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Biomedical subjects

E L Way

Publications and source records attributed to E L Way.

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

Inhibition of Ca++-ATPase of rat erythrocyte membranes by k-opioid agonists.

The effect of various opioid agonists on Ca++, Mg++-ATPase activity of rat erythrocyte membranes was studied. The Ca++-stimulated component of this enzyme (Ca++-ATPase) showed properties similar to those of the Ca++-pumping ATPase of human erythrocyte membranes, that is, high affinity for Ca++, potentiation by calmodulin, and insensitiveness to Na+. Ethylketocyclazocine (EKC) dose-dependently inhibited this Ca++-ATPase activity at a concentration less than one nM without changing basal Mg++-ATPase activity and this action was reversed by the antagonist Win 44,441. Other opioid agonists mimicked this EKC effect and the rank order of potency was dynorphin (1-13) = greater than EKC greater than levorphanol = morphine = B-endorphin = dihydromorphine greater than leu-enkephalin greater than (D-Ala)2-(D-leu)5-enkephalin (DADL) = morphiceptin. It is concluded that rat erythrocyte membranes possess k-type opioid receptors through which the Ca++-pump is inhibited.

Animals↗
PubMed1984

Dynorphin-(1-13) restores the potency of morphine on the tolerant guinea pig ileum.

Since it has been reported that dynorphin (1-13) enhances the analgetic effects of morphine in mice tolerant either to morphine or beta-endorphin and modulates opioid agonist binding in the tolerant state, experiments were initiated to determine whether comparable phenomenon could be obtained in vitro with dynorphin-(1-13) on the isolated longitudinal muscle of the guinea pig ileum. In the guinea pig ileum rendered tolerant to morphine dynorphin-(1-13) was found to cause a dose-dependent restoration in sensitivity to the inhibitory action of morphine. This phenomenon appeared specific for dynorphin-(1-13) since [Leu5]enkephalin, beta-endorphin, and ethylketocyclazocine did not change significantly the IC50 of morphine in the tolerant preparation. Furthermore, dynorphin-(1-13) attenuated the contraction elicited by naloxone in the morphine tolerant-dependent tissue. It is concluded, that dynorphin-(1-13) restores sensitivity to morphine in the tolerant-dependent state by a modulatory effect probably on the mu receptor that is distinct from its kappa action.

Animals↗
PubMed1984

Calcium transport in and out of brain nerve endings in vitro--the role of synaptosomal plasma membrane Ca2+-ATPase in Ca2+-extrusion.

The effects of cellular cations and ATP on calcium transport in and out of the nerve endings (synaptosomes) of mice brain were studied. The synaptosomes accumulated 45Ca time-dependently in the absence of ATP or other additions for at least 10 min. When ATP was present, the overall 45Ca accumulation was decreased and was maximal at about 4 min, after which it started to decline. Studies on the effects of cations with or without ATP at 4 min revealed selective activities for different cations. Mg2+ inhibited 45Ca accumulation in the absence of ATP but increased 45Ca accumulation when ATP was present. Similarly, ATP increased 45Ca accumulation only when Mg2+ was present. Na+, on the other hand, inhibited 45Ca accumulation both in the presence and absence of ATP and/or Mg2+. K+ increased 45Ca accumulation in the presence of ATP with or without Mg2+; however, K+-stimulation was not noted in the presence of 100 mM Na+, and in fact, K+ became inhibitory. The ATP-stimulated 45Ca accumulation in the presence of Mg2+ peaked within 4-6 min and then declined, suggesting release of 45Ca. Compatible with this notion, in 45Ca-loaded synaptosomes, ATP evoked 45Ca release which was accompanied by the appearance of Pi in the medium. Although ATP-activated 45Ca-release can occur in the presence of Mg2+, Mg2+ is not required and, in fact, is inhibitory. Rapid release of 45Ca was also noted when 45Ca-loaded synaptosomes were incubated in the presence of Na+ without ATP. It is concluded that Mg2+, Na+, K+ and ATP each has a specific role in regulating Ca2+ permeability of the plasma membrane, calcium binding and calcium extrusion.

Adenosine Triphosphate↗
PubMed1984

Opioid-like properties of seven dynorphin (1-10) analogs.

In order to disassociate the k action of dynorphin from its other actions, seven analogs were synthesized and evaluated for pharmacologic activity in comparison with dynorphin (1-13) and dynorphin amide (1-10). Dynorphin (1-10) was modified by protecting the terminal carboxy group, incorporating thioproline at position 10 and substituting methionine for leucine at position 5. All analogs exhibited the ability to inhibit electrically-induced twitches of the guinea pig ileum and mouse vas deferens in a manner that was dose dependent and naloxone reversible. The decapeptide terminating with a pyrrolidine group showed the highest potency in the ilea and mouse vas deferens. None of the analogs showed analgetic activity by the mouse tail flick test. Binding studies using mouse brain synaptosomes showed that all seven analogs can displace the binding of tritiated dihydromorphine (DHM), ethylketocyclazocine (EKC) and D-Ala-D-Leucine enkephalin (DADL). The alterations in chemical structure affected affinity of the analogs to the opiate receptor and their pharmacologic properties differently, suggesting that different opiate subtypes may be involved.

Animals↗
PubMed1984

Characterization of calcium-activated and magnesium-activated ATPases of brain nerve endings.

The properties of Ca2+-activated and Mg2+-activated ATPases of nerve endings from mouse brain were investigated. Ca2+ and Mg2+ each can activate ATP hydrolysis in synaptosomes and its subfractions. Both Ca2+-ATPase and Mg2+-ATPase exhibit high and low affinity for their respective cations. At millimolar concentrations of Ca2+ or Mg2+, several nucleoside triphosphates could serve as substrate for the two enzymes and their specific activities were about three to four times higher in synaptic vesicles than in synaptosomal plasma membranes (SPM). Both in SPM and in synaptic vesicles the relative activity in the presence of Ca2+ was in the order of CTP greater than UTP greater than GTP = ATP, but with Mg2+ the activity was higher with ATP than with the other three triphosphates. Mg2+-ATPase was more active than Ca2+-ATPase in SPM, but in synaptic vesicles the two enzymes exhibited similar activity. Kinetic studies revealed that Mg2+-ATPase was inhibited by excess ATP and not by excess Mg2+. The simultaneous presence of Na+ + K+ stimulated Mg2+-ATPase and inhibited Ca2+-ATPase activity in intact synaptosomes and SPM. The stimulation of Mg2+-ATPase by Na+ + K+ was further increased by increasing Mg2+ concentration and was inhibited by Ca2+ and by ouabain. When Ca2+ and Mg2+ are present together in SPM or synaptic vesicles, the total Pi liberated by the two cations may either increase or decrease, depending on their relative concentrations. Kinetic analyses indicate that Ca2+ and Mg2+ bind independently to the enzyme alone or together at different sites. The results suggest that Ca2+-ATPase and Mg2+-ATPase in SPM or synaptic vesicles may be separate and distinct systems.

Adenosine Triphosphatases↗
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

The effects of opiates on calcium accumulation on rat peritoneal mast cells.

Opiate agonists, morphine, levorphanol and beta-endorphin increased calcium accumulation in rat peritoneal mast cells. This effect was dose dependent and beta-endorphin was 10 times more potent than morphine. The stimulation was stereospecific and inhibited by naloxone. The site of the opiate action appears to be on the outer surface of the plasma membrane since lysis of the mast cell did not alter the response to morphine. Tolerance to the opiate effect was not seen after chronic morphine administration. Morphine did not stimulate histamine release even at relatively high doses in vivo or high concentrations in vitro. It is reasoned that the enhancing effects on external calcium accumulation may reduce the critical cytosol calcium level for effecting histamine release.

Animals↗
PubMed1983

K receptor activities of the three opioid peptide families.

A large number of opioid peptides derived from the three established precursors, pro-opiomelanocortin (POMC), pro-enkephalin A (pro-enk A), and pro-enkephalin B (pro-enk B) were tested for their ability to inhibit electrically induced contractions of the isolated rabbit vas deferens, a preparation sensitive to k but not to mu opioid ligands. In the presence of enzyme inhibitors, all peptides exhibit roughly similar potencies, but the shorter peptides display lower potencies in the absence of enzyme inhibitors. This suggests that the loss in activity is due to their enzymatic degradation. It is concluded that the pro-enk B gene codes peptides with high k-receptor activities, whereas peptides produced by the POMC gene are devoid of k-receptor activity and the peptides, coded by the pro-enk A gene exhibit weak to moderate k-receptor activity.

Animals↗
PubMed1983

Hypersensitivity of the opioid-tolerant guinea pig ileum to electrical stimulation after abrupt agonist removal.

Studies were initiated on the myenteric plexus of the guinea pig ileum to ascertain whether a supersensitive response by abrupt withdrawal could be elicited after tolerance development to opioids. After an optimal twitch response to a given electrical stimulus was established and its height measured, the mean concentration of an opioid to inhibit half maximally the response (IC50) was determined. Subsequently, the tissues were rendered tolerant by incubating with 1x to 5x the IC50 of the agonist for two hours. The agonist was then removed by thorough washing with Ringer solution and the twitch height to the same electrical stimulation was remeasured. The increase in twitch height after washing over that before incubation provided an index of the degree of hypersensitivity of the tolerant-dependent tissue. The degree of hypersensitivity was found to increase in relation to the concentration of morphine. Stereo-specificity of agonist action was evinced by a hypersensitive response to levorphanol and not to dextrorphan. The development of hypersensitivity was blocked by naloxone and by chloramphenicol. Ethylketocyclazocine, a k ligand, did not show a hypersensitivity response. It is concluded that physical dependence on u-opioids can be developed in vitro and that it can be quantified by a super-sensitivity response.

Animals↗
PubMed1983

Possible inhibition of Ca++ pump of rat erythrocyte ghosts by opioid k agonists.

Rat erythrocyte ghosts containing 45Ca++, EGTA, ATP and Pipes buffer (pH 7.2) were prepared for studying opioid effects on Ca++ flux. Ethylketocyclazocine and dynorphin 1-13 (dynorphin) dose-dependently inhibited La+++-sensitive outward 45Ca++ movement at nM concentrations and the effect was naloxone reversible. Morphine and beta-endorphin were also effective at higher concentrations, whereas levorphanol and leu-enkephalin were ineffective. None of the opioids studied inhibited 45Ca++ inward movement. Based on these findings, it is concluded that rat erythrocyte possesses k-type opioid receptors through which the Ca++-pump may be inhibited.

Animals↗
PubMed1983

Synthesis and properties of beta h-endorphin analogs containing the dynorphin-(1-13) sequence.

Two analogs of beta h-endorphin containing dynorphin-(5-13) or dynorphin-(6-13) sequence have been synthesized by the solid-phase method. Biological activities of the analogs have also been investigated. It was found that the opiate activity is about three times more than beta h-endorphin in the guinea pig ileum assay. Both analgesic potency and opiate receptor-binding activity of the analogs are lower when compared with beta h-endorphin. However, the analogs have interesting behavioral effects in mice.

Analgesics↗
PubMed1983

A rapid and simple method for the quantitative determination of tolerance development to opiates in the guinea-pig ileum in vitro.

The rate and degree of tolerance development of morphine, normorphine and methadone were assessed in vitro on the guinea-pig ileum. After the half-maximal concentration to inhibit electrically induced contractions (IC50) for each compound was determined, tolerance to a fixed concentration representing 0.5, 1 or 2 times its IC50 was induced by incubation at 37 degrees C for 1, 2 or 4 hr. The IC50 was then redetermined and the ratio of the IC50 after and before incubation provided a quantitative index of the degree of tolerance development to each agonist. For any given concentration or time, morphine induced the highest, whereas normorphine the intermediate and methadone the lowest degree of tolerance. Tolerance to opiates was associated with some degree of physical dependence as evidenced by the fact that the application of naloxone at the end of each experiment elicited a muscular contraction. Specificity of tolerance development to opiates was demonstrated by several experiments. Coincubation of morphine with naloxone under the same conditions resulted in an inhibition of tolerance development. The stereospecificity of the process was demonstrated by the fact that levorphanol and I-methadone induced a high degree of tolerance, whereas, under the same conditions the less active d-isomers, dextrorphan and d-methadone, did not. Moreover, subsensitivity to acetylcholine, norepinephrine and adenosine monophosphate did not develop in the presence of tolerance to morphine. The validity of this method for investigating mechanisms involved in tolerance and physical dependence was further demonstrated by obtaining data compatible with earlier experiments in vivo that cyclic AMP enhances tolerance development and cycloheximide inhibits this phenomenon. The present method should facilitate studies on the mechanisms involved in opiate tolerance and physical dependence development.

Animals↗
PubMed1982

Effect of 4-aminopyridine and verapamil on the inhibitory action of normorphine on the guinea-pig ileum.

The ability of normorphine to inhibit the muscular twitches elicited by electrical stimulation of the longitudinal muscle of the guinea-pig ileum was antagonized by 4-aminopyridine (4-AP). Incubation with 4-aminopyridine (4-AP) caused a marked increase of the normorphine IC50 and this effect was not modified by naloxone or by the Ca2+ channel blocker, verapamil. Although verapamil caused a significant decrease of the normorphine IC50 it also caused a potent reduction of the contractile effect of acetylcholine. The results suggest that 4-AP and verapamil act on different Ca2+ pools and that verapamil does not primarily affect the presynaptic Ca2+ pool involved in opiate action.

4-Aminopyridine↗
PubMed1982

Application of an irreversible opiate antagonist (beta-FNA, beta-funal-trexamine) to demonstrate dynorphin selectivity for K-opioid sites.

Application of 100 nM beta-FNA for 60 minutes to isolated longitudinal muscles-myenteric plexus preparations from the guinea pig ileum caused a marked antagonism of the inhibitory action of normorphine and leucine enkephalin without greatly affecting the inhibitory potency of dynorphin or ethylketocyclazocine. The interaction of beta-FNA with the normorphine (mu-opiate receptors) appears to be non-equilibrium. Pretreatment with beta-FNA caused a significant increase in the apparent naloxone dissociation constant for normorphine and leucine enkephalin but not for dynorphin or ethylketocyclazocine. The results lend further support to the hypothesis that normorphine and the enkephalins activate preferentially mu-opiate receptors on the ileum, whereas dynorphin interacts predominantly at k-opiate sites.

Animals↗