Contribution of Extracellular Particles Isolated from Morus sp. (Mulberry) Fruit to Their Reported Protective Health Benefits: An In Vitro Study.

Morus sp. (mulberry) has a long tradition of use as a medicinal treatment, including for cardiovascular disease and type 2 diabetes, being shown to have antioxidant properties and to promote wound healing. Extracellular vesicles (EVs) are sub-micron, membrane-enclosed particles that were first identified in mammalian bodily fluids. EV-like particles have been described in plants (PDVs) and shown to have similar characteristics to mammalian EVs. We hypothesised that some of the health benefits previously attributed to the fruit of Morus sp. could be due to the release of PDVs. We isolated PDVs from Morus nigra and Morus alba via ultracentrifugation and incubated THP-1 monocytes, differentiated THP-1 macrophages, or HMEC-1 endothelial cells with pro-oxidant compounds DMNQ (THP-1) and glucose oxidase (HMEC-1) or lipopolysaccharide (LPS) in the presence of different fractions of mulberry EVs. Mulberry EVs augmented ROS production with DMNQ in THP-1 and caused the downregulation of ROS in HMEC-1. Mulberry EVs increased LPS-induced IL-1ß secretion but reduced CCL2 and TGF-ß secretion in THP-1 macrophages. In scratch wound assays, mulberry EVs inhibited HMEC-1 migration but increased proliferation in both low and high serum conditions, suggesting that they have opposing effects in these two important aspects of wound healing. One of the limitations of plant-derived therapeutics has been overcoming the low bioavailability of isolated compounds. We propose that PDVs could provide the link between physiological dose and therapeutic benefit by protecting plant active compounds in the GIT as well as potentially delivering genetic material or proteins that contribute to previously observed health benefits.

Ubiquitin-proteasomal regulation of chromatin remodeler INO80 in the nucleus accumbens mediates persistent cocaine craving.

Neuroadaptations in the nucleus accumbens (NAc) underlie cue-induced cocaine craving that intensifies ("incubates") during abstinence and is believed to contribute to persistent relapse vulnerability. Changes in gene expression often govern perpetual behavioral abnormalities, but epigenetic plasticity during prolonged abstinence from drug exposure is poorly understood. We examined how E3 ubiquitin ligase TRIM3 dysregulates chromatin remodeler INO80 to mediate cocaine craving during prolonged abstinence. We found that INO80 expression increased in the NAc on abstinence day 30 (AD30) but not on AD1 following cocaine self-administration. Furthermore, TRIM3, which mediates degradation of INO80, was reduced on AD30, along with TRIM3-INO80 interaction. Viral-mediated gene transfer of INO80 or TRIM3 governed cocaine craving during prolonged abstinence. Lastly, chromatin immunoprecipitation followed by massively parallel DNA sequencing identified INO80-mediated transcriptional regulation of predicted pathways associated with cocaine plasticity. Together, these results demonstrate a novel ubiquitin-proteasomal-epigenetic mechanism by which TRIM3-INO80 mediates cocaine craving during prolonged abstinence.

MEF2C transcription factor is associated with the genetic and epigenetic risk architecture of schizophrenia and improves cognition in mice.

Large-scale consortia mapping the genomic risk architectures of schizophrenia provide vast amounts of molecular information, with largely unexplored therapeutic potential. We harnessed publically available information from the Psychiatric Genomics Consortium, and report myocyte enhancer factor 2C (MEF2C) motif enrichment in sequences surrounding the top scoring single-nucleotide polymorphisms within risk loci contributing by individual small effect to disease heritability. Chromatin profiling at base-pair resolution in neuronal nucleosomes extracted from prefrontal cortex of 34 subjects, including 17 cases diagnosed with schizophrenia, revealed MEF2C motif enrichment within cis-regulatory sequences, including neuron-specific promoters and superenhancers, affected by histone H3K4 hypermethylation in disease cases. Vector-induced short- and long-term Mef2c upregulation in mouse prefrontal projection neurons consistently resulted in enhanced cognitive performance in working memory and object recognition paradigms at baseline and after psychotogenic drug challenge, in conjunction with remodeling of local connectivity. Neuronal genome tagging in vivo by Mef2c-Dam adenine methyltransferase fusion protein confirmed the link between cognitive enhancement and MEF2C occupancy at promoters harboring canonical and variant MEF2C motifs. The multilayered integrative approaches presented here provide a roadmap to uncover the therapeutic potential of transcriptional regulators for schizophrenia and related disorders.

The dendritic spine morphogenic effects of repeated cocaine use occur through the regulation of serum response factor signaling.

The nucleus accumbens (NAc) is a primary brain reward region composed predominantly of medium spiny neurons (MSNs). In response to early withdrawal from repeated cocaine administration, de novo dendritic spine formation occurs in NAc MSNs. Much evidence indicates that this new spine formation facilitates the rewarding properties of cocaine. Early withdrawal from repeated cocaine also produces dramatic alterations in the transcriptome of NAc MSNs, but how such alterations influence cocaine's effects on dendritic spine formation remain unclear. Studies in non-neuronal cells indicate that actin cytoskeletal regulatory pathways in nuclei have a direct role in the regulation of gene transcription in part by controlling the access of co-activators to their transcription factor partners. In particular, actin state dictates the interaction between the serum response factor (SRF) transcription factor and one of its principal co-activators, MAL. Here we show that cocaine induces alterations in nuclear F-actin signaling pathways in the NAc with associated changes in the nuclear subcellular localization of SRF and MAL. Using in vivo optogenetics, the brain region-specific inputs to the NAc that mediate these nuclear changes are investigated. Finally, we demonstrate that regulated SRF expression, in turn, is critical for the effects of cocaine on dendritic spine formation and for cocaine-mediated behavioral sensitization. Collectively, these findings reveal a mechanism by which nuclear-based changes influence the structure of NAc MSNs in response to cocaine.

Opposing actions of the synapse-associated protein of 97-kDa molecular weight (SAP97) and Disrupted in Schizophrenia 1 (DISC1) on Wnt/ß-catenin signaling.

It has been suggested that synapse-associated protein of 97-kDa molecular weight (SAP97) is a susceptibility factor for childhood and adult neuropsychiatric disorders. SAP97 is a scaffolding protein that shares direct and indirect binding partners with the Disrupted in Schizophrenia 1 (DISC1) gene product, a gene with strong association with neuropsychiatric disorders. Here we investigated the possibility that these two proteins converge upon a common molecular pathway. Since DISC1 modifies Wnt/ß-catenin signaling via changes in glycogen synthase kinase 3 beta (GSK3ß) phosphorylation, we asked if SAP97 impacts Wnt/ß-catenin signaling and GSK3ß phosphorylation. We find that SAP97 acts as inhibitor of Wnt signaling activity and can suppress the stimulatory effects of DISC1 on ß-catenin transcriptional activity. Reductions in SAP97 abundance also decrease GSK3ß phosphorylation. In addition, we find that over expression of DISC1 leads to an increase in the abundance of SAP97, by inhibiting its proteasomal degradation. Our findings suggest that SAP97 and DISC1 contribute to maintaining Wnt/ß-catenin signaling activity within a homeostatic range by regulating GSK3ß phosphorylation.

Normalizing dopamine D2 receptor-mediated responses in D2 null mutant mice by virus-mediated receptor restoration: comparing D2L and D2S.

D2 receptor null mutant (Drd2(-/-)) mice have altered responses to the rewarding and locomotor effects of psychostimulant drugs, which is evidence of a necessary role for D2 receptors in these behaviors. Furthermore, work with mice that constitutively express only the D2 receptor short form (D2S), as a result of genetic deletion of the long form (D2L), provides the basis for a current model in which D2L is thought to be the postsynaptic D2 receptor on medium spiny neurons in the basal forebrain, and D2S the autoreceptor that regulates the activity of dopamine neurons and dopamine synthesis and release. Because constitutive genetic deletion of the D2 or D2L receptor may cause compensatory changes that influence functional outcomes, our approach is to identify aspects of the abnormal phenotype of a Drd2(-/-) mouse that can be normalized by virus-mediated D2 receptor expression. Drd2(-/-) mice are deficient in basal and methamphetamine-induced locomotor activation and lack D2 receptor agonist-induced activation of G protein-regulated inward rectifying potassium channels (GIRKs) in dopaminergic neurons. Here we show that virus-mediated expression of D2L in the nucleus accumbens significantly restored methamphetamine-induced locomotor activation, but not basal locomotor activity, compared to mice receiving the control virus. It also restored the effect of methamphetamine to decrease time spent in the center of the activity chamber in female but not male Drd2(-/-) mice. Furthermore, the effect of expression of D2S was indistinguishable from D2L. Similarly, virus-mediated expression of either D2S or D2L in substantia nigra neurons restored D2 agonist-induced activation of GIRKs. In this acute expression system, the alternatively spliced forms of the D2 receptor appear to be equally capable of acting as postsynaptic receptors and autoreceptors.

SAMITAL® improves chemo/radiotherapy-induced oral mucositis in patients with head and neck cancer: results of a randomized, placebo-controlled, single-blind Phase II study.

PURPOSE: This randomised, placebo-controlled single-blind trial investigated the safety and efficacy of SAMITAL®, a formulation of highly standardised botanical extracts, in the treatment of chemo/radiotherapy-induced oral mucositis (OM) in patients with head and neck cancer. METHODS: Patients received SAMITAL® or placebo four times daily for up to 50 days during scheduled chemo/radiotherapy. Severity of OM was monitored according to a modified WHO severity scale, and pain and quality-of-life assessments were based on the effect of symptoms of OM on relevant daily activities, according to a visual analogue scale. RESULTS: Mean scores for the severity of OM were significantly (p < 0.05 versus baseline) reduced from day 31 until the end of treatment in patients treated with SAMITAL® (n = 20). No significant improvement was observed in the placebo group (n = 10). Pain reduction was significant from day 4 till end of treatment with SAMITAL® and from days 7 to 21 in placebo patients. SAMITAL® also significantly improved quality of life, as shown by improvements in scores for relevant daily activities including eating, drinking and sleeping. All SAMITAL® patients completed the treatment period, but no placebo recipients completed treatment. No severe adverse events were observed with SAMITAL®, and systemic absorption of relevant active ingredients was undetectable. CONCLUSIONS: SAMITAL® significantly decreased the severity of chemo/radiotherapy-induced OM in patients with head and neck cancer, with no treatment-related adverse events. Pain relief lasted through the treatment period, and improvements in quality of life were reflected by the significant benefits of SAMITAL® on activities like drinking, eating and speaking.

The nucleus accumbens 5-HTR4-CART pathway ties anorexia to hyperactivity.

In mental diseases, the brain does not systematically adjust motor activity to feeding. Probably, the most outlined example is the association between hyperactivity and anorexia in Anorexia nervosa. The neural underpinnings of this 'paradox', however, are poorly elucidated. Although anorexia and hyperactivity prevail over self-preservation, both symptoms rarely exist independently, suggesting commonalities in neural pathways, most likely in the reward system. We previously discovered an addictive molecular facet of anorexia, involving production, in the nucleus accumbens (NAc), of the same transcripts stimulated in response to cocaine and amphetamine (CART) upon stimulation of the 5-HT(4) receptors (5-HTR(4)) or MDMA (ecstasy). Here, we tested whether this pathway predisposes not only to anorexia but also to hyperactivity. Following food restriction, mice are expected to overeat. However, selecting hyperactive and addiction-related animal models, we observed that mice lacking 5-HTR(1B) self-imposed food restriction after deprivation and still displayed anorexia and hyperactivity after ecstasy. Decryption of the mechanisms showed a gain-of-function of 5-HTR(4) in the absence of 5-HTR(1B), associated with CART surplus in the NAc and not in other brain areas. NAc-5-HTR(4) overexpression upregulated NAc-CART, provoked anorexia and hyperactivity. NAc-5-HTR(4) knockdown or blockade reduced ecstasy-induced hyperactivity. Finally, NAc-CART knockdown suppressed hyperactivity upon stimulation of the NAc-5-HTR(4). Additionally, inactivating NAc-5-HTR(4) suppressed ecstasy's preference, strengthening the rewarding facet of anorexia. In conclusion, the NAc-5-HTR(4)/CART pathway establishes a 'tight-junction' between anorexia and hyperactivity, suggesting the existence of a primary functional unit susceptible to limit overeating associated with resting following homeostasis rules.

Reduced activity of AMP-activated protein kinase protects against genetic models of motor neuron disease.

A growing body of research indicates that amyotrophic lateral sclerosis (ALS) patients and mouse models of ALS exhibit metabolic dysfunction. A subpopulation of ALS patients possesses higher levels of resting energy expenditure and lower fat-free mass compared to healthy controls. Similarly, two mutant copper zinc superoxide dismutase 1 (mSOD1) mouse models of familial ALS possess a hypermetabolic phenotype. The pathophysiological relevance of the bioenergetic defects observed in ALS remains largely elusive. AMP-activated protein kinase (AMPK) is a key sensor of cellular energy status and thus might be activated in various models of ALS. Here, we report that AMPK activity is increased in spinal cord cultures expressing mSOD1, as well as in spinal cord lysates from mSOD1 mice. Reducing AMPK activity either pharmacologically or genetically prevents mSOD1-induced motor neuron death in vitro. To investigate the role of AMPK in vivo, we used Caenorhabditis elegans models of motor neuron disease. C. elegans engineered to express human mSOD1 (G85R) in neurons develops locomotor dysfunction and severe fecundity defects when compared to transgenic worms expressing human wild-type SOD1. Genetic reduction of aak-2, the ortholog of the AMPK α2 catalytic subunit in nematodes, improved locomotor behavior and fecundity in G85R animals. Similar observations were made with nematodes engineered to express mutant tat-activating regulatory (TAR) DNA-binding protein of 43 kDa molecular weight. Altogether, these data suggest that bioenergetic abnormalities are likely to be pathophysiologically relevant to motor neuron disease.

ZNF217, a candidate breast cancer oncogene amplified at 20q13, regulates expression of the ErbB3 receptor tyrosine kinase in breast cancer cells.

Understanding the mechanisms underlying ErbB3 overexpression in breast cancer will facilitate the rational design of therapies to disrupt ErbB2-ErbB3 oncogenic function. Although ErbB3 overexpression is frequently observed in breast cancer, the factors mediating its aberrant expression are poorly understood. In particular, the ErbB3 gene is not significantly amplified, raising the question as to how ErbB3 overexpression is achieved. In this study we showed that the ZNF217 transcription factor, amplified at 20q13 in ∼20% of breast tumors, regulates ErbB3 expression. Analysis of a panel of human breast cancer cell lines (n = 50) and primary human breast tumors (n = 15) showed a strong positive correlation between ZNF217 and ErbB3 expression. Ectopic expression of ZNF217 in human mammary epithelial cells induced ErbB3 expression, whereas ZNF217 silencing in breast cancer cells resulted in decreased ErbB3 expression. Although ZNF217 has previously been linked with transcriptional repression because of its close association with C-terminal-binding protein (CtBP)1/2 repressor complexes, our results show that ZNF217 also activates gene expression. We showed that ZNF217 recruitment to the ErbB3 promoter is CtBP1/2-independent and that ZNF217 and CtBP1/2 have opposite roles in regulating ErbB3 expression. In addition, we identify ErbB3 as one of the mechanisms by which ZNF217 augments PI-3K/Akt signaling.

Transient viral-mediated overexpression of alpha-calcium/calmodulin-dependent protein kinase II in the nucleus accumbens shell leads to long-lasting functional upregulation of alpha-amino-3-hydroxyl-5-methyl-4-isoxazole-propionate receptors: dopamine type-1 receptor and protein kinase A dependence.

Calcium/calmodulin-dependent protein kinase II (CaMKII) activity is necessary for the long-lasting expression of locomotor sensitization and enhanced drug-taking observed in rats previously exposed to psychostimulants. Exposure to these drugs also transiently increases alphaCaMKII levels in the nucleus accumbens (NAcc), an effect that, when mimicked by transient viral-mediated overexpression of alphaCaMKII in NAcc shell neurons, leads to long-lasting enhancement in locomotor responding to amphetamine and NAcc alpha-amino-3-hydroxyl-5-methyl-4-isoxazole-propionate (AMPA). The present experiments characterized the dopamine (DA) dependence of the functional AMPA receptor upregulation observed long after transient overexpression of alphaCaMKII. Rats infected with herpes simplex virus-alphaCaMKII in the NAcc shell showed a transient increase in alphaCaMKII levels that peaked at 4 days post-infection and returned to baseline 8 days later. When challenged with AMPA (0.8 nmol/side) in the NAcc shell at 20 days post-infection, these rats showed enhanced locomotion compared with controls. This sensitized locomotor response was blocked when AMPA was coinfused with either the DA type-1 receptor antagonist SCH23390 (0.8 nmol/side) or the protein kinase A inhibitor Rp-cAMPS (80 nmol/side). Neither SCH23390 nor Rp-cAMPS produced locomotor effects when infused by itself into the NAcc shell. Furthermore, these antagonists did not block the acute non-sensitized locomotor response to AMPA observed in control rats. These findings show that transient viral-mediated overexpression of alphaCaMKII in neurons of the NAcc shell leads to long-lasting functional upregulation of AMPA receptors that is DA type-1 receptor and protein kinase A dependent. Thus, transient increases in levels of alphaCaMKII in the NAcc shell produce long-lasting changes in the way that DA and glutamate interact in this site to generate locomotor behavior.

Epithelial mesenchymal transition traits in human breast cancer cell lines.

Epithelial mesenchymal transition (EMT) has long been associated with breast cancer cell invasiveness and evidence of EMT processes in clinical samples is growing rapidly. Genome-wide transcriptional profiling of increasingly larger numbers of human breast cancer (HBC) cell lines have confirmed the existence of a subgroup of cell lines (termed Basal B/Mesenchymal) with enhanced invasive properties and a predominantly mesenchymal gene expression signature, distinct from subgroups with predominantly luminal (termed Luminal) or mixed basal/luminal (termed Basal A) features (Neve et al Cancer Cell 2006). Studies providing molecular and cellular analyses of EMT features in these cell lines are summarised, and the expression levels of EMT-associated factors in these cell lines are analysed. Recent clinical studies supporting the presence of EMT-like changes in vivo are summarised. Human breast cancer cell lines with mesenchymal properties continue to hold out the promise of directing us towards key mechanisms at play in the metastatic dissemination of breast cancer.

Modification in the technique of pancreaticoduodenectomy: supracolic division of jejunum to facilitate uncinate process dissection.

Pancreaticoduodenectomy remains the recommended procedure for periampullary and pancreatic head tumors. The dissection of the uncinate process from the superior mesenteric vessels is a key step in this surgery. We describe a modification in the existing practice of infracolic division of the jejunum in order to facilitate this step. In this modification, the duodenojejunal (DJ) flexure and the proximal jejunum are delivered into the supracolic compartment and then the jejunum is divided. This exposes the uncinate process completely and facilitates the separation from the Superior Mesenteric Artery (SMA) and the Superior Mesenteric Vein (SMV). We have successfully employed this modified technique for 33 resections since February 2004. This modification of dividing the jejunum in the supracolic compartment is based on sound anatomic and embryologic grounds. It helps in aligning the uncinate process with the jejunal mesentery thereby making the dissection of uncinate process from the superior mesenteric vessels safe and complete.

Is there a role for estrogen and progesterone receptors in gall bladder cancer?

BACKGROUND/AIMS: The concept of metaplastic and non-metaplastic types of gall bladder cancer and the likelihood of hormone receptor expression in the nuclei of tumour cells raised the possibility of a potential role for anti-estrogen therapy in gall bladder cancer. This study was carried out to determine the hormone receptors (ER/PR) expression level in gall bladder cancer using specific immunohistochemical assays and correlate it with patient and tumour histopathological characteristics. PATIENTS AND METHODS: Histopathological tumour specimens of 62 patients who underwent a radical cholecystectomy were analysed. Pronase pretreatment and primary monoclonal antibodies were used to perform immunohistochemical analysis for ER and PR. RESULTS: The histology was adenocarcinoma--predominantly, moderately to poorly differentiated (91%). Gallstones were present in 90% of the individuals. Of the 62 specimens analysed, 62 (100%) and 61 (98%) were negative for ER and PR, respectively. CONCLUSION: The high incidence of gallstone-related gall bladder cancer in India is associated with metaplasia and a tendency to poorer differentiation in the tumour histology. These tumours are consequently less likely to express hormone receptors. Thus, there does not seem to be a role for anti-hormone therapy in patients with histogenesis similar to that seen in India.

beta-Secretase cleavage of the amyloid precursor protein mediates neuronal apoptosis caused by familial Alzheimer's disease mutations.

The amyloid precursor protein (APP) is cleaved by two enzymes, beta-secretase and gamma-secretase, to generate the pathological amyloid beta (Abeta) peptide. Expression of familial Alzheimer's disease (FAD) mutants of APP in primary neurons causes both intracellular accumulation of the C-terminal beta-secretase cleavage product of APP and increased secretion of Abeta, and eventually results in apoptotic death of the cells. To determine whether either of these two processing products of APP is involved in this apoptotic pathway, we first modeled experimentally the accumulation of the beta-secretase cleavage product in neurons. The C-terminal 100 amino acids (C100) of APP, with and without a signal peptide, was expressed in cells via recombinant herpes simplex virus (HSV) vectors. Both transgene products were targeted to the membrane, and both caused apoptosis in the neurons, implicating the beta-secretase cleavage product of APP in apoptosis caused by FAD APPs. Expression in neurons of a mutant of FAD APP that inhibited beta-secretase cleavage inhibited its ability to cause apoptosis. However, expression in neurons of a mutant of FAD APP that inhibited gamma-secretase cleavage did not inhibit the ability of this mutant to cause apoptosis. These data suggested that the C-terminal beta-secretase cleavage product of APP, but not Abeta, mediates the apoptosis caused by FAD mutants of APP. Consistent with this hypothesis, C31, which is generated from the beta-secretase cleavage product, itself caused neuronal apoptosis. Inhibitors of caspases 3, 6 and 8, but not of caspase 9, inhibited the apoptosis caused by FAD mutants of APP. It may be inferred from these data that beta-secretase cleavage of FAD mutants of APP allows the appropriate caspase access to its site of action to produce C31, which directly causes neuronal apoptosis.

Dopamine D2 receptor-induced heterologous sensitization of adenylyl cyclase requires Galphas: characterization of Galphas-insensitive mutants of adenylyl cyclase V.

Whereas acute stimulation of Galphai/o-coupled receptors inhibits the activity of adenylyl cyclase, a delayed consequence of persistent activation of the receptors is heterologous sensitization, an enhanced responsiveness of adenylyl cyclase to activators such as forskolin or agonists of Galphas-coupled receptors. Galphas-insensitive mutants of adenylyl cyclase type V were used to test the hypothesis that heterologous sensitization requires Galphas-dependent activation of adenylyl cyclase. When adenylyl cyclase was stably expressed in human embryonic kidney (HEK) 293 cells with the D2L dopamine receptor, basal, forskolin-stimulated, and isoproterenol-stimulated cyclic AMP accumulation were all enhanced by 2-h pretreatment with the D2 receptor agonist quinpirole. Transient expression of wild-type adenylyl cyclase and three Galphas-insensitive mutants (F379L, R1021Q, and F1093S) in HEK293 cells stably expressing the D2L receptor demonstrated that all three mutants had little or no responsiveness to beta-adrenergic receptor-mediated activation of Galphas but that the mutants retained sensitivity to forskolin and to D2L receptor-mediated inhibition. Transiently expressed adenylyl cyclase V was robustly sensitized by 2-h pretreatment with quinpirole. In contrast, the Galphas-insensitive mutants displayed no sensitization of forskolin-stimulated cyclic AMP accumulation, indicating that responsiveness to Galphas is required for the expression of heterologous sensitization.

Altered responsiveness to cocaine and increased immobility in the forced swim test associated with elevated cAMP response element-binding protein expression in nucleus accumbens.

Drugs of abuse regulate the transcription factor cAMP response element-binding protein (CREB) in striatal regions, including the nucleus accumbens (NAc). To explore how regulation of CREB in the NAc affects behavior, we used herpes simplex virus (HSV) vectors to elevate CREB expression in this region or to overexpress a dominant-negative mutant CREB (mCREB) that blocks CREB function. Rats treated with HSV-mCREB in place conditioning studies spent more time in environments associated with cocaine, indicating increased cocaine reward. Conversely, rats treated with HSV-CREB spent less time in cocaine-associated environments, indicating increased cocaine aversion. Studies in which drug-environment pairings were varied to coincide with either the early or late effects of cocaine suggest that CREB-associated place aversions reflect increased cocaine withdrawal. Because cocaine withdrawal can be accompanied by symptoms of depression, we examined how altered CREB function in the NAc affects behavior in the forced swim test (FST). Elevated CREB expression increased immobility in the FST, an effect that is opposite to that caused by standard antidepressants and is consistent with a link between CREB and dysphoria. Conversely, overexpression of mCREB decreased immobility, an effect similar to that caused by antidepressants. Moreover, the kappa opioid receptor antagonist nor-Binaltorphimine decreased immobility in HSV-CREB- and HSV-mCREB-treated rats, suggesting that CREB-mediated induction of dynorphin (an endogenous kappa receptor ligand) contributes to immobility behavior in the FST. Exposure to the FST itself dramatically increased CREB function in the NAc. These findings raise the possibility that CREB-mediated transcription within the NAc regulates dysphoric states.