Dopaminergic Modulation of Effort-Related Choice Behavior as Assessed by a Progressive Ratio Chow Feeding Choice Task: Pharmacological Studies and the Role of Individual Differences

Mesolimbic dopamine (DA) is involved in behavioral activation and effort-related processes. Rats with impaired DA transmission reallocate their instrumental behavior away from food-reinforced tasks with high response requirements, and instead select less effortful food-seeking behaviors. In the present study, the effects of several drug treatments were assessed using a progressive ratio (PROG)/chow feeding concurrent choice task. With this task, rats can lever press on a PROG schedule reinforced by a preferred high-carbohydrate food pellet, or alternatively approach and consume the less-preferred but concurrently available laboratory chow. Rats pass through each ratio level 15 times, after which the ratio requirement is incremented by one additional response. The DA D₂ antagonist haloperidol (0.025–0.1 mg/kg) reduced number of lever presses and highest ratio achieved but did not reduce chow intake. In contrast, the adenosine A_2A antagonist MSX-3 increased lever presses and highest ratio achieved, but decreased chow consumption. The cannabinoid CB1 inverse agonist and putative appetite suppressant AM251 decreased lever presses, highest ratio achieved, and chow intake; this effect was similar to that produced by pre-feeding. Furthermore, DA-related signal transduction activity (pDARPP-32(Thr34) expression) was greater in nucleus accumbens core of high responders (rats with high lever pressing output) compared to low responders. Thus, the effects of DA antagonism differed greatly from those produced by pre-feeding or reduced CB1 transmission, and it appears unlikely that haloperidol reduces PROG responding because of a general reduction in primary food motivation or the unconditioned reinforcing properties of food. Furthermore, accumbens core signal transduction activity is related to individual differences in work output.     

Importance of pH and disulfide bridges on the structural and binding properties of human α₁-acid glycoprotein

Human α₁-acid glycoprotein (AGP) is an acute phase plasma glycoprotein containing two disulfide bridges. As a member of the lipocalin superfamily, it binds and transports several basic and neutral ligands, but a number of other activities have also been described. Thanks to its binding properties, AGP is also a good candidate for the development of biosensors and affinity chromatography media, and in this context detailed structural information is needed. The structural properties of AGP at different p²Hs and under reducing conditions were analysed by FT-IR spectroscopy. The obtained data indicate that AGP, when denatured, does not aggregate at neutral or basic p²Hs whilst it does at acidic p²Hs. Under reducing conditions the protein is remarkably less thermostable than its oxidized counterpart and presents an enhanced tendency to aggregate, even at neutral p²H. A heat-induced molten globule-like state (MG) was detected at 55 °C at p²H 7.4 and 5.5. At p²H 4.5 the MG occurred at 45 °C with an onset of formation at 40 °C. The MG was not observed under reducing conditions. A lower affinity of chlorpromazine and progesterone for the MG formed at p²H 4.5 and 40 °C was observed, suggesting that ligand(s) may be released near the negative surfaces of biological membranes. Furthermore, the reduced AGP displays an enhanced affinity for progesterone, indicating the importance of disulfide bonds for the binding capacity of AGP.     

P-gp localization in mitochondria and its functional characterization in multiple drug-resistant cell lines

Multidrug resistance (MDR) phenotype is characterized by the over-expression of P-glycoprotein (P-gp) on cell plasma membranes that extrudes several drugs out of cells. Cells that express the MDR phenotype are resistant to the mitochondrial related apoptosis and to several anticancer drugs. This study assessed the presence of P-gp in mitochondria and its role in parental drug-sensitive (P5) and in P5-derived MDR1 cells P1(0.5) hepatocellular carcinoma (HCC) cell lines and in drug-sensitive (PSI-2) and mdr1-transfected (PN1A) NIH/3T3 cells. By using Western blot analysis, confocal laser microscopy, measurements of Rhodamine 123 transport across mitochondrial membranes, MDR1 small interfering RNA and flow cytometry analysis, experiments indicate that P-gp is expressed in mitochondria of P1(0.5) and PN1A cells and it is functionally active. Rho 123 accumulation was largely reduced in mitochondria of P1(0.5) cells as compared to those of P5 cells; the reduced uptake of fluorescence in mitochondria of MDR cells was due to P-gp-mediated Rho 123 efflux. In conclusion, these data demonstrate that functionally active P-gp is expressed in the mitochondrial membrane of MDR-positive cells and pumps out anticancer drugs from mitochondria into cytosol. Therefore, P-gp could be involved in the protection of mitochondrial DNA from damage due to antiproliferative drugs.     

The Anopheles gambiae salivary protein gSG6: An anopheline-specific protein with a blood-feeding role

The Anopheles gambiae salivary gland protein 6 (gSG6) is a small protein specifically found in the salivary glands of adult female mosquitoes. We report here the expression of a recombinant form of the protein and we show that in vivo gSG6 is expressed in distal-lateral lobes and is secreted with the saliva while the female mosquito probes for feeding. Injection of gSG6 dsRNA into adult A. gambiae females results in decreased gSG6 protein levels, increased probing time and reduced blood feeding ability. gSG6 orthologs have been found so far only in the salivary glands of Anopheles stephensi and Anopheles funestus, both members of the Cellia subgenus. We report here the gSG6 sequence from five additional anophelines, four species of the A. gambiae complex and Anopheles freeborni, a member of the subgenus Anopheles. We conclude that gSG6 plays some essential blood feeding role and was recruited in the anopheline subfamily most probably after the separation of the lineage which gave origin to Cellia and Anopheles subgenera.     

Structure of the stomach cuticle in adult and larvae of the spider crab Maja brachydactyla (Brachyura,Decapoda)

The stomach of decapods is a complex organ with specialized structures that are delimited by a cuticle. The morphology and ontogeny of the stomach are largely described, but few studies have focused on the morphology of its cuticle. This study examined the morphology of the stomach cuticle of cardiac sacs, gastric mill ossicles, cardio-pyloric valve and pyloric filters, and during various stages (zoea I and II, megalopa, first juvenile, and adult) of the common spider crab Maja brachydactyla using dissection, histology and transmission electron microscopy. The results show that cuticle morphology varies among structures (e.g., cardiac sacs, urocardiac ossicle, cardio-pyloric valve, pyloric filters), within a single structure (e.g., different sides of the urocardiac ossicle) and among different life stages. The cuticle during the larval stages is very thin and the different layers (epicuticle, exocuticle, and endocuticle) are infrequently distinguishable by histology. Major changes during larval development regarding cuticle morphology are observed after the molt to megalopa, including the increment in thickness in the gastric mill ossicles and cardio-pyloric valve, and the disappearance of the long thickened setae of the cardio-pyloric valve. The cuticle of all the stomach structures in the adults is thicker than in larval and juvenile stages. The cuticle varies in thickness, differential staining affinity and morphology of the cuticle layers. The structure–function relationship of the cuticle morphology is discussed.     

Cytotoxicity and enzymatic activity inhibition in cell lines treated with novel iminosugar derivatives

Iminosugars are monosaccharide analogues that have been demonstrated to be specific inhibitors for glycosidases and are currently used therapeutically in several human disorders. N-alkylated derivatives of d-fagomine and (2R,3S,4R,5S)-2-(hydroxymethyl)-5-methylpyrrolidine-3,4-diol with aliphatic chains were tested in eight human cancer cell lines to analyze their cytotoxicity and the inhibitory effect in the activities of specific glycosidases. Results indicate that these compounds were more cytotoxic as the length of the alkyl chain increases. N-dodecyl-d-fagomine inhibited specifically the α-d-glucosidase activity in cell lysates, whereas no effect was detected in other glycosidases. The N-dodecyl derivative of (2R,3S,4R,5S)-2-(Hydroxymethyl)-5-methylpyrrolidine-3,4-diol induced specific inhibition against α-l-fucosidase in cell lysates. Our results indicated that the length of the alkyl chain linked to the iminosugars determine their cytotoxicity as well as the inhibitory effect on the enzymatic activities of specific glycosidases, in human cancer cell lines.     

Molecular-targeted therapy for Duchenne muscular dystrophy: progress and potential

Duchenne muscular dystrophy (DMD) is a lethal heritable childhood myodegenerative condition caused by a mutation within the gene encoding the dystrophin protein within the X chromosome. While, historically, patients with this condition rarely lived into their thirties, they are now living substantially longer as a result of new treatments based on multi-disciplinary care. Despite these advances, the prognosis for DMD patients is limited, and a progressive reduction in quality of life and early death in adulthood cannot be prevented using currently available treatment regimens. The best hopes for a cure lies with cellular and gene therapy approaches that target the underlying genetic defect. In the past several years, viral and nonviral gene therapy methodologies based on adeno-associated viruses, naked plasmid delivery, antisense oligonucleotides, and oligonucleotide-mediated gene editing have advanced to a high degree of sophistication, to the extent that research has moved from the laboratory setting to the clinic. Notwithstanding these accomplishments, shortcomings with each therapy remain, so more work is required to devise an appropriate therapeutic strategy for the management and eventual cure of this debilitating disease.     

Synthesis of new palladium(II) complexes containing tetradentate-nitrogen donor ligands: Combined structural studies by NMR spectroscopy and X-ray crystallography

Treatment of the tetradentate (NN′N′N) N-alkylaminopyrazole ligands 3,6-dimethyl-1,8-(3,5-dimethyl-1-pyrazolyl)-3,6-diazaoctane (ddad) and 1,4-bis[2-(3,5-dimethyl-1-pyrazolyl)ethyl]piperazine (bedp) with [PdCl₂(CH₃CN)₂] in a 1:1 M/L ratio in acetonitrile produces [Pd₂Cl₄(L)] and [PdCl₂(L)] (L = ddad and bedp). Treatment of the corresponding complex [PdCl₂(L)] (L = ddad, bedp) in the presence of AgBF₄ in CH₂Cl₂/methanol (2:1) or NaBF₄ in acetonitrile gives [Pd(L)](BF₄)₂. The Pd(II) complexes have been characterised by elemental analyses, conductivity measurements, IR and ¹H and ¹³C{¹H} NMR spectroscopies when possible. The X-ray structure of the complex [Pd(ddad)]Cl₂ · 3H₂O has been determined. The Pd(II) is coordinated to the ddad ligand by two nitrogen atoms of pyrazolyl groups and two nitrogen atoms of the amine groups, in a slightly distorted square-planar geometry.