharmacokinetic analysis revealed that the disappearance of this analog followed a two-compartment open model with an elimination half-life from the membrane fraction of 655 hr. Increasing temperature of the membrane preparation showed an increase in polarization, indicating a consequent decrease in mobility of the molecule, while addition of a fluidizing agent like ethanol caused a decrease in polarization, and thus the expected increase in fluidity.

Animals↗
PubMed1983

Isolation of opiate binding components by affinity chromatography and reconstitution of binding activities.

Rat brain membranes exhibiting stereospecific opiate binding activity were solubilized by sonication and detergent treatment. The active material could be bound to an affinity column containing 6-succinylmorphine but could not be eluted with free agonist. Although two protein peaks could be eluted with NaCl, neither possessed binding activity; however, one of the peaks (A), in combination with an acidic lipid fraction, eluted subsequently from the column, showed stereospecific binding. Opiate ligands of the mu type bound to this protein/lipid mixture with an order of affinities closely correlating with those of the original membrane but one to two orders of magnitude lower; binding of delta, kappa, and sigma prototype opioids was considerably less. The protein/lipid mixture also competed with the membranes for mu ligands. These results suggest that the isolated protein-lipid complex may be a component of the opiate receptor and, specifically, the mu receptor or binding site. However, because of the lower affinities of mu opiates for this complex, it is conceivable that some essential membrane component is still missing. Preliminary analysis of peak A indicates that it contains a broad spectrum of protein bands, but it remains to be seen which of these are essential for activity.

Animals↗
PubMed1983

Attenuation of enkephalin activity in neuroblastoma X glioma NG108-15 hybrid cells by phospholipases.

The role of membrane phospholipids in enkephalin receptor-mediated inhibition of adenylate cyclase (EC 4.6.1.1) activity in neuroblastoma X glioma NG108-15 hybrids was studied by selective hydrolysis of lipids with phospholipases. When NG108-15 cells were treated with phospholipase C from Clostridium welchii at 37 degrees C, an enzyme concentration--dependent decrease in adenylate cyclase activity was observed. The basal and prostaglandin E1 (PGE1)-stimulated adenylate cyclase activities were more sensitive to phospholipase C (EC 3.1.4.3) treatment than were the NaF-5'-guanylylimidodiphosphate (Gpp(NH)p)-sensitive adenylate cyclase activities. Further, Leu5-enkephalin inhibition of basal or PGE1-stimulated adenylate cyclase activity was attenuated by phospholipase C treatment, characterized by a decrease of enkephalin potency and of maximal inhibitory level. [3H]D-Ala2-Met5-enkephalinamide binding revealed a decrease in receptor affinity with no measurable reduction in number of binding sites after phospholipase C treatment. Although opiate receptor was still under the regulation of guanine nucleotide after phospholipase C treatment, adenylate cyclase activity was more sensitive to the stimulation of Gpp(NH)p. Thus, the reduction of opiate agonist affinity was not due to the uncoupling of opiate receptor from N-component. Further, treatment of NG108-15 hybrid cell membrane with phospholipase C at 24 degrees C produced analogous attenuation of enkephalin potency and efficacy without alteration in receptor binding. The reduction in enkephalin potency could be reversed by treating NG108-15 membrane with phosphatidylcholine, but not with phosphatidylserine, phosphatidylinositol, or cerebroside sulfate. The enkephalin activity in NG108-15 cells was not altered by treating the cells with phospholipase A2 o phospholipase C from Bacillus cereus. Hence, apparently, there was a specific lipid dependency in enkephalin inhibition of adenylate cyclase activity.

Adenylyl Cyclases↗