Virus-Mediated shRNA Knockdown of Prodynorphin in the Rat Nucleus Accumbens Attenuates Depression-Like Behavior and Cocaine Locomotor Sensitization

Dynorphins, endogenous opioid peptides that arise from the precursor protein prodynorphin (Pdyn), are hypothesized to be involved in the regulation of mood states and the neuroplasticity associated with addiction. The current study tested the hypothesis that dynorphin in the nucleus accumbens (NAcc) mediates such effects. More specifically, we examined whether knockdown of Pdyn within the NAcc in rats would alter the expression of depressive-like and anxiety-like behavior, as well as cocaine locomotor sensitization. Wistar rats were injected with adeno-associated viral (AAV) vectors encoding either a Pdyn-specific short hairpin RNA (AAV-shPdyn) or a scrambled shRNA (AAV-shScr) as control. Four weeks later, rats were tested for anxiety-like behavior in the elevated plus maze test and depressive-like behavior in the forced swim test (FST). Finally, rats received one daily injection of saline or cocaine (20 mg/kg, i.p.), followed by assessment of locomotion for 4 consecutive days. Following 3 days of abstinence, the rats completed 2 additional daily cocaine/saline locomotor trials. Pdyn knockdown in the NAcc led to a significant reduction in depressive-like behavior in the FST, but had no effect on anxiety-like behavior in the elevated plus maze. Pdyn knockdown did not alter baseline locomotor behavior, the locomotor response to acute cocaine, or the initial sensitization of the locomotor response to cocaine over the first 4 cocaine treatment days. However, following 3 days abstinence the locomotor response to the cocaine challenge returned to their original levels in the AAV-shPdyn rats while remaining heightened in the AAV-shScr rats. These results suggest that dynorphin in a very specific area of the nucleus accumbens contributes to depressive-like states and may be involved in neuroadaptations in the NAcc that contribute to the development of cocaine addiction as a persistent and lasting condition.     

Protective efficacy of BCG overexpressing an L,D-transpeptidase against M. tuberculosis infection

Background

M. bovis Bacille Calmette-Guérin (BCG), currently the only available vaccine against tuberculosis (TB), fails to adequately protect individuals from active and latent TB infection. New vaccines are desperately needed to decrease the worldwide burden of TB.

Methods and Findings

We created a recombinant strain of BCG that overproduces an L,D-transpeptidase in order to alter the bacterial peptidoglycan layer and consequently increase the ability of this immunogen to protect against virulent M. tuberculosis (Mtb). We demonstrate that this novel recombinant BCG protects mice against virulent Mtb at least as well as control BCG, as measured by its ability to reduce bacterial burden in lungs and spleen, reduce lung histopathology, and prolong survival. A nutrient starved recombinant BCG preparation, while offering comparable protection, elicited a response characterized by elevated levels of select Th1 cytokines.

Conclusions

Recombinant BCG overexpressing a L,D-transpeptidase that is nutrient starved elicits a stronger Th1 type response and is at least as protective as parent BCG. Results from this study suggest that nutrient starvation treatment of live BCG vaccines should be further investigated as a way to increase host induction of Th-1 related cytokines in the development of experimental anti-TB vaccines.     

The Arabidopsis AtOPT3 protein functions in metal homeostasis and movement of iron to developing seeds

The Arabidopsis thaliana AtOPT3 belongs to the oligopeptide transporter (OPT) family, a relatively poorly characterized family of peptide/modified peptide transporters found in archebacteria, bacteria, fungi, and plants. A null mutation in AtOPT3 resulted in embryo lethality, indicating an essential role for AtOPT3 in embryo development. In this article, we report on the isolation and phenotypic characterization of a second AtOPT3 mutant line, opt3-2, harboring a T-DNA insertion in the 5' untranslated region of AtOPT3. The T-DNA insertion in the AtOPT3 promoter resulted in reduced but sufficient AtOPT3 expression to allow embryo formation in opt3-2 homozygous seeds. Phenotypic analyses of opt3-2 plants revealed three interesting loss-of-function phenotypes associated with iron metabolism. First, reduced AtOPT3 expression in opt3-2 plants resulted in the constitutive expression of root iron deficiency responses regardless of exogenous iron supply. Second, deregulation of root iron uptake processes in opt3-2 roots resulted in the accumulation of very high levels of iron in opt3-2 tissues. Hyperaccumulation of iron in opt3-2 resulted in the formation of brown necrotic areas in opt3-2 leaves and was more pronounced during the seed-filling stage. Third, reduced AtOPT3 expression resulted in decreased accumulation of iron in opt3-2 seeds. The reduced accumulation of iron in opt3-2 seeds is especially noteworthy considering the excessively high levels of accumulated iron in other opt3-2 tissues. AtOPT3, therefore, plays a critical role in two important aspects of iron metabolism, namely, maintenance of whole-plant iron homeostasis and iron nutrition of developing seeds.