The natural axis of transmitter receptor distribution in the human cerebral cortex.

Transmitter receptors constitute a key component of the molecular machinery for intercellular communication in the brain. Recent efforts have mapped the density of diverse transmitter receptors across the human cerebral cortex with an unprecedented level of detail. Here, we distill these observations into key organizational principles. We demonstrate that receptor densities form a natural axis in the human cerebral cortex, reflecting decreases in differentiation at the level of laminar organization and a sensory-to-association axis at the functional level. Along this natural axis, key organizational principles are discerned: progressive molecular diversity (increase of the diversity of receptor density); excitation/inhibition (increase of the ratio of excitatory-to-inhibitory receptor density); and mirrored, orderly changes of the density of ionotropic and metabotropic receptors. The uncovered natural axis formed by the distribution of receptors aligns with the axis that is formed by other dimensions of cortical organization, such as the myelo- and cytoarchitectonic levels. Therefore, the uncovered natural axis constitutes a unifying organizational feature linking multiple dimensions of the cerebral cortex, thus bringing order to the heterogeneity of cortical organization.

Evidence for Dual Binding Sites for 1,1,1-Trichloro-2,2-bis(p-chlorophenyl)ethane (DDT) in Insect Sodium Channels.

1,1,1-Trichloro-2,2-bis(p-chlorophenyl)ethane (DDT), the first organochlorine insecticide, and pyrethroid insecticides are sodium channel agonists. Although the use of DDT is banned in most of the world due to its detrimental impact on the ecosystem, indoor residual spraying of DDT is still recommended for malaria control in Africa. Development of resistance to DDT and pyrethroids is a serious global obstacle for managing disease vectors. Mapping DDT binding sites is necessary for understanding mechanisms of resistance and modulation of sodium channels by structurally different ligands. The pioneering model of the housefly sodium channel visualized the first receptor for pyrethroids, PyR1, in the II/III domain interface and suggested that DDT binds within PyR1. Previously, we proposed the second pyrethroid receptor, PyR2, at the I/II domain interface. However, whether DDT binds to both pyrethroid receptor sites remains unknown. Here, using computational docking of DDT into the Kv1.2-based mosquito sodium channel model, we predict that two DDT molecules can bind simultaneously within PyR1 and PyR2. The bulky trichloromethyl group of each DDT molecule fits snugly between four helices in the bent domain interface, whereas two p-chlorophenyl rings extend into two wings of the interface. Model-driven mutagenesis and electrophysiological analysis confirmed these propositions and revealed 10 previously unknown DDT-sensing residues within PyR1 and PyR2. Our study proposes a dual DDT-receptor model and provides a structural background for rational development of new insecticides.

On the origin of the triplet puzzle of homologies in receptor heteromers: Toll-like receptor triplets in different types of receptors.

Based on our theory, we have discovered main triplets of amino acid residues in the GABAB1 receptor and several other neural receptors, which seem to originate from toll-like receptors and appear also as homologies in receptor heteromers. The obtained results strengthen our hypothesis that these triplets may 'guide-and-clasp' receptor-receptor interactions.

Stratification of the channel domain in neurotransmitter receptors.

Analyses of the ionic pore of ligand-gated ion channels at the amino acid level reveal a structural and functional stratification of the M2 channel domain. Mutations in the equatorial and outer regions affect channel gating, whereas mutations of other amino acid rings alter ionic permeability or selectivity.

Two chicken neuromedin U receptors: characterization of primary structure, biological activity and tissue distribution.

Neuromedin U (NMU) is a bioactive peptide that is involved in a variety of physiological functions. Two of its receptors, NMUR1 and NMUR2, have been identified and characterized in mammals. In this study, we performed cDNA cloning of chicken NMUR1 and NMUR2, and characterized their primary structure, biological activity, and expression patterns in chicken tissues. The chicken NMUR1 and NMUR2 cDNAs encoded 438 and 395 amino acid sequences, respectively. Chicken NMUR1 showed 54.8%-56.5% sequence identity with human, rat, and mouse NMUR1, and NMUR2 shared 67.3%-70.1% sequence identity with mammalian orthologs. Both chicken receptors have typical characteristics of G-protein-coupled receptors with seven transmembrane domains and the D/ERY motif. An increase in intracellular Ca(2+) mobilization was observed in HEK293 cells transfected with chicken NMUR1 or NMUR2 cDNA and treated with chicken or rat NMU. Real-time PCR analysis revealed that NMUR1 mRNA was preferentially expressed in the intestinal tissues such as the duodenum, jejunum, ileum, cecum, and colon/rectum, and brain regions such as the midbrain and optic lobe, and the ovary in adult hens. NMUR2 mRNA was exclusively expressed in the brain regions such as the cerebrum and midbrain. These results indicate that NMUR1 and NMUR2 mRNAs, which encode functional receptor proteins, are expressed in chicken tissues with different distribution patterns.

[Architecture of receptor-operated ionic channels of biological membranes].

Ion channels of biological membranes are the key proteins, which provide bioelectric functioning of living systems. These proteins are homo- or heterooligomers assembled from several identical or different subunits. Understanding the architectural organization and functioning of ion channels has been significantly extended due to resolving the crystal structure of several types of voltage-gated and receptor-operated channels. This review summarizes the information obtained from crystal structures of potassium, nicotinic acetylcholine receptor, P2X, and other ligand-gated ion channels. Despite the differences in the function, topology, ionic selectivity, and the subunit stoichiometry, a high similarity in the principles of organization of these macromolecular complexes has been revealed.

Atomic structure and dynamics of pentameric ligand-gated ion channels: new insight from bacterial homologues.

Pentameric ligand-gated ion channels (pLGICs) are widely expressed in the animal kingdom and are key players of neurotransmission by acetylcholine (ACh), gamma-amminobutyric acid (GABA), glycine and serotonin. It is now established that this family has a prokaryotic origin, since more than 20 homologues have been discovered in bacteria. In particular, the GLIC homologue displays a ligand-gated ion channel function and is activated by protons. The prokaryotic origin of these membrane proteins facilitated the X-ray structural resolution of the first members of this family. ELIC was solved at 3.3 A in a closed-pore conformation, and GLIC at up to 2.9 A in an apparently open-pore conformation. These data reveal many structural features, notably the architecture of the pore, including its gate and its selectivity filter, and the interactions between the protein and lipids. In addition, comparison of the structures of GLIC and ELIC hints at a mechanism of channel opening, which consists of both a quaternary twist and a tertiary deformation. This mechanism couples opening-closing motions of the channel with a global reorganization of the protein, including the subunit interface that holds the neurotransmitter binding sites in eukaryotic pLGICs.

The subunit arrangement and assembly of ionotropic receptors.

Ionotropic receptors mediate rapid communication between neurons. These receptors are oligomers and are usually assembled from multiple subunit types. Receptors built from different subunit combinations have distinct functional properties, such as single-channel conductances, rates of desensitization and sensitivities to activators and inactivators; they can also have different intracellular locations. Methods are now available for determining not only the subunit stoichiometry but also the subunit arrangement within ionotropic receptors. This information will inform experiments designed to understand the molecular basis of receptor assembly and function. It will also permit the modelling of potential ligand-binding sites at the interfaces between the subunits and should lead to a more rational approach to drug development.

Synthetic polyamines: an overview of their multiple biological activities.

The binding of polyamines to a variety of receptors and other defined recognition sites has been widely reported. It is well-known that polyamines interact with aspartate, glutamate, and aromatic residues of a given receptor and/or enzyme mainly through the formation of ion bonds, since at physiological pH, protonation of amino groups is nearly complete. From this, the hypothesis arises that a polyamine may be a universal template able to recognize different receptor systems. This hypothesis suggests that both affinity and selectivity may be fine-tuned by inserting appropriate substituents onto the amine functions and by varying the methylene chain lengths between them on the polyamine backbone. In this paper, we detail several application of this design strategy aimed at discovering potent and selective polyamines able to bind neurotransmitter receptors and enzymes, such as muscarinic receptor subtypes, muscle-type nicotinic receptors and acethylcholinesterase.

Trk: a neuromodulator of age-specific behavioral and neurochemical responses to cocaine in mice.

Responses to psychostimulants vary with age, but the molecular etiologies of these differences are largely unknown. The goal of the present research was to identify age-specific behavioral and molecular adaptations to cocaine and to elucidate the mechanisms involved therein. Postweanling, periadolescent, and adult male CD-1 mice were exposed to cocaine (20 mg/kg) for 7 d. The rewarding effects of cocaine were assessed, as were the response to a Trk antagonist and the regulation of dopamine and cAMP-regulated phosphoprotein, 32 kDa (DARPP-32). Cocaine was rewarding in both periadolescent and adult mice using a conditioned place preference procedure. In contrast, postweanling mice failed to demonstrate significant cocaine-induced place preference. Because components of the neurotrophin system including brain-derived neurotrophic factor and TrkB are developmentally regulated, their role in the age-specific effects of cocaine was determined using the Trk receptor antagonist K252a. Postweanling mice that received K252a before daily cocaine showed a significant place preference to the cocaine-paired environment that was not seen in the absence of K252a. DARPP-32 protein levels were significantly upregulated in the lateral region of the caudate-putamen exclusively in postweanling mice after chronic cocaine. Daily pretreatment with K252a attenuated the induction of DARPP-32 in the postweanling striatum. These data indicate that Trk neurotransmission plays a role in age-specific behavioral and molecular responses to cocaine and concurrently modulates DARPP-32 levels.

Nicotinic receptors, allosteric proteins and medicine.

The nicotinic acetylcholine receptor (nAChR) was the first ion channel and membrane receptor of a neurotransmitter to be isolated and chemically identified and is one of the best known membrane proteins involved in signal transduction. Subsequently, nAChRs have been a target for drug discovery because of their potential to impact numerous brain diseases and disorders. Here, we consider recent developments in our understanding of nAChR structure and of the conformational transitions that link the acetylcholine (ACh)-binding site and the ion channel to mediate fast neurotransmission. The knowledge of such allosteric mechanisms is essential to understand pathologies such as congenital myasthenia, autosomal dominant nocturnal frontal lobe epilepsies, sudden infant death syndrome, attention deficit hyperactivity disorder and nicotine addiction and to design novel therapies.

Subunit identification and reconstitution of the N-type Ca2+ channel complex purified from brain.

Calcium channels play an important role in regulating various neuronal processes, including synaptic transmission and cellular plasticity. The N-type calcium channels, which are sensitive to omega-conotoxin, are involved in the control of transmitter release from neurons. A functional N-type calcium channel complex was purified from rabbit brain. The channel consists of a 230-kilodalton subunit (alpha 1B) that is tightly associated with a 160-kilodalton subunit (alpha 2 delta), a 57-kilodalton subunit (beta 3), and a 95-kilodalton glycoprotein subunit. The complex formed a functional calcium channel with the same pharmacological properties and conductance as those of the native omega-conotoxin-sensitive calcium channel in neurons.

Molecular diversity of glutamate receptors and implications for brain function.

The glutamate receptors mediate excitatory neurotransmission in the brain and are important in memory acquisition, learning, and some neurodegenerative disorders. This receptor family is classified in three groups: the N-methyl-D-aspartate (NMDA), alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionate (AMPA)-kainate, and metabotropic receptors. Recent molecular studies have shown that many receptor subtypes exist in all three groups of the receptors and exhibit heterogeneity in function and expression patterns. This article reviews the molecular and functional diversity of the glutamate receptors and discusses their implications for integrative brain function.

Ca2+ permeability of KA-AMPA--gated glutamate receptor channels depends on subunit composition.

NMDA (N-methyl-D-aspartate) receptors and non-NMDA receptors represent the two major classes of ion channel-linked glutamate receptors. Unlike the NMDA receptor channels, non-NMDA receptor channels have usually been thought to conduct monovalent cations only. Non-NMDA receptor ion channels that can be gated by kainic acid (KA) and alpha-amino-3-hydroxy-5-methyl-4-isoxazole propionic acid (AMPA) are formed by the glutamate receptor subunits GluR1, GluR2, and GluR3. These subunits were expressed in various combinations in Xenopus oocytes so that their permeability to divalent cations could be studied. At physiological resting potentials, KA and AMPA elicited inward calcium currents in oocytes expressing GluR1, GluR3, and GluR1 plus GluR3. In contrast, oocytes expressing GluR1 plus GluR2 or GluR3 plus GluR2 showed no such permeability. Thus, in neurons expressing certain KA-AMPA receptor subunits, glutamate may trigger calcium-dependent intracellular events by activating non-NMDA receptors.

Cloning, expression, and gene structure of a G protein-coupled glutamate receptor from rat brain.

A complementary DNA encoding a G protein-coupled glutamate receptor from rat brain, GluGR, was cloned by functional expression in Xenopus oocytes. The complementary DNA encodes a protein of 1199 amino acids containing a seven-transmembrane motif, flanked by large amino- and carboxyl-terminal domains. This receptor lacks any amino acid sequence similarity with other G protein-coupled receptors, suggesting that it may be a member of a new subfamily. The presence of two introns flanking the central core suggests that GluGR may have evolved by exon shuffling. Expressed in oocytes, GluGR is activated by quisqualate greater than glutamate greater than ibotenate greater than trans-1-aminocyclopentyl-1,3-dicarboxylate, and it is inhibited by 2-amino-3-phosphonopropionate. Activation is blocked by Bordella pertussis toxin. These properties are typical of some metabotropic glutamate receptors.

Signaling specificity by Frizzled receptors in Drosophila.

Wnt-Frizzled (Fz) signaling pathways play recurring important roles during the development and homeostasis of vertebrates and invertebrates. Fz receptors can signal through beta-catenin-dependent and -independent pathways. In Drosophila, Fz and Fz2 are redundant receptors for Wg. In addition, Fz conveys signals through a distinct pathway to organize planar polarization of epithelial structures. We demonstrate that the cytoplasmic sequences of Fz2 and Fz preferentially activate the beta-catenin and planar polarity cascade, respectively. Both receptors activate either pathway, but with different efficiencies. Intrinsic differences in signaling efficiency in closely related receptors might be a general mechanism for generating signaling specificity in vivo.

G protein signaling from activated rat frizzled-1 to the beta-catenin-Lef-Tcf pathway.

The frizzled receptors, which mediate development and display seven hydrophobic, membrane-spanning segments, are cell membrane-localized. We constructed a chimeric receptor with the ligand-binding and transmembrane segments from the beta2-adrenergic receptor (beta2AR) and the cytoplasmic domains from rat Frizzled-1 (Rfz1). Stimulation of mouse F9 clones expressing the chimera (beta2AR-Rfz1) with the beta-adrenergic agonist isoproterenol stimulated stabilization of beta-catenin, activation of a beta-catenin-sensitive promoter, and formation of primitive endoderm. The response was blocked by inactivation of pertussis toxin-sensitive, heterotrimeric guanine nucleotide-binding proteins (G proteins) and by depletion of Galphaq and Galphao. Thus, G proteins are elements of Wnt/Frizzled-1 signaling to the beta-catenin-lymphoid-enhancer factor (LEF)-T cell factor (Tcf) pathway.

Molecular modeling of neurological membrane proteins - from binding sites to synapses.

The field of molecular mechanics studies of proteins has developed enormously since its origin in the 1970's, and many applications and methodologies have branched from the original idea of the force field. The applications of such methodologies are far spread and commonplace in neuroscience research today. In this mini-review, we outline the main methodologies applied when studying events ranging from ligands binding within small binding sites, through overall large-scale conformational changes, to the even larger-scale oligomerization events of neurological membrane proteins. The limitations and caveats of the methods are discussed, while examples of recent applications are described and their implications discussed. We have chosen to focus on the monoamine transporters throughout, with a few examples from neurological membrane proteins such as ionotropic and metabotropic neurotransmitter receptors.

Primary structure and expression of the gamma 2 subunit of the glutamate receptor channel selective for kainate.

The presence and primary structure of a novel subunit of the mouse glutamate receptor channel, designated as gamma 2, have been revealed by cloning and sequencing the cDNA. The gamma 2 subunit has structural characteristics common to the neurotransmitter-gated ion channel family and shares a high amino acid sequence identity with the rat KA-1 subunit, thus constituting the gamma subfamily of the glutamate receptor channel. Expression of the gamma 2 subunit together with the beta 2 subunit in Xenopus oocytes yields functional glutamate receptor channels selective for kainate.

Distinct agonist- and antagonist-binding sites on the glycine receptor.

The distinction between receptor-binding sites for agonists and antagonists underpins the pharmacological differences between these two classes of ligands. In the glycine receptor, antagonist (strychnine) binding requires an interaction with residues Lys-200 and Tyr-202. We now demonstrate that the agonist-binding site of this receptor is located at the residue Thr-204. The agonist-binding site interaction is thus likely to be mediated by hydrogen bonding and not by ionic interactions. Our results demonstrate that, in contrast to other studies of ligand-gated ion channel receptors, agonist- and antagonist-binding sites are composed of distinct amino acid residues.