Dopamine Binding Research Articles

Sensing ultra-trace dopamine by restoration of fluorescence on locally acidified gold nanoparticles.

An ultra-sensitive sensor was fabricated to measure dopamine through quenching and restoring FITC fluorescence by the competitive binding of dopamine and N-acetylneuraminic acid with mercaptophenylboronic acid anchored on the gold nanoparticles. Assessed by quantifying dopamine in human urine samples, it reached a limit of detection of as low as 50 pM dopamine over a linear range from 10-10 M to 10-7 M. It also features low technical barriers and operation cost, and the strategy proposed could be extendable to other analytes and not restricted to the gold nanoparticles.

M68 - THE RELATIONSHIP BETWEEN FUNCTIONAL DOPAMINE D2 RECEPTOR HAPLOTYPES AND COGNITIVE OUTCOMES, AS MEDIATED BY REGIONAL BRAIN STRUCTURE: A COMPARISON OF CONTROL, AT-RISK, AND BIPOLAR DISORDER SUBJECTS

Background Bipolar Disorder (BD) is a complex and highly heritable mood disorder associated with a range of cognitive deficits across all illness states. Previous research has demonstrated associations between the dopamine D2 receptor gene (DRD2) and BD diagnosis, structural and functional brain changes, and cognitive dysfunction. Specific SNPs impacting DRD2 function have been identified: rs1800497 (Taq1A1, minor T allele) is associated with reduced striatal D2R dopamine binding; and rs1076560 and rs2283265 (minor A/T alleles) are associated with alternative splicing of DRD2 exon 6, resulting in increased expression of DRD2-short mRNA compared to DRD2-long isoform, and reduced inhibition of glutamate release. The present study adopted a cognitive-neurogenic approach to explore the mediating role of brain structure in the relationship between DRD2 functional haplotypes and cognitive outcomes. Methods Participants comprised 26 BD cases (BD-I or BD-II), 62 at-risk subjects (unaffected first-degree relatives of BD probands), and 55 control subjects with 3 T MRI, genetic and cognitive data. DRD2 haplotypes were defined by three functional SNPs (rs1800497, rs1076560, rs2283265). Cortical thickness, surface area and volume were extracted for three functionally defined regions of interest (ROIs) that were parcellated on the basis of resting state functional connectivity data – Dorsal Attention (DA), Ventral Attention (VA), and Fronto-Parietal (FP). Working memory and attention were assessed using Repeatable Battery for Assessment of Neuropsychological Status (RBANS), and intelligence (IQ) was determined using Wechsler Abbreviated Scale of Intelligence (WASI). A series of simple and multiple mediation models were implemented using SPSS PROCESS macro. These models predicted the effects of the functional DRD2 haplotype on working memory, attention and intelligence, as independently and jointly mediated by cortical area, thickness and volume in the three functionally defined ROIs. Results The frequency of CCG homozygotes versus TAA carriers was significantly higher in at-risk subjects compared to controls (freq=0.758 vs 0.564, p=0.013) with a trend in BD (freq=0.692, p=0.1). In the control group, CCG homozygotes exhibited an increase in RBANS attention scores as a result of the mediating effects of DA area and volume (of 1.6 and 1.7 standard deviations). Simple mediation effects were also observed in the control group for IQ and RBANS immediate memory in the absence of a significant total effect. In contrast, simple mediation effects were not apparent in the slightly larger at-risk group nor the BD group. While multiple mediation models reinforced many simple mediation effects observed in controls, an additional mediation effect was observed in the BD group – TAT haplotype carriers exhibited a 0.6 SD decrease in RBANS attention scores as a result of the mediating effects of FP volume. Discussion The DRD2 mediation effects observed in controls were absent in the at-risk and BD groups, suggesting that the normally ameliorative effect of the CCG haplotype on cognitive outcomes is either negated by other genetic and familial risk factors for BD, or is mediated by alternate brain structures. Additionally, evidence is provided for the reversal of typically observed DRD2 effects on cognitive outcomes by the significant mediation effect for the BD group in the multiple mediation model of attention, where carriers of the TAT haplotype demonstrated improved attention.

Enhanced Sensitivity of Dopamine Biosensors: An Electrochemical Approach Based on Nanocomposite Electrodes Comprising Polyaniline, Nitrogen-Doped Graphene, and DNA-Functionalized Carbon Nanotubes

A new, highly-exfoliated nitrogen-doped graphene is electrochemically synthesized, which enhances the catalytic activity of poly(anilineboronic acid) nanocomposite electrodes for dopamine detection in the presence of excess ascorbic acid. The sensing approach is made up of poly(anilineboronic acid) nanocomposites electrodeposited on the surface of a glassy carbon electrode via in-situ electrochemical polymerization of anilineboronic acid monomers using cyclic voltammetry. A thin layer of DNA-functionalized carbon nanotubes, and nitrogen-doped graphene is coated on the electrode surface prior to electro-polymerization. During the electro-polymerization the π-π stacking and electrostatic interactions between DNA-coated carbon nanostructures and monomers anchors anilineboronic acid monomers on the electrode surface. This molecular anchoring increases electrodeposition of the respective nanocomposites on electrode; thus, greatly enhances the density of boronic acid receptors for dopamine binding. The coordinate covalent bonds between nitrogen atoms of graphene and boron atoms of anilineboronic acid monomers further increase the density of boronic acid groups for target analyte detection. The developed highly-sensitive and highly-selective biosensor is capable of dopamine detection in a wide linear range from 0.02-1μM, along with a detection limit of 14nM, which is a very significant step forward for dopamine detection and paves the way for molecular diagnosis of neurological illnesses such as Parkinson's disease.

Enzyme-Free Plasmonic Biosensor for Direct Detection of Neurotransmitter Dopamine from Whole Blood

Complex biological fluids without pretreatment, separation, or purification impose stringent limitations on the practical deployment of label-free plasmonic biosensors for advanced assays needed in point of care applications. In this work, we present an enzyme-free plasmonic neurotransmitter dopamine biosensor integrated with a microfluidic plasma separator. This integrated device allows the in-line separation of plasma directly from the bloodstream and channels it to the active detection area, where inorganic cerium oxide nanoparticles function as local selective dopamine binding sites through strong surface redox reaction. A thorough understanding and engineering of the nanoparticles is carried out to maximize its dopamine sensitivity and selectivity. We obtain detection of dopamine at 100 fM concentration in simulated body fluid and 1 nM directly from blood without any prior sample preparation. The detection selectivity is found to be at least five-times higher compared to the common interfering species. This demonstration shows the feasibility of the practical implementation of the proposed plasmonic system in detection of variety of biomarkers directly from the complex biological fluids.

Molecularly Imprinted Fluorescent Test Strip for Direct, Rapid, and Visual Dopamine Detection in Tiny Amount of Biofluid

Paper-based assays for detection of physiologically important species are needed in medical theranostics owning to their superiorities in point of care testing, daily monitoring, and even visual readout by using chromogenic materials. In this work, a facile test strip is developed for visual detection of a neurotransmitter dopamine (DA) based on dual-emission fluorescent molecularly imprinted polymer nanoparticles (DE-MIPs). The DE-MIPs, featured with tailor-made DA affinity and good anti-interference, exhibit DA concentration-dependent fluorescent colors, due to the variable ratios of dual-emission fluorescence caused by DA binding and quenching. By facile coating DE-MIPs on a filter paper, the DA test strips are obtained. The resultant test strip, like the simplicity of a pH test paper, shows the potential for directly visual detection of DA levels just by dripping a tiny amount of biofluid sample on it. The test result of real serum samples demonstrates that the DA strip enables to visually and semiquantitatively detect DA within 3 min by using only 10 µL of serum samples and with a low detection limit ((100-150) × 10-9 m) by naked eye. This work thus offers a facile and efficient strategy for rapid, visual, and on-site detection of biofluids in clinic.

Left-shifting prism adaptation boosts reward-based learning

Visuospatial cognition has an inherent lateralized bias. Individual differences in the direction and magnitude of this bias are associated with asymmetrical D2/3 dopamine binding and dopamine system genotypes. Dopamine level affects feedback-based learning and dopamine signaling asymmetry is related to differential learning from reward and punishment. High D2 binding in the left hemisphere is associated with preference for reward. Prism adaptation (PA) is a simple sensorimotor technique, which modulates visuospatial bias according to the direction of the deviation. Left-deviating prism adaptation (LPA) induces rightward bias in healthy subjects. It is therefore possible that the right side of space increases in saliency along with left hemisphere dopaminergic activity. Right-deviating prism adaptation (RPA) has been used mainly as a control condition because it does not modulate behavior in healthy individuals. Since LPA induces a rightward visuospatial bias as a result of left hemisphere modulation, and higher dopaminergic activity in the left hemisphere is associated with preference for rewarding events we hypothesized that LPA would increase the preference for learning with reward. Healthy volunteers performed a computer-based probabilistic classification task before and after LPA or RPA. Consistent with our predictions, PA altered the preference for rewarded versus punished learning, with the LPA group exhibiting increased learning from reward. These results suggest that PA modulates dopaminergic activity in a lateralized fashion.

In vitro and in silico analysis of the effects of D2 receptor antagonist target binding kinetics on the cellular response to fluctuating dopamine concentrations.

Background and PurposeTarget binding kinetics influence the time course of the drug effect (pharmacodynamics) both (i) directly, by affecting the time course of target occupancy, driven by the pharmacokinetics of the drug, competition with endogenous ligands and target turnover, and (ii) indirectly, by affecting signal transduction and homeostatic feedback. For dopamine D2 receptor antagonists, it has been hypothesized that fast receptor binding kinetics cause fewer side effects, because part of the dynamics of the dopaminergic system is preserved by displacement of these antagonists.Experimental ApproachTarget binding kinetics of D2 receptor antagonists and signal transduction after dopamine and D2 receptor antagonist exposure were measured in vitro. These data were integrated by mechanistic modelling, taking into account competitive binding of endogenous dopamine and the antagonist, the turnover of the second messenger cAMP and negative feedback by PDE turnover.Key ResultsThe proposed signal transduction model successfully described the cellular cAMP response for 17 D2 receptor antagonists with widely different binding kinetics. Simulation of the response to fluctuating dopamine concentrations revealed that a significant effect of the target binding kinetics on the dynamics of the signalling only occurs at endogenous dopamine concentration fluctuations with frequencies below 1 min−1.Conclusions and ImplicationsSignal transduction and feedback are important determinants of the time course of drug effects. The effect of the D2 receptor antagonist dissociation rate constant (koff) is limited to the maximal rate of fluctuations in dopamine signalling as determined by the dopamine koff and the cAMP turnover.

In Vivo Use of a Multi-DNA Aptamer-Based Payload/Targeting System To Study Dopamine Dysregulation in the Central Nervous System.

The delivery of therapeutics across the blood-brain barrier remains a considerable challenge in investigating central nervous system related processes. In this work, a liposome vehicle was surface-modified with an aptamer that binds to the transferrin receptor and was loaded with two different dopamine-binding aptamer payloads. This system was effectively used to promote the delivery of the aptamer cargo from the peripheral injection site into the brain. The effect of these delivered aptamers on behavior was investigated in vivo in a locomotor task. The first dopamine binding aptamer assessed was a DNA aptamer, the binding of which had been previously validated through the aptamer-based biosensor development reported by several independent research groups. The second aptamer investigated was the result of a novel in vitro selection experiment described herein. Our data suggest that systemic administration of the modified liposomes led to delivery of the dopamine aptamers into the brain. Fluorescence microscopy revealed differential distribution of fluorescence based on the presence or absence of the transferrin receptor aptamer on the surface of fluorescently modified liposomes. In a behavioral experiment using cocaine administration to induce elevated concentrations of neural dopamine, systemic pretreatment with the dopamine aptamer-loaded liposomes reduced cocaine-induced hyperlocomotion. Multiple controls including a transferrin-negative liposome control and transferrin-positive liposomes loaded with either a nonbinding, base-substituted dopamine aptamer or a random oligonucleotide were investigated. None of these controls altered cocaine-induced hyperlocomotion. Chronic systemic administration of the modified liposomes produced no deleterious neurobehavioral or neural degenerative effects. Importantly, this work is one example of an application for this versatile multiaptamer payload/targeting system. Its general application is limited only by the availability of aptamers for specific neural targets.

Photoinduced holes transfer based visual determination of dopamine in human serum.

A simple unexplored strategy was followed to construct ratiometric fluorescence-based sensing system for the detection of dopamine (DA) in human serum. Ratiometric fluorescence system was constructed through bonding energy transfer (TBET) by conjugating carboxyl functionalized CdTe quantum dots (QDs) and amine-capped Carbon quantum dots (CQDs). The photophysical properties of sensing system were characterized by standard methods. Photoluminescence (PL) of sensing system under excitonic wavelength (350 nm) depends on dual emission at 440 and 595 nm that corresponds to CQDs and CdTe QDs respectively. The developed sensing system was utilized for visual determination of DA, an unquenched blue fluorescence of CQDs in ratiometric system reveals the visual color differentiation for DA binding with CdTe QDs among the possible interferences (Alanine, Glycine, Glucose, Sucrose, Urea and Ascorbic acid). The limit of detection (LOD) and quantification (LOQ) was calculated as 8.1 and 27.2 nm respectively by using regression analysis. Photoinduced holes transfer (PHT) might have attributed the possible sensing mechanism for DA that quench the photoluminescence sequentially to enhance the sensing performance of QDs. The matrix interferences and reliability of the developed sensing platform were evaluated by testing DA spiked human serum and the sensing response was found to be field deployable.

Dual molecular recognition strategy for highly sensitive electrochemical detection of dopamine based on amplification of DNA–Au bio–bar codes

We had reported an electrochemical assay for dopamine with high sensitivity and selectivity based on dual molecular recognition strategy and amplification of DNA–Au bio–bar codes. In this assay, carbon nanotubes were first immobilized on the glass carbon electrode surface. Subsequently, a boronic acid-functionalized pyrene probe was self-assembled on the sidewalls of carbon nanotubes via π–π stacking interactions as a capture probe. 3,3′-Dithiodipropionic acid di(N-hydroxysuccinimide ester) (DSP) and double strands DNA functionalized AuNPs (DSP/AuNPs/dsDNA) were designed as signal probes and characterized by transmission electron microscopy and spectroscopic techniques. Upon the binding of dopamine with boronic acid followed by DSP/AuNPs/dsDNA, the AuNPs were captured on the electrode surface to enhance the signal. RuHex, an electroactive complex, which could bind to the anionic phosphate of DNA strands through electrostatic interaction, serves as signaling transducer. Signal amplification and dual recognition were used to quantify dopamine concentration from 30 pM to 30 nM, with a detection limit of 18 pM. Other small molecules could be measured by this assay after modification with differential recognition molecules.

Negatively Charged Gangliosides Promote Membrane Association of Amphipathic Neurotransmitters

Lipophilic neurotransmitters (NTs) such as dopamine are chemical messengers enabling neurotransmission by adhering onto the extracellular surface of the post-synaptic membrane in a synapse, followed by binding to their receptors. Previous studies have shown that the strength of the NT–membrane association is dependent on the lipid composition of the membrane. Negatively charged lipids such as phosphatidylserine, phosphatidylglycerol, and phosphatidic acid have been indicated to promote NT–membrane binding, however these anionic lipids reside almost exclusively in the intracellular leaflet of the post-synaptic membrane instead of the extracellular leaflet facing the synaptic cleft. Meanwhile, the extracellular leaflet is relatively rich in biologically relevant anionic gangliosides such as monosialotetrahexosylganglioside (GM1), yet the role of gangliosides in NT–membrane association is not clear. Here, we explored the role of GM1 in modulating the binding of dopamine and histamine (as amphipathic/cationic NTs) as well as acetylcholine (as a hydrophilic/cationic NT) with the post-synaptic membrane surface. Atomistic molecular dynamics simulations and free energy calculations indicated that GM1 fosters membrane association of histamine and dopamine. For acetylcholine, this effect was not observed. The in silico results suggest that gangliosides form a charge-based vestibule in front of the post-synaptic membrane, attracting amphipathic NTs to the vicinity of the membrane. The results also stress the importance to understand the significance of the structural details of NTs, as exemplified by the GM1–acetylcholine interaction. In a larger context, the NT–membrane adherence, coupled to lateral diffusion in the membrane plane, is proposed to improve neurotransmission efficiency by advancing NT entry into the membrane-embedded ligand-binding sites.

(Invited) Near Infrared Chemical Imaging of Cellular Communication Using Carbon Nanotubes

Intercellular chemical communication proceeds with both spatial and temporal components. A very prominent example is neurotransmitter release from neurons. But so far there are no analytical tools available to detect release and propagation of signaling molecules on small length scales and fast time scales. We have developed fluorescent nanosensors to monitor interactions of cells with their environment e.g. for chemical imaging of small molecules released by cells. For that purpose we used near infrared (nIR) fluorescent single-walled carbon nanotubes (SWCNTs) and chemically rendered them selective for small signaling molecules such as the neurotransmitter dopamine. When dopamine binds the quantum yield of the sensors increased by up to 400 % and we were able to report single dopamine binding events. Parallel imaging of many of those sensors provides a nIR image (900 nm -1300 nm) of the dopamine concentration around cells. Furthermore, we used a stochastic kinetic Monte-Carlo simulation to predict the collective image of many sensors and how on and off rate constants affect the spatial and temporal resolution of this approach. This technique was used to image hot spots and directionality of dopamine release from neurons and other cells. We observed localized, unlabeled release sites that correlate with membrane curvature and cell morphology. We furthermore tailored SWCNTs to bind cell surface receptors such as integrins to monitor and investigate cell adhesion and migration on surfaces. These results illustrate how to use fluorescent nanosensors for quantitative chemical imaging of complex biological processes.

Elucidation of self‐mediated enhancement of dopamine transport by the dopamine transporter which can be modulated by extracellular gate and N‐terminal residues

The dopamine (DA) and serotonin (5‐HT) transporters (DAT and SERT respectively) are members of the SLC6 family of neurotransmitter:sodium symporters (NSSs). DAT and SERT clear substrate from perisynaptic spaces by the generally accepted alternating access mechanism in which the transporters transition between outward‐facing and inward‐facing conformational states. This mechanism involves a sequential closing and opening of extracellular (EC) and intracellular (IC) gates along the permeation pathway. The EC and IC gates are almost exclusively composed of acidic and basic residues capable of interacting through salt bridge formation, closing the primary substrate binding site to the intracellular and extracellular milieus. Previously, we identified that the human SERT acidic/basic residue pair is critical for the recognition of 3,4‐methylenedioxymethamphetamine (MDMA) as a substrate. In contrast, mutations in the EC gate had no effect on 5‐HT recognition. Our current efforts focus on determining if the EC gate of the human DAT has similar mechanistic characteristics for substrate selectivity. While investigating that possibility, we identified a previously unreported phenomenon of self‐mediated enhancement of DA transport in hDAT expressing cells. In homologous competitive uptake experiments utilizing [3H]DA, a 200% to 300% increase in [3H]DA uptake is observed upon addition of micromolar concentrations of unlabeled DA. This enhancement is further modulated by the identity of the acidic EC gate residue. Substituting the Asp with the polar amino acid Asn (D476N) produces a 600% increase in [3H]DA uptake. Experiments utilizing heterologous competition with [3H]DA and the known hDAT substrates 1‐methyl‐4‐phenylpyridinium (MPP+), 4‐(4‐dimethylamino)phenyl‐1‐methylpyridinium(APP+) and tyramine indicate that the enhancement effect is not exclusive to DA nor is it observed with all DAT substrates. Notably, elimination of phosphorylation at N‐terminal residue Thr53 virtually eliminates the DA self‐enhancement phenotype. The apparent ability of DA to enhance its own uptake is consistent with reports proposing a second substrate binding site (S2) that can act as a trigger for release of substrate from the primary DA binding site (S1). However, the presence of this S2 site remains controversial. Continued investigation of this phenomenon may provide insight into molecular determinants on both the substrate and transporter that are important for substrate identification and translocation.This abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

Positive Allosteric Modulators of the D1 Dopamine Receptor Act at Diverse Binding Sites

The D1 dopamine receptor (D1R) and its signaling is associated with several important neurological processes and neuropsychiatric disorders. Enhancing D1R signaling in the prefrontal cortex is associated with increased cognition and therefore is an appealing treatment strategy for the cognitive decline observed in schizophrenia, Alzheimer's disease, and other disorders. Because of the clinical liability inherent with D1R agonists, positive allosteric modulators of the D1R have been proposed as an alternative, due to their potential for high selectivity and larger therapeutic windows. Currently, the location and characteristics of potential binding site(s) for allosteric modulators on the D1R is unknown. We have identified two structurally diverse D1R positive allosteric modulators, MLS1082 and MLS6585, via a high throughput screen of the NIH Molecular Libraries Program. The compounds potentiate dopamine‐stimulated G‐protein‐ (cAMP stimulation, 3–5 fold) and β‐arrestin‐mediated (6–8 fold) signaling pathways and increase the binding affinity of dopamine for the D1R (3–6 fold). Neither compound displayed agonist activity in the absence of dopamine. Taking advantage of our structurally distinct potentiators, G‐protein signaling and β‐arrestin recruitment experiments using maximally effective concentrations of MLS6585 and MLS1082 in combination were used to determine if the compounds act at similar or separate sites. In combination, MLS1082 + MLS6585 caused an additive potentiation of the potency of dopamine beyond that caused by either compound alone for both β‐arrestin recruitment (11–20 fold) and cAMP accumulation (4–6 fold). This suggests that the two compounds are acting at separate sites on the receptor. We also observed similar results using analogs of the two compounds, with analogs of MLS6585 having additive activity with MLS1082 and vice‐versa. However, analogs are not additive with their parent compound, meaning the parent compounds and their analogs act at similar sites. The combination experiments were repeated with Compound B, a known D1R positive allosteric modulator. Compound B was additive with MLS6585 but not MLS1082, further suggesting that the D1R has two separate positive allosteric modulator binding sites. Point mutation studies are currently underway to identify the location of these putative sites as well as whether these sites may contribute separately to potentiation of signaling efficacy versus dopamine binding affinity.Support or Funding InformationNINDS IRPThis abstract is from the Experimental Biology 2018 Meeting. There is no full text article associated with this abstract published in The FASEB Journal.

α2A- and α2C-Adrenoceptors as Potential Targets for Dopamine and Dopamine Receptor Ligands.

The poor norepinephrine innervation and high density of Gi/o-coupled α2A- and α2C-adrenoceptors in the striatum and the dense striatal dopamine innervation have prompted the possibility that dopamine could be an effective adrenoceptor ligand. Nevertheless, the reported adrenoceptor agonistic properties of dopamine are still inconclusive. In this study, we analyzed the binding of norepinephrine, dopamine, and several compounds reported as selective dopamine D2-like receptor ligands, such as the D3 receptor agonist 7-OH-PIPAT and the D4 receptor agonist RO-105824, to α2-adrenoceptors in cortical and striatal tissue, which express α2A-adrenoceptors and both α2A- and α2C-adrenoceptors, respectively. The affinity of dopamine for α2-adrenoceptors was found to be similar to that for D1-like and D2-like receptors. Moreover, the exogenous dopamine receptor ligands also showed high affinity for α2A- and α2C-adrenoceptors. Their ability to activate Gi/o proteins through α2A- and α2C-adrenoceptors was also analyzed in transfected cells with bioluminescent resonance energy transfer techniques. The relative ligand potencies and efficacies were dependent on the Gi/o protein subtype. Furthermore, dopamine binding to α2-adrenoceptors was functional, inducing changes in dynamic mass redistribution, adenylyl cyclase activity, and ERK1/2 phosphorylation. Binding events were further studied with computer modeling of ligand docking. Docking of dopamine at α2A- and α2C-adrenoceptors was nearly identical to its binding to the crystallized D3 receptor. Therefore, we provide conclusive evidence that α2A- and α2C-adrenoceptors are functional receptors for norepinephrine, dopamine, and other previously assumed selective D2-like receptor ligands, which calls for revisiting previous studies with those ligands.

Flipped Phenyl Ring Orientations of Dopamine Binding with Human and Drosophila Dopamine Transporters: Remarkable Role of Three Nonconserved Residues.

Molecular modeling and molecular dynamics simulations were performed in the present study to examine the modes of dopamine binding with human and Drosophila dopamine transporters (hDAT and dDAT). The computational data revealed flipped binding orientations of dopamine in hDAT and dDAT due to the major differences in three key residues (S149, G153, and A423 of hDAT vs A117, D121, and S422 of dDAT) in the binding pocket. These three residues dictate the binding orientation of dopamine in the binding pocket, as the aromatic ring of dopamine tends to take an orientation with both the para- and meta-hydroxyl groups being close to polar residues and away from nonpolar residues of the protein. The flipped binding orientations of dopamine in hDAT and dDAT clearly demonstrate a generally valuable insight concerning how the species difference could drastically affect the protein-ligand binding modes, demonstrating that the species difference, which is a factor rarely considered in early drug design stage, must be accounted for throughout the ligand/drug design and discovery processes in general.

Perceived fatigue and energy are independent unipolar states: Supporting evidence

Persistent fatigue is a common problem (∼20–45% of U.S. population), with higher prevalence and severity in people with medical conditions such as cancer, depression, fibromyalgia, heart failure, sleep apnea and multiple sclerosis. There are few FDA-approved treatments for fatigue and great disagreement on how to measure fatigue, with over 250 instruments used in research. Many instruments define fatigue as “a lack of energy”, thus viewing energy and fatigue states as opposites on a single bipolar continuum. In this paper, we hypothesize that energy and fatigue are distinct perceptual states, should be measured using separate unipolar scales, have different mechanisms, and deficits should be treated using tailored therapies. Energy and fatigue independence has been found in both exploratory and confirmatory factor analysis studies. Experiments in various fields, including behavioral pharmacology and exercise science, often find changes in energy and not fatigue, or vice versa. If the hypothesis that energy and fatigue are independent is correct, there are likely different mechanisms that drive energy and fatigue changes. Energy could be increased by elevated dopamine and norepinephrine transmission and binding. Fatigue could be increased by elevated brain serotonin and inflammatory cytokines and reduced histamine binding. The hypothesis could be tested by an experiment that attempts to produce simultaneously high ratings of energy and fatigue (such as with two drugs using a randomized, double-blind, placebo-controlled design), which would offer strong evidence against the common viewpoint of a bipolar continuum. If the hypothesis is correct, prior literature using bipolar instruments will be limited, and research on the prevalence, mechanisms, and treatment of low energy and elevated fatigue as separate conditions will be needed. In the immediate future, measuring both energy and fatigue using unipolar measurement tools may improve our understanding of these states and improve therapeutic outcomes.

Optical Control of Dopamine Receptors Using a Photoswitchable Tethered Inverse Agonist.

Family A G protein-coupled receptors (GPCRs) control diverse biological processes and are of great clinical relevance. Their archetype rhodopsin becomes naturally light sensitive by binding covalently to the photoswitchable tethered ligand (PTL) retinal. Other GPCRs, however, neither bind covalently to ligands nor are light sensitive. We sought to impart the logic of rhodopsin to light-insensitive Family A GPCRs in order to enable their remote control in a receptor-specific, cell-type-specific, and spatiotemporally precise manner. Dopamine receptors (DARs) are of particular interest for their roles in motor coordination, appetitive, and aversive behavior, as well as neuropsychiatric disorders such as Parkinson's disease, schizophrenia, mood disorders, and addiction. Using an azobenzene derivative of the well-known DAR ligand 2-(N-phenethyl-N-propyl)amino-5-hydroxytetralin (PPHT), we were able to rapidly, reversibly, and selectively block dopamine D1 and D2 receptors (D1R and D2R) when the PTL was conjugated to an engineered cysteine near the dopamine binding site. Depending on the site of tethering, the ligand behaved as either a photoswitchable tethered neutral antagonist or inverse agonist. Our results indicate that DARs can be chemically engineered for selective remote control by light and provide a template for precision control of Family A GPCRs.

A Fluorescence Lifetime Imaging Microscopy Supported Investigation on Temperature-Dependent Penetration of Dopamine in a 1,2-Ditetradecanoyl-sn-glycero-3-phospho-(1′-rac-glycerol) Lipid Bilayer

Distribution of dopamine, an essential neurotransmitter in mammalian central and peripheral nervous systems, in a lipid bilayer and at the surface of 1,2-ditetradecanoyl-sn-glycero-3-phospho-(1'-rac-glycerol) vesicles has been studied herein. To track the progress of dopamine through different regions of the lipid vesicle, these were synthesized using 7-nitrobenz-2-oxa-1,3-diazol-4-yl (NBD)-labeled phospholipid molecules tagged to either the headgroup (NBDPE) or the acyl chain (NBDPG). Dopamine-induced quenching of NBD fluorescence in the lipid vesicles demonstrates that dopamine has a preference to diffuse into the lipid bilayer. The change in the excited state lifetime obtained for NBDPG clearly indicates the preference in dopamine binding. The propositions were supported by fluorescence lifetime imaging microscopy.

A DNA sequence obtained by replacement of the dopamine RNA aptamer bases is not an aptamer

A unique specificity of the aptamer-ligand biorecognition and binding facilitates bioanalysis and biosensor development, contributing to discrimination of structurally related molecules, such as dopamine and other catecholamine neurotransmitters. The aptamer sequence capable of specific binding of dopamine is a 57 nucleotides long RNA sequence reported in 1997 (Biochemistry, 1997, 36, 9726). Later, it was suggested that the DNA homologue of the RNA aptamer retains the specificity of dopamine binding (Biochem. Biophys. Res. Commun., 2009, 388, 732). Here, we show that the DNA sequence obtained by the replacement of the RNA aptamer bases for their DNA analogues is not able of specific biorecognition of dopamine, in contrast to the original RNA aptamer sequence. This DNA sequence binds dopamine and structurally related catecholamine neurotransmitters non-specifically, as any DNA sequence, and, thus, is not an aptamer and cannot be used neither for in vivo nor in situ analysis of dopamine in the presence of structurally related neurotransmitters.