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Inhibitory effects of tramadol on nicotinic acetylcholine receptors in adrenal chromaffin cells and in Xenopus oocytes expressing alpha 7 receptors.

ABSTRACT: 1. Tramadol has been used clinically as an analgesic; however, the mechanism of its analgesic effects is still unknown. 2. We used bovine adrenal chromaffin cells to investigate effects of tramadol on catecholamine secretion, nicotine-induced cytosolic Ca(2+) concentration ([Ca(2+)](i)) increases and membrane current changes. We also investigated effects of tramadol on alpha7 nicotinic acetylcholine receptors (AChRs) expressed in Xenopus oocytes. 3. Tramadol concentration-dependently suppressed carbachol-induced catecholamine secretion to 60% and 27% of the control at the concentration of 10 and 100 microM, respectively, whereas it had little effect on veratridine- or high K(+)-induced catecholamine secretion. 4. Tramadol also suppressed nicotine-induced ([Ca(2+)](i)) increases in a concentration-dependent manner. Tramadol inhibited nicotine-induced inward currents, and the inhibition was unaffected by the opioid receptor antagonist naloxone. 5. Tramadol inhibited nicotinic currents carried by alpha7 receptors expressed in Xenopus oocytes. 6. Tramadol inhibited both alpha-bungarotoxin-sensitive and -insensitive nicotinic currents in bovine adrenal chromaffin cells. 7. In conclusion, tramadol inhibits catecholamine secretion partly by inhibiting nicotinic AChR functions in a naloxone-insensitive manner and alpha7 receptors are one of those inhibited by tramadol.

SUBMITTER: Shiraishi M

PROVIDER: S-EPMC1573343 | biostudies-other | 2002 May

REPOSITORIES: biostudies-other

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Monkey adrenal chromaffin cells express ?6?4* nicotinic acetylcholine receptors.

Project description:Nicotinic acetylcholine receptors (nAChRs) that contain ?6 and ?4 subunits have been demonstrated functionally in human adrenal chromaffin cells, rat dorsal root ganglion neurons, and on noradrenergic terminals in the hippocampus of adolescent mice. In human adrenal chromaffin cells, ?6?4* nAChRs (the asterisk denotes the possible presence of additional subunits) are the predominant subtype whereas in rodents, the predominant nAChR is the ?3?4* subtype. Here we present molecular and pharmacological evidence that chromaffin cells from monkey (Macaca mulatta) also express ?6?4* receptors. PCR was used to show the presence of transcripts for ?6 and ?4 subunits and pharmacological characterization was performed using patch-clamp electrophysiology in combination with ?-conotoxins that target the ?6?4* subtype. Acetylcholine-evoked currents were sensitive to inhibition by BuIA[T5A,P6O] and MII[H9A,L15A]; ?-conotoxins that inhibit ?6-containing nAChRs. Two additional agonists were used to probe for the expression of ?7 and ?2-containing nAChRs. Cells with currents evoked by acetylcholine were relatively unresponsive to the ?7-selctive agonist choline but responded to the agonist 5-I-A-85380. These studies provide further insights into the properties of natively expressed ?6?4* nAChRs.

| S-EPMC3984115 | biostudies-literature

Expression of α3β2β4 nicotinic acetylcholine receptors by rat adrenal chromaffin cells determined using novel conopeptide antagonists.

Project description:Adrenal chromaffin cells release neurotransmitters in response to stress and may be involved in conditions such as post-traumatic stress and anxiety disorders. Neurotransmitter release is triggered, in part, by activation of nicotinic acetylcholine receptors (nAChRs). However, despite decades of use as a model system for studying exocytosis, the nAChR subtypes involved have not been pharmacologically identified. Quantitative real-time PCR of rat adrenal medulla revealed an abundance of mRNAs for α3, α7, β2, and β4 subunits. Whole-cell patch-clamp electrophysiology of chromaffin cells and subtype-selective ligands were used to probe for nAChRs derived from the mRNAs found in adrenal medulla. A novel conopeptide antagonist, PeIA-5469, was created that is highly selective for α3β2 over other nAChR subtypes heterologously expressed in Xenopus laevis oocytes. Experiments using PeIA-5469 and the α3β4-selective α-conotoxin TxID revealed that rat adrenal medulla contain two populations of chromaffin cells that express either α3β4 nAChRs alone or α3β4 together with the α3β2β4 subtype. Conclusions were derived from observations that acetylcholine-gated currents in some cells were sensitive to inhibition by PeIA-5469 and TxID, while in other cells, currents were sensitive only to TxID. Expression of functional α7 nAChRs was determined using three α7-selective ligands: the agonist PNU282987, the positive allosteric modulator PNU120596, and the antagonist α-conotoxin [V11L,V16D]ArIB. The results of these studies identify for the first time the expression of α3β2β4 nAChRs as well as functional α7 nAChRs by rat adrenal chromaffin cells.

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Ketamine and its preservative, benzethonium chloride, both inhibit human recombinant alpha7 and alpha4beta2 neuronal nicotinic acetylcholine receptors in Xenopus oocytes.

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Native ?6?4* nicotinic receptors control exocytosis in human chromaffin cells of the adrenal gland.

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| S-EPMC3250250 | biostudies-literature

α-Conotoxins Identify the α3β4* Subtype as the Predominant Nicotinic Acetylcholine Receptor Expressed in Human Adrenal Chromaffin Cells.

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| S-EPMC4613940 | biostudies-literature

Diverse inhibitory actions of quaternary ammonium cholinesterase inhibitors on Torpedo nicotinic ACh receptors transplanted to Xenopus oocytes.

Project description:BACKGROUND AND PURPOSE: This work was aimed at comparing and analysing the effects and mechanisms of action of the quaternary ammonium cholinesterase inhibitors (QChEIs) BW284c51, decamethonium and edrophonium, on nicotinic ACh receptor (nAChR) function. EXPERIMENTAL APPROACH: nAChRs purified from Torpedo electroplax were transplanted to oocytes and currents elicited by ACh (I(ACh)) either alone or in presence of these QChEIs were recorded. KEY RESULTS: None of the QChEIs, by itself, elicited changes in membrane conductance; however, when co-applied with ACh, all of them decreased I(ACh) in a concentration-dependent way. The mechanisms of nAChR inhibition were different for these QChEIs. BW284c51 blockade was non-competitive and voltage-dependent, although it also affected the n(H) of the dose-response curve. By contrast, decamethonium and edrophonium inhibition, at -60 mV, was apparently competitive and did not modify either desensitisation or n(H). Decamethonium effects were voltage-independent and washed out slowly after its removal; by contrast, edrophonium blockade had strong voltage dependence and its effects disappeared quickly after its withdrawal. Analysis of the voltage-dependent blockade indicated that BW284c51 bound to a shallow site into the channel pore, whereas edrophonium bound to a deeper locus. Accordingly, additive inhibitory effects on I(ACh) were found among any pairs of these QChEIs. CONCLUSIONS AND IMPLICATIONS: The tested QChEIs bound to the nAChR at several and different loci, which might account for their complex inhibitory behaviour, acting both as allosteric effectors and, in the case of BW284c51 and edrophonium, as open channel blockers.

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Formation of functional alpha3beta4alpha5 human neuronal nicotinic receptors in Xenopus oocytes: a reporter mutation approach.

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| S-EPMC1573006 | biostudies-literature

Palmitoylation of nicotinic acetylcholine receptors.

Project description:It is well established that nicotinic acetylcholine receptors (nAChRs) undergo a number of different posttranslational modifications, such as disulfide bond formation, glycosylation, and phosphorylation. Recently, our laboratory has developed more sensitive assays of protein palmitoylation that have allowed us and others to detect the palmitoylation of relatively low abundant proteins such as ligand-gated ion channels. Here, we present evidence that palmitoylation is prevalent on many subunits of different nAChR subtypes, both muscle-type nAChRs and the neuronal "alpha(4)beta(2)" and "alpha(7)" subtypes most abundant in brain. The loss of ligand binding sites that occurs when palmitoylation is blocked with the inhibitor bromopalmitate suggests that palmitoylation of alpha(4)beta(2) and alpha(7) subtypes occurs during subunit assembly and regulates the formation of ligand binding sites. However, additional experiments are needed to test whether nAChR subunit palmitoylation is involved in other aspects of nAChR trafficking or whether palmitoylation regulates nAChR function. Further investigation would be aided by identifying the sites of palmitoylation on the subunits, and here we propose a mass spectrometry strategy for identification of these sites.

| S-EPMC3523180 | biostudies-literature

Nicotinic acetylcholine receptors and cancer.

Project description:Nicotine, the primary addictive constituent of cigarettes, is believed to contribute to cancer promotion and progression through the activation of nicotinic acetylcholine receptors (nAChRs), which are membrane ligand-gated cation channels. nAChRs activation can be triggered by the neurotransmitter Ach, or certain other biological compounds, such as nicotine. In recent years, genome-wide association studies have indicated that allelic variation in the α5-α3-β4 nAChR cluster on chromosome 15q24-15q25.1 is associated with lung cancer risk. The role of nAChRs in other types of cancer has also been reported. The present review highlights the role of nAChRs in types of human cancer.

| S-EPMC4840641 | biostudies-literature

Functional reconstitution of Haemonchus contortus acetylcholine receptors in Xenopus oocytes provides mechanistic insights into levamisole resistance.

Project description:BACKGROUND AND PURPOSE: The cholinergic agonist levamisole is widely used to treat parasitic nematode infestations. This anthelmintic drug paralyses worms by activating a class of levamisole-sensitive acetylcholine receptors (L-AChRs) expressed in nematode muscle cells. However, levamisole efficacy has been compromised by the emergence of drug-resistant parasites, especially in gastrointestinal nematodes such as Haemonchus contortus. We report here the first functional reconstitution and pharmacological characterization of H. contortus L-AChRs in a heterologous expression system. EXPERIMENTAL APPROACH: In the free-living nematode Caenorhabditis elegans, five AChR subunit and three ancillary protein genes are necessary in vivo and in vitro to synthesize L-AChRs. We have cloned the H. contortus orthologues of these genes and expressed them in Xenopus oocytes. We reconstituted two types of H. contortus L-AChRs with distinct pharmacologies by combining different receptor subunits. KEY RESULTS: The Hco-ACR-8 subunit plays a pivotal role in selective sensitivity to levamisole. As observed with C. elegans L-AChRs, expression of H. contortus receptors requires the ancillary proteins Hco-RIC-3, Hco-UNC-50 and Hco-UNC-74. Using this experimental system, we demonstrated that a truncated Hco-UNC-63 L-AChR subunit, which was specifically detected in a levamisole-resistant H. contortus isolate, but not in levamisole-sensitive strains, hampers the normal function of L-AChRs, when co-expressed with its full-length counterpart. CONCLUSIONS AND IMPLICATIONS: We provide the first functional evidence for a putative molecular mechanism involved in levamisole resistance in any parasitic nematode. This expression system will provide a means to analyse molecular polymorphisms associated with drug resistance at the electrophysiological level.

| S-EPMC3221097 | biostudies-literature

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