Persistent binding at dopamine transporters determines sustained psychostimulant effects.

Psychostimulants interacting with the dopamine transporter (DAT) can be used illicitly or for the treatment of specific neuropsychiatric disorders. However, they can also produce severe and persistent adverse events. Often, their pharmacological properties in vitro do not fully correlate to their pharmacological profile in vivo. Here, we investigated the pharmacological effects of enantiomers of pyrovalerone, α-pyrrolidinovalerophenone, and 3,4-methylenedioxypyrovalerone as compared to the traditional psychostimulants cocaine and methylphenidate, using a variety of in vitro, computational, and in vivo approaches. We found that in vitro drug-binding kinetics at DAT correlate with the time-course of in vivo psychostimulant action in mice. In particular, a slow dissociation (i.e., slow koff) of S-enantiomers of pyrovalerone analogs from DAT predicts their more persistent in vivo effects when compared to cocaine and methylphenidate. Overall, our findings highlight the critical importance of drug-binding kinetics at DAT for determining the in vivo profile of effects produced by psychostimulant drugs.

Reversing anterior insular cortex neuronal hypoexcitability attenuates compulsive behavior in adolescent rats.

Development of self-regulatory competencies during adolescence is partially dependent on normative brain maturation. Here, we report that adolescent rats as compared to adults exhibit impulsive and compulsive-like behavioral traits, the latter being associated with lower expression of mRNA levels of the immediate early gene zif268 in the anterior insula cortex (AIC). This suggests that underdeveloped AIC function in adolescent rats could contribute to an immature pattern of interoceptive cue integration in decision making and a compulsive phenotype. In support of this, we report that layer 5 pyramidal neurons in the adolescent rat AIC are hypoexcitable and receive fewer glutamatergic synaptic inputs compared to adults. Chemogenetic activation of the AIC attenuated compulsive traits in adolescent rats supporting the idea that in early stages of AIC maturity there exists a suboptimal integration of sensory and cognitive information that contributes to inflexible behaviors in specific conditions of reward availability.

Synthetic Placement of Active Sites in MFI Zeolites for Selective Toluene Methylation to para-Xylene.

Brønsted acidic zeolites are ubiquitous catalysts in fuel and chemical production. Broadening the catalytic diversity of a given zeolite requires strategies to manipulate the acid site placement at framework positions within distinct microporous locations. Here, we combine experiment and theory to elucidate how intermolecular interactions between organic structure-directing agents (OSDAs) and framework Al centers influence the placement of H+ sites in distinct void environments of MFI zeolites and demonstrate the catalytic consequences of active site location on kinetically controlled (403 K) toluene methylation to xylene regioisomers. Kinetic measurements, interpreted using mechanism-derived rate expressions and transition state theory, alongside density functional theory (DFT) calculations show that larger intersection environments similarly stabilize all three xylene isomer transition states without altering well-established aromatic substitution patterns (ortho/para/meta ∼ 60%:30%:10%), while smaller channel environments preferentially destabilize transition states that form bulkier ortho- and meta-isomers, thereby resulting in high intrinsic para-xylene selectivity (∼80%). DFT calculations reveal that the flexibility of nonconventional OSDAs (e.g., 1,4-diazabicyclo[2.2.2]octane) to reorient within MFI intersections and their ability to hydrogen-bond to form protonated complexes favor the placement of Al in smaller channel environments compared to conventional quaternary OSDAs (e.g., tetra-n-propylammonium). These molecular-level insights establish a mechanistic link between OSDA structure, active site placement, and transition state stability in MFI zeolites and provide active site design strategies that are orthogonal to crystallite design approaches harnessing complex reaction-diffusion phenomena to enhance regioisomer selectivity in the industrial production of valuable polymer precursors.

Muscarinic Acetylcholine M2 Receptors Regulate Lateral Habenula Neuron Activity and Control Cocaine Seeking Behavior.

The lateral habenula (LHb) balances reward and aversion by opposing activation of brain reward nuclei and is involved the inhibition of responding for cocaine in a model of impulsive behavior. Previously, we reported that the suppression of cocaine seeking was prevented by LHb inactivation or nonselective antagonism of LHb mAChRs. Here, we investigate mAChR subtypes mediating the effects of endogenous acetylcholine in this model of impulsive drug seeking and define cellular mechanisms in which mAChRs alter LHb neuron activity. Using in vitro electrophysiology, we find that LHb neurons are depolarized or hyperpolarized by the cholinergic agonists oxotremorine-M (Oxo-M) and carbachol (CCh), and that mAChRs inhibit synaptic GABA and glutamatergic inputs to these cells similarly in male and female rats. Synaptic effects of CCh were blocked by the M2-mAChR (M2R) antagonist AFDX-116 and not by pirenzepine, an M1-mAChR (M1R) antagonist. Oxo-M-mediated depolarizing currents were also blocked by AFDX-116. Although M2R activation inhibited excitatory and inhibitory inputs to LHb neurons, the effect on excitation was greater, suggesting a shift in excitatory-inhibitory balance toward net inhibition. Activation of VTA inhibitory inputs to LHb neurons, via channelrhodopsin-2 expression, evoked IPSCs that were inhibited by M2Rs. Finally, we measured LHb-dependent operant response inhibition for cocaine and found it impaired by antagonism of M2Rs, and not M1Rs. In summary, we show that a cholinergic signal to LHb and activation of M2Rs are critical to enable inhibition of responding for cocaine, and we define cellular mechanisms through which this may occur.Significance Statement:The lateral habenula (LHb) is a brain region receiving information from brain areas involved in decision-making, and its output influences motivation, reward, and movement. This interface between thoughts, emotions, and actions is how the LHb permits adaptive behavior, and LHb dysfunction is implicated in psychiatric and drug use disorders. Silencing the LHb impairs control over cocaine seeking in rats, and mAChRs are also implicated. Here, we measured cocaine seeking while blocking different mAChRs and examined mechanisms of mAChR effects on LHb neurons. M2-mAChRs were necessary for control of cocaine seeking, and these receptors altered LHb neuron activity in several ways. Our study reveals that LHb M2-mAChRs represent a potential target for treating substance use disorders.

A new paradigm for regulation of protein phosphatase 2A function via Src and Fyn kinase-mediated tyrosine phosphorylation.

Protein phosphatase 2A (PP2A) is a major phospho-Ser/Thr phosphatase and a key regulator of cellular signal transduction pathways. While PP2A dysfunction has been linked to human cancer and neurodegenerative disorders such as Alzheimer's disease (AD), PP2A regulation remains relatively poorly understood. It has been reported that the PP2A catalytic subunit (PP2Ac) is inactivated by a single phosphorylation at the Tyr307 residue by tyrosine kinases such as v-Src. However, multiple mass spectrometry studies have revealed the existence of other putative PP2Ac phosphorylation sites in response to activation of Src and Fyn, two major Src family kinases (SFKs). Here, using PP2Ac phosphomutants and novel phosphosite-specific PP2Ac antibodies, we show that cellular pools of PP2Ac are instead phosphorylated on both Tyr127 and Tyr284 upon Src activation, and on Tyr284 following Fyn activation. We found these phosphorylation events enhanced the interaction of PP2Ac with SFKs. In addition, we reveal SFK-mediated phosphorylation of PP2Ac at Y284 promotes dissociation of the regulatory Bα subunit, altering PP2A substrate specificity; the phosphodeficient Y127/284F and Y284F PP2Ac mutants prevented SFK-mediated phosphorylation of Tau at the CP13 (pSer202) epitope, a pathological hallmark of AD, and SFK-dependent activation of ERK, a major growth regulatory kinase upregulated in many cancers. Our findings demonstrate a novel PP2A regulatory mechanism that challenges the existing dogma on the inhibition of PP2A catalytic activity by Tyr307 phosphorylation. We propose dysregulation of SFK signaling in cancer and AD can lead to alterations in PP2A phosphorylation and subsequent deregulation of key PP2A substrates, including ERK and Tau.

How Water and Ion Mobility Affect the NMR Fingerprints of the Hydrated JBW Zeolite: A Combined Computational-Experimental Investigation.

An important aspect within zeolite synthesis is to make fully tunable framework materials with controlled aluminium distribution. A major challenge in characterising these zeolites at operating conditions is the presence of water. In this work, we investigate the effect of hydration on the 27 Al NMR parameters of the ultracrystalline K,Na-compensated aluminosilicate JBW zeolite using experimental and computational techniques. The JBW framework, with Si/Al ratio of 1, is an ideal benchmark system as a stepping stone towards more complicated zeolites. The presence and mobility of water and extraframework species directly affect NMR fingerprints. Excellent agreement between theoretical and experimental spectra is obtained provided dynamic methods are employed with hydrated structural models. This work shows how NMR is instrumental in characterising aluminium distributions in zeolites at operating conditions.

Striatal Rgs4 regulates feeding and susceptibility to diet-induced obesity.

Consumption of high fat, high sugar (western) diets is a major contributor to the current high levels of obesity. Here, we used a multidisciplinary approach to gain insight into the molecular mechanisms underlying susceptibility to diet-induced obesity (DIO). Using positron emission tomography (PET), we identified the dorsal striatum as the brain area most altered in DIO-susceptible rats and molecular studies within this region highlighted regulator of G-protein signaling 4 (Rgs4) within laser-capture micro-dissected striatonigral (SN) and striatopallidal (SP) medium spiny neurons (MSNs) as playing a key role. Rgs4 is a GTPase accelerating enzyme implicated in plasticity mechanisms of SP MSNs, which are known to regulate feeding and disturbances of which are associated with obesity. Compared to DIO-resistant rats, DIO-susceptible rats exhibited increased striatal Rgs4 with mRNA expression levels enriched in SP MSNs. siRNA-mediated knockdown of striatal Rgs4 in DIO-susceptible rats decreased food intake to levels comparable to DIO-resistant animals. Finally, we demonstrated that the human Rgs4 gene locus is associated with increased body weight and obesity susceptibility phenotypes, and that overweight humans exhibit increased striatal Rgs4 protein. Our findings highlight a novel role for involvement of Rgs4 in SP MSNs in feeding and DIO-susceptibility.

Atomistic insight in the flexibility and heat transport properties of the stimuli-responsive metal-organic framework MIL-53(Al) for water-adsorption applications using molecular simulations.

To exploit the full potential of metal-organic frameworks as solid adsorbents in water-adsorption applications, many challenges remain to be solved. A more fundamental insight into the properties of the host material and the influence that water exerts on them can be obtained by performing molecular simulations. In this work, the prototypical flexible MIL-53(Al) framework is modelled using advanced molecular dynamics simulations. For different water loadings, the presence of water is shown to affect the relative stability of MIL-53(Al), triggering a phase transition from the narrow-pore to the large-pore phase at the highest considered loading. Furthermore, the effect of confinement on the structural organisation of the water molecules is also examined for different pore volumes of MIL-53(Al). For the framework itself, we focus on the thermal conductivity, as this property plays a decisive role in the efficiency of adsorption-based technologies, due to the energy-intensive adsorption and desorption cycles. To this end, the heat transfer characteristics of both phases of MIL-53(Al) are studied, demonstrating a strong directional dependence for the thermal conductivity.

Identification of vanadium dopant sites in the metal-organic framework DUT-5(Al).

Studying the structural environment of the VIV ions doped in the metal-organic framework (MOF) DUT-5(Al) ((AlIIIOH)BPDC) with electron paramagnetic resonance (EPR) reveals four different vanadium-related spectral components. The spin-Hamiltonian parameters are derived by analysis of X-, Q- and W-band powder EPR spectra. Complementary Q-band Electron Nuclear DOuble Resonance (ENDOR) experiments, Scanning Electron Microscopy (SEM), Energy Dispersive X-ray spectroscopy (EDX), X-Ray Diffraction (XRD) and Fourier Transform InfraRed (FTIR) measurements are performed to investigate the origin of these spectral components. Two spectral components with well resolved 51V hyperfine structure are visible, one corresponding to VIV[double bond, length as m-dash]O substitution in a large (or open) pore and one to a narrow (or closed) pore variant of this MOF. Furthermore, a broad structureless Lorentzian line assigned to interacting vanadyl centers in each other's close neighborhood grows with increasing V-concentration. The last spectral component is best visible at low V-concentrations. We tentatively attribute it to (VIV[double bond, length as m-dash]O)2+ linked with DMF or dimethylamine in the pores of the MOF. Simulations using these four spectral components convincingly reproduce the experimental spectra and allow to estimate the contribution of each vanadyl species as a function of V-concentration.

Engineering a Highly Defective Stable UiO-66 with Tunable Lewis- Brønsted Acidity: The Role of the Hemilabile Linker.

The stability of metal-organic frameworks (MOFs) typically decreases with an increasing number of defects, limiting the number of defects that can be created and limiting catalytic and other applications. Herein, we use a hemilabile (Hl) linker to create up to a maximum of six defects per cluster in UiO-66. We synthesized hemilabile UiO-66 (Hl-UiO-66) using benzene dicarboxylate (BDC) as linker and 4-sulfonatobenzoate (PSBA) as the hemilabile linker. The PSBA acts not only as a modulator to create defects but also as a coligand that enhances the stability of the resulting defective framework. Furthermore, upon a postsynthetic treatment in H2SO4, the average number of defects increases to the optimum of six missing BDC linkers per cluster (three per formula unit), leaving the Zr-nodes on average sixfold coordinated. Remarkably, the thermal stability of the materials further increases upon this treatment. Periodic density functional theory calculations confirm that the hemilabile ligands strengthen this highly defective structure by several stabilizing interactions. Finally, the catalytic activity of the obtained materials is evaluated in the acid-catalyzed isomerization of α-pinene oxide. This reaction is particularly sensitive to the Brønsted or Lewis acid sites in the catalyst. In comparison to the pristine UiO-66, which mainly possesses Brønsted acid sites, the Hl-UiO-66 and the postsynthetically treated Hl-UiO-66 structures exhibited a higher Lewis acidity and an enhanced activity and selectivity. This is further explored by CD3CN spectroscopic sorption experiments. We have shown that by tuning the number of defects in UiO-66 using PSBA as the hemilabile linker, one can achieve highly defective and stable MOFs and easily control the Brønsted to Lewis acid ratio in the materials and thus their catalytic activity and selectivity.

Rigid Arrangements of Ionic Charge in Zeolite Frameworks Conferred by Specific Aluminum Distributions Preferentially Stabilize Alkanol Dehydration Transition States.

Zeolite reactivity depends on the solvating environments of their micropores and the proximity of their Brønsted acid sites. Turnover rates (per H+ ) for methanol and ethanol dehydration increase with the fraction of H+ sites sharing six-membered rings of chabazite (CHA) zeolites. Density functional theory (DFT) shows that activation barriers vary widely with the number and arrangement of Al (1-5 per 36 T-site unit cell), but cannot be described solely by Al-Al distance or density. Certain Al distributions yield rigid arrangements of anionic charge that stabilize cationic intermediates and transition states via H-bonding to decrease barriers. This is a key feature of acid catalysis in zeolite solvents, which lack the isotropy of liquid solvents. The sensitivity of polar transition states to specific arrangements of charge in their solvating environments and the ability to position such charges in zeolite lattices with increasing precision herald rich catalytic diversity among zeolites of varying Al arrangement.

A Supramolecular View on the Cooperative Role of Brønsted and Lewis Acid Sites in Zeolites for Methanol Conversion.

A systematic molecular level and spectroscopic investigation is presented to show the cooperative role of Brønsted acid and Lewis acid sites in zeolites for the conversion of methanol. Extra-framework alkaline-earth metal containing species and aluminum species decrease the number of Brønsted acid sites, as protonated metal clusters are formed. A combined experimental and theoretical effort shows that postsynthetically modified ZSM-5 zeolites, by incorporation of extra-framework alkaline-earth metals or by demetalation with dealuminating agents, contain both mononuclear [MOH]+ and double protonated binuclear metal clusters [M(µ-OH)2M]2+ (M = Mg, Ca, Sr, Ba, and HOAl). The metal in the extra-framework clusters has a Lewis acid character, which is confirmed experimentally and theoretically by IR spectra of adsorbed pyridine. The strength of the Lewis acid sites (Mg > Ca > Sr > Ba) was characterized by a blue shift of characteristic IR peaks, thus offering a tool to sample Lewis acidity experimentally. The incorporation of extra-framework Lewis acid sites has a substantial influence on the reactivity of propene and benzene methylations. Alkaline-earth Lewis acid sites yield increased benzene methylation barriers and destabilization of typical aromatic intermediates, whereas propene methylation routes are less affected. The effect on the catalytic function is especially induced by the double protonated binuclear species. Overall, the extra-framework metal clusters have a dual effect on the catalytic function. By reducing the number of Brønsted acid sites and suppressing typical catalytic reactions in which aromatics are involved, an optimal propene selectivity and increased lifetime for methanol conversion over zeolites is obtained. The combined experimental and theoretical approach gives a unique insight into the nature of the supramolecular zeolite catalyst for methanol conversion which can be meticulously tuned by subtle interplay of Brønsted and Lewis acid sites.

(-)-Phenserine and the prevention of pre-programmed cell death and neuroinflammation in mild traumatic brain injury and Alzheimer's disease challenged mice.

Mild traumatic brain injury (mTBI) is a risk factor for neurodegenerative disorders, such as Alzheimer's disease (AD) and Parkinson's disease (PD). TBI-derived neuropathologies are promoted by inflammatory processes: chronic microgliosis and release of pro-inflammatory cytokines that further promote neuronal dysfunction and loss. Herein, we evaluated the effect on pre-programmed cell death/neuroinflammation/synaptic integrity and function of (-)-Phenserine tartrate (Phen), an agent originally developed for AD. This was studied at two clinically translatable doses (2.5 and 5.0 mg/kg, BID), in a weight drop (concussive) mTBI model in wild type (WT) and AD APP/PSEN1 transgenic mice. Phen mitigated mTBI-induced cognitive impairment, assessed by Novel Object Recognition and Y-maze behavioral paradigms, in WT mice. Phen fully abated mTBI-induced neurodegeneration, evaluated by counting Fluoro-Jade C-positive (FJC+) cells, in hippocampus and cortex of WT mice. In APP/PSEN1 mice, degenerating cell counts were consistently greater across all experimental groups vs. WT mice. mTBI elevated FJC+ cell counts vs. the APP/PSEN1 control (sham) group, and Phen similarly mitigated this. Anti-inflammatory effects on microglial activation (IBA1-immunoreactivity (IR)) and the pro-inflammatory cytokine TNF-α were evaluated. mTBI increased IBA1-IR and TNF-α/IBA1 colocalization vs. sham, both in WT and APP/PSEN1 mice. Phen decreased IBA1-IR throughout hippocampi and cortices of WT mice, and in cortices of AD mice. Phen, likewise, reduced levels of IBA1/TNF-α-IR colocalization volume across all areas in WT animals, with a similar trend in APP/PSEN1 mice. Actions on astrocyte activation by mTBI were followed by evaluating GFAP, and were similarly mitigated by Phen. Synaptic density was evaluated by quantifying PSD-95+ dendritic spines and Synaptophysin (Syn)-IR. Both were significantly reduced in mTBI vs. sham in both WT and APP/PSEN1 mice. Phen fully reversed the PSD-95+ spine loss in WT and Syn-IR decrease in both WT and APP/PSEN1 mice. To associate immunohistochemical changes in synaptic markers with function, hippocampal long term potentiation (LTP) was induced in WT mice. LTP was impaired by mTBI, and this impairment was mitigated by Phen. In synopsis, clinically translatable doses of Phen ameliorated mTBI-mediated pre-programmed cell death/neuroinflammation/synaptic dysfunction in WT mice, consistent with fully mitigating mTBI-induced cognitive impairments. Phen additionally demonstrated positive actions in the more pathologic brain microenvironment of AD mice, further supporting consideration of its repurposing as a treatment for mTBI.

Cannabinoid disruption of learning mechanisms involved in reward processing.

The increasing use of cannabis, its derivatives, and synthetic cannabinoids for medicinal and recreational purposes has led to burgeoning interest in understanding the addictive potential of this class of molecules. It is estimated that ∼10% of marijuana users will eventually show signs of dependence on the drug, and the diagnosis of cannabis use disorder (CUD) is increasing in the United States. The molecule that sustains the use of cannabis is Δ9-tetrahydrocannabinol (Δ9-THC), and our knowledge of its effects, and those of other cannabinoids on brain function has expanded rapidly in the past two decades. Additionally, the identification of endogenous cannabinoid (endocannabinoid) systems in brain and their roles in physiology and behavior, demonstrate extensive involvement of these lipid signaling molecules in regulating CNS function. Here, we examine roles for endogenous cannabinoids in shaping synaptic activity in cortical and subcortical brain circuits, and we discuss mechanisms in which exogenous cannabinoids, such as Δ9-THC, interact with endocannabinoid systems to disrupt neuronal network oscillations. We then explore how perturbation of the interaction of this activity within brain reward circuits may lead to impaired learning. Finally, we propose that disruption of cellular plasticity mechanisms by exogenous cannabinoids in cortical and subcortical circuits may explain the difficulty in establishing viable cannabinoid self-administration models in animals.

Enduring Loss of Serotonergic Control of Orbitofrontal Cortex Function Following Contingent and Noncontingent Cocaine Exposure.

Clinical descriptions of cocaine addiction include compulsive drug seeking and maladaptive decision-making despite substantial aversive consequences. Research suggests that this may result from altered orbitofrontal cortex (OFC) function and its participation in outcome-based behavior. Clinical and animal studies also implicate serotonin in the regulation of OFC function in addiction and other neuropsychiatric disorders. Here we test the hypothesis that exposure to cocaine, through self-administration (CSA) or yoked-administration (CYA), alters the regulation of OFC function by 5-HT. Using whole-cell electrophysiology in brain slices from naïve rats we find that 5-HT1A receptors generate hyperpolarizing outward currents in layer-V OFC pyramidal neurons, and that 5-HT2A receptors increase glutamate release onto these cells. Following extended withdrawal from CSA or CYA, this 5-HT regulation of OFC activity is largely lost. In-situ hybridization of 5-HT receptor transcripts reveals that 5-HT1A receptor mRNA is unaffected and 5-HT2A receptor mRNA is significantly elevated after CSA or CYA. These results demonstrate that 5-HT control of OFC neurons is disrupted for extended periods following cocaine exposure. We hypothesize that this dysregulation of 5-HT signaling leads to enduring disruptions of OFC network activity that this is involved in impaired decision-making associated with cocaine addiction.

The protein serine/threonine phosphatases PP2A, PP1 and calcineurin: A triple threat in the regulation of the neuronal cytoskeleton.

The microtubule, F-actin and neurofilament networks play a critical role in neuronal cell morphogenesis, polarity and synaptic plasticity. Significantly, the assembly/disassembly and stability of these cytoskeletal networks is crucially modulated by protein phosphorylation and dephosphorylation events. Herein, we aim to more closely examine the role played by three major neuronal Ser/Thr protein phosphatases, PP2A, PP1 and calcineurin, in the homeostasis of the neuronal cytoskeleton. There is strong evidence that these enzymes interact with and dephosphorylate a variety of cytoskeletal proteins, resulting in major regulation of neuronal cytoskeletal dynamics. Conversely, we also discuss how multi-protein cytoskeletal scaffolds can also influence the regulation of these phosphatases, with important implications for neuronal signalling and homeostasis. Not surprisingly, deregulation of these cytoskeletal scaffolds and phosphatase dysfunction are associated with many neurological diseases.

Disruption of hippocampal synaptic transmission and long-term potentiation by psychoactive synthetic cannabinoid 'Spice' compounds: comparison with Δ9 -tetrahydrocannabinol.

There has been a marked increase in the availability of synthetic drugs designed to mimic the effects of marijuana. These cannabimimetic drugs, sold illicitly as 'Spice' and related products, are associated with serious medical complications in some users. In vitro studies suggest that synthetic cannabinoids in these preparations are potent agonists at central cannabinoid CB1 receptors (CB1Rs), but few investigations have delineated their cellular effects, particularly in comparison with the psychoactive component of marijuana, Δ9 -tetrahydrocannabinol (Δ9 -THC). We compared the ability of three widely abused synthetic cannabinoids and Δ9 -THC to alter glutamate release and long-term potentiation in the mouse hippocampus. JWH-018 was the most potent inhibitor of hippocampal synaptic transmission (EC50 ~15 nM), whereas its fluoropentyl derivative, AM2201, inhibited synaptic transmission with slightly lower potency (EC50 ~60 nM). The newer synthetic cannabinoid, XLR-11, displayed much lower potency (EC50 ~900 nM) that was similar to Δ9 -THC (EC50 ~700 nM). The effects of all compounds occurred via activation of CB1Rs, as demonstrated by reversal with the selective antagonist/inverse agonist AM251 or the neutral CB1R antagonist PIMSR1. Moreover, AM2201 was without effect in the hippocampus of transgenic mice lacking the CB1R. Hippocampal slices exposed to either synthetic cannabinoids or Δ9 -THC exhibited significantly impaired long-term potentiation (LTP). We find that, compared with Δ9 -THC, the first-generation cannabinoids found in Spice preparations display higher potency, whereas a recent synthetic cannabinoid is roughly equipotent with Δ9 -THC. The disruption of synaptic function by these synthetic cannabinoids is likely to lead to profound impairments in cognitive and behavioral function.

No Effect of Lifelong Methylmercury Exposure on Oxidative Status in Zebra Finches (Taeniopygia guttata): A Demonstration of Methylmercury-Induced Selection?

Songbirds exposed to methylmercury (MeHg) often exhibit reduced reproductive success and cognitive abilities. To better understand whether oxidative stress plays a role, we dosed zebra finches (Taeniopygia guttata) with a contaminated (1.2 ppm MeHg-cysteine) or control diet for their entire lives, including during development in the egg. Levels of antioxidant enzymes [superoxide dismutase (SOD1 and SOD2)], oxidative damage (4-hydroxynonenal; 4-HNE), and antioxidant transcription factors [nuclear factor (erythroid-derived 2)-like 2; Nrf2] were measured in the liver and pectoralis muscle of adults. MeHg treatment did not affect levels of 4-HNE or liver SOD2 or Nrf2. Birds in the MeHg treatment differed significantly from controls in pectoralis SOD1 and Nrf2, and tended to differ in liver SOD1 and pectoralis SOD2; however, we detected no overall pattern of effect of MeHg on oxidative status in dosed finches. We suspect that this is a consequence of the differential survival of MeHg-tolerant birds.

Norepinephrine activates dopamine D4 receptors in the rat lateral habenula.

The lateral habenula (LHb) is involved in reward and aversion and is reciprocally connected with dopamine (DA)-containing brain regions, including the ventral tegmental area (VTA). We used a multidisciplinary approach to examine the properties of DA afferents to the LHb in the rat. We find that >90% of VTA tyrosine hydroxylase (TH) neurons projecting to the LHb lack vesicular monoamine transporter 2 (VMAT2) mRNA, and there is little coexpression of TH and VMAT2 protein in this mesohabenular pathway. Consistent with this, electrical stimulation of LHb did not evoke DA-like signals, assessed with fast-scan cyclic voltammetry. However, electrophysiological currents that were inhibited by L741,742, a DA-D4-receptor antagonist, were observed in LHb neurons when DA uptake or degradation was blocked. To prevent DA activation of D4 receptors, we repeated this experiment in LHb slices from DA-depleted rats. However, this did not disrupt D4 receptor activation initiated by the dopamine transporter inhibitor, GBR12935. As the LHb is also targeted by noradrenergic afferents, we examined whether GBR12935 activation of DA-D4 receptors occurred in slices depleted of norepinephrine (NE). Unlike DA, NE depletion prevented the activation of DA-D4 receptors. Moreover, direct application of NE elicited currents in LHb neurons that were blocked by L741,742, and GBR12935 was found to be a more effective blocker of NE uptake than the NE-selective transport inhibitor nisoxetine. These findings demonstrate that NE is released in the rat LHb under basal conditions and that it activates DA-D4 receptors. Therefore, NE may be an important regulator of LHb function.

Dopamine D4 receptor excitation of lateral habenula neurons via multiple cellular mechanisms.

Glutamatergic lateral habenula (LHb) output communicates negative motivational valence to ventral tegmental area (VTA) dopamine (DA) neurons via activation of the rostromedial tegmental nucleus (RMTg). However, the LHb also receives a poorly understood DA input from the VTA, which we hypothesized constitutes an important feedback loop regulating DA responses to stimuli. Using whole-cell electrophysiology in rat brain slices, we find that DA initiates a depolarizing inward current (I(DAi)) and increases spontaneous firing in 32% of LHb neurons. I(DAi) was also observed upon application of amphetamine or the DA uptake blockers cocaine or GBR12935, indicating involvement of endogenous DA. I(DAi) was blocked by D4 receptor (D4R) antagonists (L745,870 or L741,742), and mimicked by a selective D4R agonist (A412997). I(DAi) was associated with increased whole-cell conductance and was blocked by Cs+ or a selective blocker of hyperpolarization-activated cyclic nucleotide-gated (HCN) ion channel, ZD7288. I(DAi) was also associated with a depolarizing shift in half-activation voltage for the hyperpolarization-activated cation current (Ih) mediated by HCN channels. Recordings from LHb neurons containing fluorescent retrograde tracers revealed that I(DAi) was observed only in cells projecting to the RMTg and not the VTA. In parallel with direct depolarization, DA also strongly increased synaptic glutamate release and reduced synaptic GABA release onto LHb cells. These results demonstrate that DA can excite glutamatergic LHb output to RMTg via multiple cellular mechanisms. Since the RMTg strongly inhibits midbrain DA neurons, activation of LHb output to RMTg by DA represents a negative feedback loop that may dampen DA neuron output following activation.