Humans ; Dynorphins/metabolism* ; Receptors, Opioid, kappa/metabolism* ; Cryoelectron Microscopy

Amygdala ; drug effects ; metabolism ; Animals ; Conditioning (Psychology) ; Morphine ; adverse effects ; Rats ; Receptors, Opioid, kappa ; metabolism

A series of aralkyl-ketone-4-piperidol derivatives were synthesized and tested for their analgesic activities. All of the novel 30 compounds were prepared from 4-piperidone and alpha-halo-aralkyl-ketone through five steps, including Boc protection, nucleophilic addition in presence of CeCl3/NaI catalyst, deprotection, condensation and salification. Their structures were confirmed by 1H NMR and HRMS. Preliminary in vivo pharmacological trials showed that most of the synthesized compounds revealed analgesic effects. Among the tested compounds, 8, 13 and 22 exhibited potent analgesic activities in both mice writhing and mice hot plate model. The three compounds have low affinity for mu, delta, kappa receptors, which is a chance to find a better precursor of non-opioid analgesic for further optimization.
Analgesics, Non-Narcotic ; chemical synthesis ; chemistry ; pharmacology ; Animals ; Mice ; Molecular Structure ; Pain Measurement ; Pain Threshold ; drug effects ; Piperidones ; chemical synthesis ; chemistry ; pharmacology ; Receptors, Opioid, delta ; metabolism ; Receptors, Opioid, kappa ; metabolism ; Receptors, Opioid, mu ; metabolism ; Structure-Activity Relationship

AIMTo design and synthesize new chiral 8-(substituted) amino-analogues of 3-[(tetrahydro-2-furanyl)methyl] benzomorphans, to expand knowledge of the structure-activity relationship (SAR) for 8-aminobenzomorphan.

METHODSTarget compounds were synthesized from the 8-triflate of the optically active 3-[(tetrahydro-2-furanyl)methyl]-2,6-methano-benzomorphans using Pd-catalyzed aminations. Opioid receptor binding experiments were performed to evaluate their biological activities.

RESULTSBoth 8-amino and 8-phenylamino analogues showed lower binding affinity for mu, delta and kappa receptors than corresponding 8-hydroxy-3-[(tetrahydro-2-furanyl)methyl]-2,6-methano-benzomorphan in vitro.

CONCLUSIONThe relative poor binding affinity of the target compounds did not warrant conducting the in vivo studies to determine if they have the profile(kappa agonist/mu antagonist) that will be potentially useful in the treatment of drug addiction. Further study is in progress.

Animals ; Benzomorphans ; chemical synthesis ; chemistry ; pharmacology ; Brain ; metabolism ; Furans ; chemical synthesis ; chemistry ; pharmacology ; Guinea Pigs ; Molecular Structure ; Narcotic Antagonists ; chemical synthesis ; chemistry ; pharmacology ; Radioligand Assay ; Receptors, Opioid ; metabolism ; Receptors, Opioid, delta ; metabolism ; Receptors, Opioid, kappa ; metabolism ; Receptors, Opioid, mu ; metabolism ; Structure-Activity Relationship

This study investigates whether kappa-opioid receptor and ORL1 receptor may interact to form a heterodimer. In immunofluorescence and co-immunoprecipitation experiments, differentially epitope-tagged receptors, colocalization and heterodimerization of kappa-opioid receptor and ORL1 receptor were used and examined in primary culturing rat neurons, Chinese hamster ovary (CHO) or human embryonic kidney 293 (HEK293) cells. The results show that fluorescence of both kappa-opioid receptor and ORL1 receptor were overlapping in primary culturing hippocampal and cortical neurons. Similarly in co-expressing CHO or HEK293 cells, HA-KOR and Myc-ORL1 were almost exclusively confined to the membranes, revealing extensive colocalization. When Flag-KOR and Myc-ORL1 were co-expressing in CHO cells, heterodimerization was identified to have the ability to co-immunoprecipitate ORL1-receptors with kappa-opioid receptor and vice versa. In the current study, further evidence was provided for the direct interaction of two subtypes of opioid receptors, kappa-opioid receptor and ORL1-receptor, to form the heterodimerization. The finding represents the novel pharmacological mechanism for modulation of opioid receptor function as well as diversity of G protein-coupled receptors.
Animals ; CHO Cells ; Cells, Cultured ; Cerebral Cortex ; cytology ; metabolism ; Cricetinae ; Cricetulus ; Dimerization ; Female ; HEK293 Cells ; Hippocampus ; cytology ; metabolism ; Humans ; Immunoprecipitation ; Male ; Neurons ; cytology ; metabolism ; Rats ; Rats, Wistar ; Receptors, Opioid ; metabolism ; Receptors, Opioid, kappa ; metabolism

AIMTo observe the neuroprotective effect of limb ischemic post-conditioning (LIPC) on local brain ischemia and reperfusion injury in rat, and to investigate whether mitochondrial ATP sensitive potassium channel (mito K(ATP)) and kappa-opioid receptor were involved in the neuroprotection.

METHODSRats were randomly divided into 6 groups that were ischemia/reperfusion group, unilateral hindlimb ischemia group (uLIPC), bilateral hindlimbs ischemia group (bLIPC), bLIPC + antagonist of kappa-opioid receptor nor-binaltorphimine (nor-BNI) group, bLIPC + mito K(ATP) blocker 5-hydroxydecanoate(5-HD) group, bLIPC + extracorporeal circulation of bilateral hindlimbs via femoral arteries (EC) group. Cerebral ischemia was induced by middle cerebral artery occlusion (MCAO), neurological scores, plasma levels of dynorphin and enkephalin, the brain infarct areas were determined after reperfusion.

RESULTSUnilateral LIPC partially improved the neurological score after local brain ischemia and reperfusion injury in rat (P < 0.05), and decreased the infarct area compared with the untreated group undergoing brain ischemia and reperfusion (P < 0.01). Bilateral LIPC significantly improved the neurological score after local brain ischemia and reperfusion injury (P < 0.01), and decreased the infarct area (P < 0.01). The neurological scores of bilateral LIPC group were significant higher than those of unilateral LIPC (P < 0.05). The plasma level of dynorphin was significantly increased (P < 0.01) at 5, 15, 30 min, 1 and 2 h after bilateral LIPC, however, it deceased to the normal level at 12 h after bilateral LIPC. The plasma level of enkephalin showed no obvious change after bilateral LIPC (P > 0.05). nor-BNI (25 nmol/L) and 5-HD (10 mg/kg) abolished the effect of bilateral LIPC (P < 0.01).

CONCLUSIONLIPC protects rat from local brain ischemia and reperfusion injury. Mito K(ATP) may be involved in the neuroprotection, and kappa-opioid receptor may also participate in the protective effect.

Animals ; Brain Ischemia ; physiopathology ; Extremities ; blood supply ; Ischemic Postconditioning ; methods ; Male ; Potassium Channels ; metabolism ; Random Allocation ; Rats ; Rats, Sprague-Dawley ; Receptors, Opioid, kappa ; metabolism ; Reperfusion Injury ; prevention & control

OBJECTIVETo investigate the cardiac effect of interleukin-2 (IL-2) and to explore the underlying mechanism.

METHODSThe video tracking system and spectrofluorometric method were used to measure the cell contraction and intracellular calcium. Fura-2/AM was used as a calcium fluorescence probe. Langendorff perfusion technique was used to determine the effect of IL-2 on the intact heart.

RESULTSCompared with the control group, IL-2 5 U/ml, 50 U/ml significantly decreased cell contraction amplitude [(74.95+/-4.79) vs (98.09+/-5.02)%, (64.30+/-5.24) vs (97.38+/-4.05)%], peak velocity of cell shortening [(70.23+/-4.85)% vs (98.09+/-5.46)%, (61.15+/-5.20)% vs (97.38+/-6.85)%], peak velocity of cell relengthening [(71.22+/-4.79)% vs (98.32+/-6.08)%, (68.16+/-5.24)% vs (97.55+/-5.00)%] and end- diastolic cell length [(88.28+/-5.84)% vs (97.95+/-5.52)%, (84.18+/-6.52)% vs (98.94+/-6.76)%]. IL-2 (5 U/ml, 50 U/ml) also markedly inhibited intracellular calcium transient [(74.94+/-4.90)% vs (98.09+/-3.74)%,(71.00+/-5.28)% vs (97.38+/-5.52)%], and elevated end-diastolic calcium level of ventricular myocytes [(113.91+/-5.93)% vs (100.10+/-3.02)%, (119.09+/-7.12)% vs (100.52+/-6.00)%], which were attenuated by the opioid receptor antagonist naloxone (Nal,10 nmol/L). In the isolated perfused rat heart,when compared with the control group, IL-2 50 U/ml markedly decreased left ventricular developed pressure [(79.91+/-2.18) vs (93.84+/-2.94)mmHg], maximal rate of rise of left ventricular pressure [(2370.7358.29) vs (2591.50+/-62.81)mmHg] maximal rate of fall of left ventricular [-(1460.95+/-38.6) vs -(1634.24+/-54.05) mmHg/s] and heart rate [(217.35+/-10.56) vs (244.52+/-11.23) beats/min], but increased left ventricular end-diastolic pressure (11.44+/-1.02 vs 9.23+/-0.46). Pretreatment with Nal (10 nmol/L) antagonized the cardiac depression and left ventricular end-diastolic pressure elevation induced by IL-2.

CONCLUSIONThe cardiac effect of IL-2 is mediated by opioid receptors on the membrane of cardiomyocytes.

Animals ; Calcium ; metabolism ; Depression, Chemical ; In Vitro Techniques ; Interleukin-2 ; pharmacology ; Male ; Myocardial Contraction ; drug effects ; Naloxone ; pharmacology ; Rats ; Rats, Sprague-Dawley ; Receptors, Opioid, kappa ; drug effects ; physiology

OBJECTIVETo investigate the effect of kappa-Opioid receptors in the cardioprotection elicited by ischemic postconditioning and the underlying mechanism.

METHODSThe isolated perfused hearts of male Sprague-Dawley rats were subjected to 30 min of global ischemia followed by 120 min of reperfusion. formazan content of myocardium was measured spectrophotometrically, and the level of lactate dehydrogenase (LDH) in the coronary effluent was also measured. In isolated ventricular myocytes hypoxia postconditioning was achieved by 3 cycles of 5 min reoxygenation/5 min hypoxia starting at the beginning of reoxygenation, and cell viability was measured.

RESULTIn the Langendorff perfused rat heart model, ischemic postconditioning (6 cycles of 10 s reperfusion/10 s global ischemia starting at the beginning of reperfusion) increased formazan content, reduced LDH release, improved the recovery of the left ventricular developed pressure, maximal rise/fall rate of left ventricular pressure, left ventricular end-diastolic pressure and rate pressure product (left ventricular developed pressure multiplied by heart rate), attenuated the decrease of coronary flow during reperfusion and increased the isolated cell viability. Pretreatment with nor-BNI, an antagonist of kappa-Opioid receptors and mitoK(ATP) blocker 5-HD attenuated the effect of ischemic/hypoxic postconditioning.

CONCLUSIONPostconditioning may protect myocardium against ischemia/reperfusion injury via activating kappa-Opioid receptors and mitoK(KATP).

Animals ; In Vitro Techniques ; Ischemic Preconditioning, Myocardial ; methods ; L-Lactate Dehydrogenase ; metabolism ; Male ; Myocardial Reperfusion Injury ; metabolism ; physiopathology ; prevention & control ; Myocardium ; metabolism ; pathology ; Potassium Channels ; metabolism ; physiology ; Rats ; Rats, Sprague-Dawley ; Receptors, Opioid, kappa ; metabolism ; physiology

AIMIn order to study the effects of kappa-opioid receptor activation, we used cultured cardiomyocytes to study the inhibitory effects of U50,488H on cellular proliferation, protein content in the presence or absence of nor-binaltorphimine (nor-BNI).

METHODSThe cellular proliferation was determined with crystal violet staining and the protein content was assayed with Lowry's method.

RESULTSA kappa-opioid receptor agonist U50,488H at 0.1 micromol/L-10 micromol/L inhibited the cellular proliferation and protein content of cultured myocardial cells in a dose-dependent manner. The inhibitory effects of U50,488H were completely blocked by pretreatment with nor-BNI, a specific kappa-opioid receptor antagonist at 1 micromol/L.

CONCLUSIONU50,488H inhibited the cultured myocardial cells' growth. The inhibitory effects of U50,488H are involved in mediating the action of kappa-opioid receptor stimulation.

3,4-Dichloro-N-methyl-N-(2-(1-pyrrolidinyl)-cyclohexyl)-benzeneacetamide, (trans)-Isomer ; pharmacology ; Animals ; Cell Growth Processes ; Cells, Cultured ; Myocytes, Cardiac ; metabolism ; Rats ; Rats, Sprague-Dawley ; Receptors, Opioid, kappa ; metabolism

To investigate the relaxation effect and underlying mechanism of U50,488H (a selective kappa-opioid receptor agonist) on aorta in the rat, isolated aortic ring was perfused and the tension of the vessel was measured. It was shown: (1) kappa-opioid receptor stimulation with U50,488H relaxed rat aorta dose-dependently; (2) the relaxation effect of U50,488H on aorta was partially endothelium-dependent; (3) the relaxation effect of U50,488H was significantly attenuated in the presence of glybenclamide and glipizide, two ATP-sensitive K(+) channel (K(ATP)) blockers; and (4) the relaxation effect of U50,488H on vessel bore no relationship to muscarinic-receptor, beta-adrenoceptor, prostaglandin and nitric oxide (NO). These results indicate that kappa-opioid receptor stimulation with U50,488H relaxes the aortic artery at least partially via K(ATP) channel in the rat.
3,4-Dichloro-N-methyl-N-(2-(1-pyrrolidinyl)-cyclohexyl)-benzeneacetamide, (trans)-Isomer ; pharmacology ; Animals ; Aorta ; physiology ; In Vitro Techniques ; KATP Channels ; metabolism ; Male ; Rats ; Rats, Sprague-Dawley ; Receptors, Opioid, kappa ; agonists ; physiology ; Vasodilation ; physiology