INTRODUCTION: Serotonin dysfunction has been linked to a variety of psychiatric diseases; however, an adequate SPECT radioligand to probe the serotonin transporter system has not been successfully developed. The purpose of this study was to characterize and determine the in vivo selectivity of iodine-123-labeled 2beta-carbomethoxy-3beta-(4'-((Z)-2-iodoethenyl)phenyl)nortropane, [(123)I]p ZIENT, in nonhuman primate brain. METHODS: Two ovariohysterectomized female baboons participated in nine studies (one bolus and eight bolus to constant infusion at a ratio of 9.0 h) to evaluate [(123)I]p ZIENT. To evaluate the selectivity of [(123)I]p ZIENT, the serotonin transporter blockers fenfluramine (1.5, 2.5 mg/kg) and citalopram (5 mg/kg), the dopamine transporter blocker methylphenidate (0.5 mg/kg) and the norepinephrine transporter blocker nisoxetine (1 mg/kg) were given at 8 h post-radiotracer injection. RESULTS: In the bolus to constant infusion studies, equilibrium was established by 4-8 h. [(123)I]p ZIENT was 93% and 90% protein bound in the two baboons and there was no detection of lipophilic radiolabeled metabolites entering the brain. In the high-density serotonin transporter regions (diencephalon and brainstem), fenfluramine and citalopram resulted in 35-71% and 129-151% displacement, respectively, whereas methylphenidate and nisoxetine did not produce significant changes (<10%). CONCLUSION: These findings suggest that [(123)I]p ZIENT is a favorable compound for in vivo SPECT imaging of serotonin transporters with negligible binding to norepinephrine and dopamine transporters. Copyright © 2010 Elsevier Inc. All rights reserved.

INTRODUCTION: Single photon emission computed tomography (SPECT) imaging of the serotonin transporter (SERT) in the brain is a useful tool for examining normal physiological functions and disease states involving the serotonergic system. The goal of this study was to develop an improved SPECT radiotracer with faster kinetics than the current leading SPECT tracer, [(123)I]ADAM, for selective SERT imaging. METHODS: The in vitro binding affinities of (2-(2′-((dimethylamino)methyl)-4′-iodophenylthio)benzenamine) (FlipADAM) (1c), were determined using Hampshire pig kidney cells stably overexpressing the serotonin, norepinephrine (NET) or dopamine transporter (DAT). Localization of [(125)I]FlipADAM (1c) was evaluated through biodistribution and autoradiography in male Sprague Dawley rats, and the specificity of binding was assessed by injecting selective SERT or NET inhibitors prior to [(125)I]FlipADAM (1c). RESULTS: FlipADAM (1c) displayed a high binding affinity for SERT (K(i)=1.0 nM) and good selectivity over NET and DAT binding (43-fold and 257-fold, respectively). [(125)I]FlipADAM (1c) successfully penetrated the blood brain barrier, as evidenced by the brain uptake at 2 min (1.75% dose/g). [(125)I]FlipADAM(1c) also had a good target to non-target (hypothalamus/cerebellum) ratio of 3.35 at 60 min post-injection. In autoradiography studies, [(125)I]FlipADAM (1c) showed selective localization in SERT-rich brain regions such as the thalamic nuclei, amygdala, dorsal raphe nuclei and other areas. CONCLUSION: [(125)I]FlipADAM (1c) exhibited faster clearance from the brain and time to binding equilibrium when compared to [(125)I]2-(2′-((dimethylamino)methyl)-phenylthio)-5-iodophenylamine [(125)I]ADAM (1b) and a higher target to non-target ratio when compared to [(125)I]5-iodo-2-(2′-((dimethylamino)methyl)-phenylthio)benzyl alcohol [(125)I]IDAM (1a). Therefore, [(123)I]FlipADAM (1c) may be an improved SPECT tracer for imaging SERT. Copyright © 2010 Elsevier Inc. All rights reserved.

Abstract Carvacrol (5-isopropyl-2-methylphenol) is a monoterpenic phenol present in the essential oil of many plants. It is the major component of the essential oil fraction of oregano and thyme. In this study, the effect of carvacrol was investigated in two behavioral models, the forced swimming and tail suspension tests in mice, to investigate the possible antidepressant effect of this substance. Additionally, the mechanisms involved in the antidepressant-like effect of carvacrol in mice were also assessed. Carvacrol (cvc) was administered orally at single doses of 12.5, 25 and 50 mg/kg. The acute treatment of cvc decreased the immobility time in the forced swimming and tail suspension tests without accompanying changes in ambulation in the open-field test. The anti-immobility effect of carvacrol (25 mg/kg) was not prevented by pretreatment of mice with p-chlorophenylalanine, prazosin and yohimbine. On the other hand, the pretreatment of mice with SCH23390 or sulpiride completely blocked the antidepressant-like effect of carvacrol (25 mg/kg) in the forced swimming test. These results show that carvacrol presents antidepressant effects in the forced swimming and tail suspension tests; this effect seems to be dependent on its interaction with the dopaminergic system, but not with the serotonergic and noradrenergic systems. Keywords: Carvacrol; Antidepressant; Forced swimming; Tail suspension; Dopaminergic system.

Gene transfer of dopamine-synthesizing enzymes into the striatal neurons has led to behavioral recovery in animal models of Parkinson’s disease (PD). We evaluated the safety, tolerability, and potential efficacy of adeno-associated virus (AAV) vector-mediated gene delivery of aromatic L-amino acid decarboxylase (AADC) into the putamen of PD patients. Six PD patients were evaluated at baseline and at 6 months, using multiple measures, including the Unified Parkinson’s Disease Rating Scale (UPDRS), motor state diaries, and positron emission tomography (PET) with 6-[(18)F]fluoro-L-m-tyrosine (FMT), a tracer for AADC. The short-duration response to levodopa was measured in three patients. The procedure was well tolerated. Six months after surgery, motor functions in the OFF-medication state improved an average of 46% based on the UPDRS scores, without apparent changes in the short-duration response to levodopa. PET revealed a 56% increase in FMT activity, which persisted up to 96 weeks. Our findings provide class IV evidence regarding the safety and efficacy of AADC gene therapy and warrant further evaluation in a randomized, controlled, phase 2 setting.

Schizophrenia is a devastating neurodevelopmental disorder whose genetic influences remain elusive. Recent genome-wide scans revealed that rare structural variants disrupted multiple genes in neurodevelopmental pathways, which strongly implicate nitric oxide (NO) signaling in schizophrenia. NO acts as a second messenger of N-methyl-D aspartate receptor activation, which further interacts with both dopaminergic and serotonergic pathways. NO is mainly synthesized by neuronal nitric oxide synthase (NOS1) in the brain, and its gene locus, 12q24.2, has attracted much attention as a major linkage region for schizophrenia. Genetic variations of NOS1 have also been associated with schizophrenia, and differential expression of NOS1 was observed in the postmortem brain of schizophrenic patients. Here, we explored the hypothesis that a putative cis-acting G-84A single nucleotide polymorphism (SNP; rs41279104) in the exon 1c promoter region of the NOS1 gene is associated with the levels of NOS1 immunoreactivity in postmortem prefrontal cortex specimens regardless of disease phenotype. Individuals with the A-allele of this SNP showed significantly lower levels of NOS1 immunoreactivity than did GG homozygotes (p=0.002). Furthermore, a case-control study using 720 individuals in a Japanese population revealed a significant association between the SNP and schizophrenia (genotypic p=0.0013 and allelic p=0.0011). Additionally, the average of onset age in schizophrenic patients with the A-allele was significantly earlier than GG homozygotes (p=0.018). When the analyses took gender into account, this significance was more significant for female. These findings provide further evidences that NOS1 is associated with a biological susceptibility gene to schizophrenia. Copyright © 2010 Elsevier B.V. All rights reserved.

The aim of the present study was to investigate potential interrelationships between immune and neural elements of Peyer’s patches in normal pigs (n=8) and in pigs infected experimentally with Brachyspira hyodysenteriae and suffering from swine dysentery (n=8). Assessment of tissue concentration of neuropeptides by enzyme linked immunosorbent assay revealed increased levels of galanin (GAL) and substance P (SP) in samples from the infected animals. In contrast, concentrations of vasoactive intestinal polypeptide (VIP) and somatostatin (SOM) were similar in both groups. Immunohistochemistry demonstrated reactivity of nerve fibres with antibodies specific for dopamine beta hydroxylase, vesicular acetylcholine transporter, SOM, GAL, VIP and SP in the interfollicular region and peripheral areas of the Peyer’s patch lymphoid follicles. In the dysenteric pigs, the GAL-positive nerve fibres were more numerous and more intensely labelled than those in the normal animals. Flow cytometry revealed a decreased percentage of CD21(+) lymphocytes and lymphocytes expressing T-cell receptor (TCR)-gamma, with or without CD8 (TCR-gamma(+)CD8(-) and TCR-gamma(+)CD8(+)), in the dysenteric pigs as compared with the normal animals. Percentages of other lymphocyte subsets (CD2(+), CD4(+), CD5(+), CD8(+), CD5(-)CD8(+)) were comparable between the groups. Immunohistochemical investigations generally correlated with results obtained by flow cytometry related to lymphocyte subpopulations. Swine dysentery can therefore affect neuroimmunomodulatory processes in the ileal Peyer’s patch, in addition to the large intestine. GAL and SP may play a specific role in this neuroimmune cross-talk. Copyright © 2010 Elsevier Ltd. All rights reserved.

The mesocorticolimbic dopamine system, which governs components of reward and goal-directed behaviors, undergoes final maturation during adolescence. Adolescent social stress contributes to adult behavioral dysfunction, and is linked to adult psychiatric and addiction disorders. Here, behavioral, corticosterone, and limbic dopamine responses to amphetamine were examined in adult male rats previously exposed to repeated social defeat stress during mid-adolescence. Amphetamine (2.5 mg/kg, ip) was administered after a novel environment test, with behavior observed in the same context for 90 min thereafter. Adult rats that had been defeated in adolescence showed increased locomotion in the novel environment, but reduced amphetamine-induced locomotion relative to non-defeated age matched controls. Monoamine and corticosterone responses to amphetamine were examined following a second amphetamine injection 3 days later. In previously defeated rats, corticosterone and medial prefrontal cortex dopamine responses to amphetamine were blunted while dopamine responses in the nucleus accumbens core were elevated. Our results suggest that experience of social defeat stress during adolescent development can contribute to altered behavioral and endocrine responses to amphetamine in adulthood. Furthermore, these effects are paralleled by changes in amphetamine-induced dopamine responses in corticolimbic systems implicated in addiction disorders. Copyright © 2010. Published by Elsevier B.V.

Striatal D2-type dopamine receptors (D2Rs) have been implicated in the pathophysiology of neuropsychiatric disorders, including Parkinson’s disease and schizophrenia. Although these receptors regulate striatal synaptic plasticity, the mechanisms underlying dopaminergic modulation of glutamatergic synapses are unclear. We combined optogenetics, two-photon microscopy and glutamate uncaging to examine D2R-dependent modulation of glutamatergic synaptic transmission in mouse striatopallidal neurons. We found that D2R activation reduces corticostriatal glutamate release and attenuates both synaptic- and action potential-evoked Ca(2+) influx into dendritic spines by approximately 50%. Modulation of Ca(2+) signaling was mediated by a protein kinase A (PKA)-dependent regulation of Ca(2+) entry through NMDA-type glutamate receptors that was inhibited by D2Rs and enhanced by activation of 2A-type adenosine receptors (A2ARs). D2Rs also produced a PKA- and A2AR-independent reduction in Ca(2+) influx through R-type voltage-gated Ca(2+) channels. These findings reveal that dopamine regulates spine Ca(2+) by multiple pathways and that competitive modulation of PKA controls NMDAR-mediated Ca(2+) signaling in the striatum.

The BIO 14.6 hamster (DV), an animal model of limb-girdle muscular dystrophy, has elevated angiotensin AT1 receptors that may affect ventilation. Moreover, AT1 receptors may modulate expression of dopamine D1 receptors. We investigated if chronic treatment of BIO 14.6 hamsters (DL) with losartan, an AT1 receptor blocker, affects D1 receptor density in the striatum and nucleus tractus solitarius (NTS) and normalizes ventilation during exposure to air, hypoxia, following hypoxia, and hypercapnia, Ventilation was evaluated using plethysmography. Compared to the golden Syrian hamsters (GS), DV hamsters exhibited lower hypercapnic and hypoxic responsiveness and ventilation during hypercapnic exposure. Relative to GS, DL hamsters increased breathing frequency in air and maintained ventilation during hypercapnia. Post-hypoxic minute ventilation decline occurred in DV but not in DL or GS hamsters. DL hamsters exhibited higher D1 receptor density in the striatum and NTS relative to DV hamsters. Thus, in dystrophic hamsters chronic losartan treatment stimulated frequency of breathing and increased the density of D1 receptors. Copyright © 2010. Published by Elsevier B.V.

We explored the protective mechanisms of human neuronal mitochondrial uncoupling protein-5 (UCP5) in MPP(+)- and dopamine-induced toxicity after its stable overexpression in SH-SY5Y cells. We raised specific polyclonal antibodies. Overexpressed UCP5 localized in mitochondria but not in cytosol. UCP5 overexpression increased proton leak, decreased mitochondrial membrane potential (MMP), reduced ATP production, and increased overall oxygen consumption (demonstrating uncoupling activity). UCP5 overexpression did not affect other neuronal UCP expression (UCP2 and UCP4). Overexpressing UCP5 is protective against MPP(+)- and dopamine-induced toxicity. MPP(+) and dopamine exposure for 6h reduced MMP and increased superoxide levels. ATP levels in UCP5-overexpressing cells were preserved under MPP(+) and dopamine toxicity, comparable to levels in untreated vector controls. At 24h, UCP5 overexpression preserved MMP, ATP levels, and cell survival; attenuated superoxide generation; and maintained oxidative phosphorylation as indicated by lower lactate levels. MPP(+) and dopamine exposure induced UCP5 mRNA transcription but did not decrease transcript degradation, as inhibition of transcription by actinomycin-D abolished induction by either toxin. Compared with our previous studies on UCP4, we observed functional differences between UCP4 and UCP5 in enhancing mitochondrial efficiency. These neuronal UCP homologues may work synergistically to maintain oxidative balance (through uncoupling activities) and ATP production (by modifying MMP). Copyright © 2010. Published by Elsevier Inc.