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RATIONALE: Neuropeptide S (NPS) displays unique pharmacological properties and induces both anxiolytic and pro-stress/arousal activities. Previous studies performed using Wistar rats demonstrated that NPS facilitated alcohol and cocaine seeking but did not affect alcohol or cocaine consumption. OBJECTIVES: Here, we investigated the effects of NPS in Marchigian Sardinian alcohol-preferring (msP) rats, a rat strain characterized by excessive alcohol consumption comorbid with heightened anxiety and depressive-like phenotypes. Specifically, we evaluated the effect of NPS on operant alcohol self-administration by msP rats compared to Wistar rats. The effect of NPS on cue-induced reinstatement of alcohol seeking in msP rats was also evaluated. Finally, using the open field test (OFT) and the elevated plus maze (EPM), we evaluated the effects of NPS on locomotor activity and anxiety. RESULTS: NPS reduced alcohol self-administration but did not affect cue-induced reinstatement in the msP rat. In addition, NPS induced reinstatement of extinguished alcohol seeking in Wistar rats without affecting alcohol intake. In the EPM task, NPS, in accordance with its anxiolytic activity, increased the time spent in the open arm of the arena by msP rats, although this effect was not observed in Wistar rats. CONCLUSIONS: These data suggest that the effect of NPS is strongly influenced by the genetic background of the animal. In Wistar rats, NPS acts as a pro-arousal agent to promote the reinstatement of alcohol seeking. However, when alcohol drinking is motivated by or associated with a state of pathological anxiety, NPS attenuates alcohol consumption and seeking due to its anxiolytic activity.

Corticotropin releasing factor (CRF) is the primary mediator of stress responses, and nociceptin/orphanin FQ (N/OFQ) plays an important role in the modulation of these stress responses. Thus, in this multidisciplinary study, we explored the relationship between the N/OFQ and the CRF systems in response to stress. Using in situ hybridization (ISH), we assessed the effect of body restraint stress on the gene expression of CRF and N/OFQ-related genes in various subdivisions of the amygdala, a critical brain structure involved in the modulation of stress response and anxiety-like behaviors. We found a selective upregulation of the NOP and downregulation of the CRF1 receptor transcripts in the CeA and in the BLA after body restraint. Thus, we performed intracellular electrophysiological recordings of GABAA-mediated IPSPs in the central nucleus of the amygdala (CeA) to explore functional interactions between CRF and N/OFQ systems in this brain region. Acute application of CRF significantly increased IPSPs in the CeA, and this enhancement was blocked by N/OFQ. Importantly, in stress-restraint rats, baseline CeA GABAergic responses were elevated and N/OFQ exerted a larger inhibition of IPSPs compared with unrestraint rats. The NOP antagonist [Nphe1]-nociceptin(1-13)NH2 increased the IPSP amplitudes in restraint rats but not in unrestraint rats, suggesting a functional recruitment of the N/OFQ system after acute stress. Finally, we evaluated the anxiety-like response in rats subjected to restraint stress and nonrestraint rats after N/OFQ microinjection into the CeA. Intra-CeA injections of N/OFQ significantly and selectively reduced anxiety-like behavior in restraint rats in the elevated plus maze. These combined results demonstrate that acute stress increases N/OFQ systems in the CeA and that N/OFQ has antistress properties.

Evidence shows that the endocannabinoid system modulates the addictive properties of nicotine. In the present study, we hypothesized that spontaneous withdrawal resulting from removal of chronically implanted transdermal nicotine patches is regulated by the endocannabinoid system. A 7-day nicotine dependence procedure (5.2 mg/rat/day) elicited occurrence of reliable nicotine abstinence symptoms in Wistar rats. Somatic and affective withdrawal signs were observed at 16 and 34 hours following removal of nicotine patches, respectively. Further behavioral manifestations including decrease in locomotor activity and increased weight gain also occurred during withdrawal. Expression of spontaneous nicotine withdrawal was accompanied by fluctuation in levels of the endocannabinoid anandamide (AEA) in several brain structures including the amygdala, the hippocampus, the hypothalamus and the prefrontal cortex. Conversely, levels of 2-arachidonoyl-sn-glycerol were not significantly altered. Pharmacological inhibition of fatty acid amide hydrolase (FAAH), the enzyme responsible for the intracellular degradation of AEA, by URB597 (0.1 and 0.3 mg/kg, i.p.), reduced withdrawal-induced anxiety as assessed by the elevated plus maze test and the shock-probe defensive burying paradigm, but did not prevent the occurrence of somatic signs. Together, the results indicate that pharmacological strategies aimed at enhancing endocannabinoid signaling may offer therapeutic advantages to treat the negative affective state produced by nicotine withdrawal, which is critical for the maintenance of tobacco use.

BackgroundEndocrine disruptors include plasticizers, pesticides, detergents, and pharmaceuticals. Turbot and other flatfish are used to characterize the presence of chemicals in the marine environment. Unfortunately, there are relatively few genes of turbot and other flatfish in GenBank, which limits the use of molecular tools such as microarrays and quantitative reverse-transcriptase polymerase chain reaction (qRT-PCR) to study disruption of endocrine responses in sentinel fish captured by regulatory agencies.ObjectivesWe fabricated a multigene cross-species microarray as a diagnostic tool to screen the effects of environmental chemicals in fish, for which there is minimal genomic information. The array included genes that are involved in the actions of adrenal and sex steroids, thyroid hormone, and xenobiotic responses. This microarray will provide a sensitive tool for screening for the presence of chemicals with adverse effects on endocrine responses in coastal fish species.MethodsWe used a custom multispecies microarray to study gene expression in wild hornyhead turbot (Pleuronichthys verticalis) collected from polluted and clean coastal waters and in laboratory male zebrafish (Danio rerio) after exposure to estradiol and 4-nonylphenol. We measured gene-specific expression in turbot liver by qRT-PCR and correlated it to microarray data.ResultsMicroarray and qRT-PCR analyses of livers from turbot collected from polluted areas revealed altered gene expression profiles compared with those from nonaffected areas.ConclusionsThe agreement between the array data and qRT-PCR analyses validates this multispecies microarray. The microarray measurement of gene expression in zebrafish, which are phylogenetically distant from turbot, indicates that this multispecies microarray will be useful for measuring endocrine responses in other fish.

The A2A adenosine receptor (A2AAR) is one of the four subtypes activated by nucleoside adenosine, and the molecules able to selectively counteract its action are attractive tools for neurodegenerative disorders. In order to find novel A2AAR ligands, two series of compounds based on purine and triazolotriazine scaffolds were synthesized and tested at ARs. Compound 13 was also tested in an in vitro model of neuroinflammation. Some compounds were found to possess high affinity for A2AAR, and it was observed that compound 13 exerted anti-inflammatory properties in microglial cells. Molecular modeling studies results were in good agreement with the binding affinity data and underlined that triazolotriazine and purine scaffolds are interchangeable only when 5- and 2-positions of the triazolotriazine moiety (corresponding to the purine 2- and 8-positions) are substituted. [ABSTRACT FROM AUTHOR]

KEY WORDS papyri; ancient DNA; racemization; human mummies; Egypt The writing sheets made with strips from the stem (caulis) of papyri (Cyperus papyrus) are one of the most ingenious products of ancient technology. We extracted DNA from samples of modern papyri varying in age from 0-100 years BP and from ancient specimens from Egypt, with an age-span from 1,300-3,200 years BP. The copy number of the plant chloroplast DNA in the sheets was determined using a competitive PCR system designed on the basis of a short (90 bp) tract of the chloroplast's ribulose bisphosphate carboxylase large subunit (rbcL) gene sequence. The results allowed us to establish that the DNA half-life in papyri is about 19-24 years. This means that the last DNA fragments will vanish within no more than 532-672 years from the sheets being manufactured. In a parallel investigation, we checked the archaeological specimens for the presence of residual DNA and determined the extent of racemization of aspartic (Asp) acid in both modern and ancient specimens, as a previous report (Poinar et al. [1996], Science 272:864-866) showed that racemization of aspartic acid and DNA decay are linked. The results confirmed the complete loss of authentic DNA, even in the less ancient (8th century AD) papyri. On the other hand, when the regression for Asp racemization rates in papyri was compared with that for human and animal remains from Egyptian archaeological sites, it proved, quite surprisingly, that the regressions are virtually identical. Our study provides an indirect argument against the reliability of claims about the recovery of authentic DNA from Egyptian mummies and bone remains. Am J Phys Anthropol 117:310-318, 2002.