SR9883 is a novel small-molecule enhancer of α4β2* nicotinic acetylcholine receptor signaling that decreases intravenous nicotine self-administration in rats.

Background: Most smokers attempting to quit will quickly relapse to tobacco use even when treated with the most efficacious smoking cessation agents currently available. This highlights the need to develop effective new smoking cessation medications. Evidence suggests that positive allosteric modulators (PAM) and other enhancers of nicotinic acetylcholine receptor (nAChR) signaling could have therapeutic utility as smoking cessation agents.

A new synthetic medium for the optimization of docosahexaenoic acid production in Crypthecodinium cohnii.

The heterotrophic microalgae Crypthecodinium cohnii was usually cultivated in complex medium containing glucose, yeast extract and sea salt. For the preparation of DHA with highest purity, a new defined medium without the yeast extract was developed. Different inoculated densities, C/N ratios, temperatures, culture volumes and glucose additions were investigated to optimize the algal growth rate and DHA production.

Biosynthesis of uniformly carbon isotope-labeled docosahexaenoic acid in Crypthecodinium cohnii.

Docosahexaenoic acid (DHA) enriched in brain can yield many important degradation products after the attack of hydroxyl radicals, which is known to serve as a nutraceutical and neuroprotective effects. Oxidative stress is a commonly observed feature of Alzheimer's disease (AD). Therefore, uniformly radiolabeled DHA plays an important role in studying the oxidative fate of DHA in vivo and vitro.

Promoting activity of (α4)(β2) nicotinic cholinergic receptors reduces ethanol consumption.

There is increasing interest in developing drugs that act at α4β2 nicotinic acetylcholine receptors (nAChRs) to treat alcohol use disorder. The smoking cessation agent varenicline, a partial agonist of α4β2 nAChRs, reduces alcohol intake, but its use can be limited by side effects at high therapeutic doses. There are two stoichiometric forms of α4β2 nAChRs, (α4)(β2) and (α4)(β2).

Discovery of an intrasubunit nicotinic acetylcholine receptor-binding site for the positive allosteric modulator Br-PBTC.

Nicotinic acetylcholine receptor (nAChR) ligands that lack agonist activity but enhance activation in the presence of an agonist are called positive allosteric modulators (PAMs). nAChR PAMs have therapeutic potential for the treatment of nicotine addiction and several neuropsychiatric disorders. PAMs need to be selectively targeted toward certain nAChR subtypes to tap this potential.

Unorthodox Acetylcholine Binding Sites Formed by α5 and β3 Accessory Subunits in α4β2* Nicotinic Acetylcholine Receptors.

All nicotinic acetylcholine receptors (nAChRs) evolved from homomeric nAChRs in which all five subunits are involved in forming acetylcholine (ACh) binding sites at their interfaces. Heteromeric α4β2* nAChRs typically have two ACh binding sites at α4/β2 interfaces and a fifth accessory subunit surrounding the central cation channel. β2 accessory subunits do not form ACh binding sites, but α4 accessory subunits do at the α4/α4 interface in (α4β2)α4 nAChRs.

A Novel α2/α4 Subtype-selective Positive Allosteric Modulator of Nicotinic Acetylcholine Receptors Acting from the C-tail of an α Subunit.

Positive allosteric modulators (PAMs) of nicotinic acetylcholine receptors (nAChR) are important therapeutic candidates as well as valuable research tools. We identified a novel type II PAM, (R)-7-bromo-N-(piperidin-3-yl)benzo[b]thiophene-2-carboxamide (Br-PBTC), which both increases activation and reactivates desensitized nAChRs. This compound increases acetylcholine-evoked responses of α2* and α4* nAChRs but is without effect on α3* or α6* nAChRs (* indicates the presence of other nAChR subunits).

An Accessory Agonist Binding Site Promotes Activation of α4β2* Nicotinic Acetylcholine Receptors.

Neuronal nicotinic acetylcholine receptors containing α4, β2, and sometimes other subunits (α4β2* nAChRs) regulate addictive and other behavioral effects of nicotine. These nAChRs exist in several stoichiometries, typically with two high affinity acetylcholine (ACh) binding sites at the interface of α4 and β2 subunits and a fifth accessory subunit. A third low affinity ACh binding site is formed when this accessory subunit is α4 but not if it is β2.

Nicotinic receptors in non-human primates: Analysis of genetic and functional conservation with humans.

Nicotinic acetylcholine receptors (nAChRs) are highly conserved between humans and non-human primates. Conservation exists at the level of genomic structure, protein structure and epigenetics. Overall homology of nAChRs at the protein level is 98% in macaques versus 89% in mice, which is highly relevant for evaluating subtype-specific ligands that have different affinities in humans versus rodents.

Expression of cloned α6* nicotinic acetylcholine receptors.

Nicotinic acetylcholine receptors (AChRs) are ACh-gated ion channels formed from five homologous subunits in subtypes defined by their subunit composition and stoichiometry. Some subtypes readily produce functional AChRs in Xenopus oocytes and transfected cell lines. α6β2β3* AChRs (subtypes formed from these subunits and perhaps others) are not easily expressed.

Efficient expression of functional (α6β2)2β3 AChRs in Xenopus oocytes from free subunits using slightly modified α6 subunits.

Human (α6β2)(α4β2)β3 nicotinic acetylcholine receptors (AChRs) are essential for addiction to nicotine and a target for drug development for smoking cessation. Expressing this complex AChR is difficult, but has been achieved using subunit concatamers. In order to determine what limits expression of α6* AChRs and to efficiently express α6* AChRs using free subunits, we investigated expression of the simpler (α6β2)2β3 AChR.

Acute activation, desensitization and smoldering activation of human acetylcholine receptors.

The behavioral effects of nicotine and other nicotinic agonists are mediated by AChRs in the brain. The relative contribution of acute activation versus chronic desensitization of AChRs is unknown. Sustained "smoldering activation" occurs over a range of agonist concentrations at which activated and desensitized AChRs are present in equilibrium.

Chemical chaperones exceed the chaperone effects of RIC-3 in promoting assembly of functional α7 AChRs.

Functional α7 nicotinic acetylcholine receptors (AChRs) do not assemble efficiently in cells transfected with α7 subunits unless the cells are also transfected with the chaperone protein RIC-3. Despite the presence of RIC-3, large amounts of these subunits remain improperly assembled. Thus, additional chaperone proteins are probably required for efficient assembly of α7 AChRs.

Acetylcholine receptor (AChR) α5 subunit variant associated with risk for nicotine dependence and lung cancer reduces (α4β2)₂α5 AChR function.

Genomic studies have identified a D398N variation in the α5 subunit of nicotinic acetylcholine receptors (AChRs) that increases risk of nicotine dependence and lung cancer. (α4β2)₂α5 AChRs are a significant brain presynaptic subtype in brain. Their high sensitivity to activation by nicotine and high Ca²+ permeability give them substantial functional impact.

Specific immunotherapy of experimental myasthenia gravis by a novel mechanism.

Objective: Myasthenia gravis (MG) and its animal model, experimental autoimmune myasthenia gravis (EAMG), are antibody (Ab)-mediated autoimmune diseases, in which autoantibodies bind to and cause loss of muscle nicotinic acetylcholine receptors (AChRs) at the neuromuscular junction. To develop a specific immunotherapy of MG, we treated rats with ongoing EAMG by intraperitoneal injection of bacterially-expressed human muscle AChR constructs.

Methods: Rats with ongoing EAMG received intraperitoneal treatment with the constructs weekly for 5 weeks beginning after the acute phase.

Mutations of cytosolic loop residues impair assembly and maturation of alpha7 nicotinic acetylcholine receptors.

Mechanisms that regulate early events in the biogenesis of the alpha7 nicotinic acetylcholine receptor (alpha7 AChR) are not well understood. Data presented here show that single amino acid mutations in the cytoplasmic loop of the alpha7 AChR, between position 335 and 343, abolish or attenuate expression of mature pentameric alpha7 AChRs in both human embryonic kidney tsA201 (HEK) and neuronal SH-SY5Y cells. Although the number of mature alpha7 AChRs is increased significantly in the presence of the chaperone protein resistant to inhibitors of cholineesterase-3 in HEK cells, sucrose gradient sedimentation reveals that the vast majority of alpha7 subunits are aggregated or improperly assembled.

Myasthenia gravis and the tops and bottoms of AChRs: antigenic structure of the MIR and specific immunosuppression of EAMG using AChR cytoplasmic domains.

The main immunogenic region (MIR), against which half or more of the autoantibodies to acetylcholine receptors (AChRs) in myasthenia gravis (MG) or experimental autoimmune MG (EAMG) are directed, is located at the extracellular end of alpha1 subunits. Rat monoclonal antibodies (mAbs) to the MIR efficiently compete with MG patient autoantibodies for binding to human muscle AChRs. Antibodies bound to the MIR do not interfere with cholinergic ligand binding or AChR function, but target complement and trigger antigenic modulation.

Beta3 subunits promote expression and nicotine-induced up-regulation of human nicotinic alpha6* nicotinic acetylcholine receptors expressed in transfected cell lines.

Nicotinic acetylcholine receptors (AChRs) containing alpha6 subunits are typically found at aminergic nerve endings where they play important roles in nicotine addiction and Parkinson's disease. alpha6* AChRs usually contain beta3 subunits. beta3 subunits are presumed to assemble only in the accessory subunit position within AChRs where they do not participate in forming acetylcholine binding sites.

Analogs of alpha-conotoxin MII are selective for alpha6-containing nicotinic acetylcholine receptors.

Neuronal nicotinic acetylcholine receptors (nAChRs) both mediate direct cholinergic synaptic transmission and modulate synaptic transmission by other neurotransmitters. Novel ligands are needed as probes to discriminate among structurally related nAChR subtypes. Alpha-conotoxin MII, a selective ligand that discriminates among a variety of nAChR subtypes, fails to discriminate well between some subtypes containing the closely related alpha3 and alpha6 subunits.

Human alpha4beta2 acetylcholine receptors formed from linked subunits.

We prepared concatamers of alpha4 and beta2 subunits for human nicotinic acetylcholine receptors (AChRs), in which the C terminus of alpha4 was linked to the N terminus of beta2, or vice versa, via a tripeptide sequence repeated 6 or 12 times, and expressed them in Xenopus oocytes. Linkage did not substantially alter channel amplitude or channel open-duration. Linkage at the C terminus of alpha4 prevented AChR potentiation by 17-beta-estradiol by disruption of its binding site.

Alpha-conotoxin PIA is selective for alpha6 subunit-containing nicotinic acetylcholine receptors.

Until now, there have been no antagonists to discriminate between heteromeric nicotinic acetylcholine receptors (nAChRs) containing the very closely related alpha6 and alpha3 subunits. nAChRs containing alpha3, alpha4, or alpha6 subunits in combination with beta2, occasionally beta4, and sometimes beta3 or alpha5 subunits, are thought to play important roles in cognitive function, pain perception, and the reinforcing properties of nicotine. We cloned a novel gene from the predatory marine snail Conus purpurascens.

Alternate stoichiometries of alpha4beta2 nicotinic acetylcholine receptors.

Two functional types of nicotinic acetylcholine receptors (nAChRs) are expressed when human embryonic kidney cells are permanently transfected with equal amounts of human alpha4 and beta2 subunit cDNAs. Most (82%) of these nAChRs exhibit an EC(50) of 74 +/- 6 microM for ACh, a much lower sensitivity than the remaining fraction (EC(50) of 0.7 +/- 0.

Alcohol modulation of neuronal nicotinic acetylcholine receptors is alpha subunit dependent.

Background: We have previously shown that n-alcohols exert a dual action on the alpha4beta2-type neuronal nicotinic acetylcholine (ACh) receptors (AChRs), with shorter-chain alcohols potentiating and longer-chain alcohols inhibiting ACh-induced currents. Ethanol potentiates the current in alpha4beta2 receptors, yet it has little or no effect on the alpha3beta2 receptors. Because the alpha4 AChRs are present predominantly in the brain, whereas the alpha3 AChRs are present predominantly in the peripheral ganglia, the differential action of ethanol on the alpha4beta2 and alpha3beta2 AChRs may contribute to its differential effects on the brain and the peripheral nervous system.

First and second transmembrane segments of alpha3, alpha4, beta2, and beta4 nicotinic acetylcholine receptor subunits influence the efficacy and potency of nicotine.

The first three transmembrane segments (M1-M3) of human nicotinic acetylcholine receptors (nAChRs) have been implicated in determining the efficacy of nicotine by studies of alpha3/alpha4 subunit chimeras. Nicotine has full efficacy on the alpha4beta2 nAChR and partial efficacy on the alpha3beta2 nAChR. Now, we have exchanged individually three amino acids between the alpha4 and the alpha3 subunits at positions 226(M1), 258(M2), and 262(M2).