Third-generation in situ hybridization chain reaction: multiplexed, quantitative, sensitive, versatile, robust

In situ hybridization based on the mechanism of the hybridization chain reaction (HCR) has addressed multi-decade challenges that impeded imaging of mRNA expression in diverse organisms, offering a unique combination of multiplexing, quantitation, sensitivity, resolution and versatility. Here, with third-generation in situ HCR, we augment these capabilities using probes and amplifiers that combine to provide automatic background suppression throughout the protocol, ensuring that reagents will not generate amplified background even if they bind non-specifically within the sample. Automatic background suppression dramatically enhances performance and robustness, combining the benefits of a higher signal-to-background ratio with the convenience of using unoptimized probe sets for new targets and organisms. In situ HCR v3.0 enables three multiplexed quantitative analysis modes: (1) qHCR imaging – analog mRNA relative quantitation with subcellular resolution in the anatomical context of whole-mount vertebrate embryos; (2) qHCR flow cytometry – analog mRNA relative quantitation for high-throughput expression profiling of mammalian and bacterial cells; and (3) dHCR imaging – digital mRNA absolute quantitation via single-molecule imaging in thick autofluorescent samples

Monoamines, BDNF, Dehydroepiandrosterone, DHEA-Sulfate, and Childhood Depression—An Animal Model Study

Basal levels of monoamines and DHEA in four main limbic brain regions were measured in prepubertal Wistar Kyoto (WKY) rats (a putative animal model of childhood depression). Basal levels of “Brain-Derived Neurotrophic Factor (BDNF)” were also determined in two regions in the hippocampus, compared with Wistar strain controls. In the second phase, we examined the responsiveness of prepubertal WKY rats to different types of chronic antidepressant treatments: Fluoxetine, Desipramine, and dehydroepiandrosterone sulfate (DHEAS). WKY prepubertal rats exhibited different monoamine levels in the limbic system, reduced DHEA levels in the VTA and lower levels of BDNF in the hippocampus CA3 region compared to controls. In prepubertal WKY rats, only treatment with DHEAS produced a statistically significant decrease in immobility, compared to saline-administered controls in the forced swim test. Wistar controls were not affected by any antidepressant. The results imply that DHEA(S) and BDNF may be involved in the pathophysiology and pharmacotherapy of childhood depression

A new family of high nuclearity CoII/DyIII coordination clusters possessing robust and unseen topologies

Mixing Co(NO3)2·6H2O/Dy(NO3)3·6H2O/(E)-4-(2-hydroxy-3-methoxybenzylideneamino)-2,3-dimethyl-1-phenyl-1,2-dihydropyrazol-5-one (HL)/pivalic acid/Et3N in various solvents results in the synthesis of seven compounds formulated as [CoII2DyIII2(μ3-MeO)2(L)2(piv)4(NO3)2] (3), [CoIIDyIII3(μ3-MeO)2(μ2-MeO)2(L)2(piv)2(NO3)3]·2(CH3OH) (4·2CH3OH), 2[CoII4DyIII4(μ2-O)2(μ3-OH)4(L)4(piv)8][CoII2DyIII5(μ3-OH)6(L)2(piv)8(NO3)4] (5), [CoII4DyIII4(μ2-O)2(μ3-OH)4(L)4(piv)8]·2(CH3CN) (6·2CH3CN), [CoII2DyIII5(μ3-OH)6(L)2(piv)8(NO3)4]·4(CH3CN) (7·4CH3CN), [CoII2DyIII2(μ3-OH)2(L)2(piv)2(NO3)2(EtOH)2(H2O)2](NO3)2·(EtOH) (8·EtOH) and [CoII4DyIII4(μ2-O)2(μ3-OH)4(L)4(piv)8] (9) with robust and unseen topologies. These show that the temperature and reaction time influence the formation of the final product. Preliminary magnetic studies, performed for 6 and 7 in the temperature range 2-300 K, are indicative of Single Molecule Magnet (SMM) behaviour. Moreover, analysis of the catalytic properties of compound 3 as an efficient catalyst for the synthesis of trans-4,5-diaminocyclopent-2-enones from 2-furaldehyde and primary amines has been carried out

Neuroactive steroids in depression and anxiety disorders: Clinical studies

Certain neuroactive steroids modulate ligand-gated ion channels via non-genomic mechanisms. Especially 3 alpha-reduced pregnane steroids are potent positive allosteric modulators of the gamma-aminobutyric acid type A (GABA(A)) receptor. During major depression, there is a disequilibrium of 3 alpha-reduced neuroactive steroids, which is corrected by clinically effective pharmacological treatment. To investigate whether these alterations are a general principle of successful antidepressant treatment, we studied the impact of nonpharmacological treatment options on neuroactive steroid concentrations during major depression. Neither partial sleep deprivation, transcranial magnetic stimulation, nor electroconvulsive therapy affected neuroactive steroid levels irrespectively of the response to these treatments. These studies suggest that the changes in neuroactive steroid concentrations observed after antidepressant pharmacotherapy more likely reflect distinct pharmacological properties of antidepressants rather than the clinical response. In patients with panic disorder, changes in neuroactive steroid composition have been observed opposite to those seen in depression. However, during experimentally induced panic induction either with cholecystokinine-tetrapeptide or sodium lactate, there was a pronounced decline in the concentrations of 3 alpha-reduced neuroactive steroids in patients with panic disorder, which might result in a decreased GABAergic tone. In contrast, no changes in neuroactive steroid concentrations could be observed in healthy controls with the exception of 3 alpha,5 alpha-tetrahydrodeoxycorticosterone. The modulation of GABA(A) receptors by neuroactive steroids might contribute to the pathophysiology of depression and anxiety disorders and might offer new targets for the development of novel anxiolytic compounds. Copyright (c) 2006 S. Karger AG, Basel

Third-generation in situ hybridization chain reaction: multiplexed, quantitative, sensitive, versatile, robust

In situ hybridization based on the mechanism of the hybridization chain reaction (HCR) has addressed multi-decade challenges that impeded imaging of mRNA expression in diverse organisms, offering a unique combination of multiplexing, quantitation, sensitivity, resolution and versatility. Here, with third-generation in situ HCR, we augment these capabilities using probes and amplifiers that combine to provide automatic background suppression throughout the protocol, ensuring that reagents will not generate amplified background even if they bind non-specifically within the sample. Automatic background suppression dramatically enhances performance and robustness, combining the benefits of a higher signal-to-background ratio with the convenience of using unoptimized probe sets for new targets and organisms. In situ HCR v3.0 enables three multiplexed quantitative analysis modes: (1) qHCR imaging – analog mRNA relative quantitation with subcellular resolution in the anatomical context of whole-mount vertebrate embryos; (2) qHCR flow cytometry – analog mRNA relative quantitation for high-throughput expression profiling of mammalian and bacterial cells; and (3) dHCR imaging – digital mRNA absolute quantitation via single-molecule imaging in thick autofluorescent samples

Tetranuclear Zn/4f coordination clusters as highly efficient catalysts for Friedel-Crafts alkylation

A series of custom-designed, high yield, isoskeletal tetranuclear Zn/4f coordination clusters showing high efficiency as catalysts with low catalytic loadings in Friedel-Crafts alkylation are described for the first time. The possibility of altering the 4f centers in these catalysts without altering the core topology allows us to further confirm their stability via EPR and NMR, as well to gain insights into the plausible reaction mechanism, showcasing the usefulness of these bimetallic systems as catalysts

Selective complexation of divalent cations by a cyclic α,β-peptoid hexamer: a spectroscopic and computational study

We describe the qualitative and quantitative analysis of the complexation properties towards cations of a cyclic peptoid hexamer composed of alternating α- and β-peptoid monomers, which bear exclusively chiral (S)-phenylethyl side chains (spe) that have no noticeable chelating properties. The binding of a series of monovalent and divalent cations was assessed by 1H NMR, circular dichroism, fluorescence and molecular modelling. In contrast to previous studies on cations binding by 18-membered α-cyclopeptoid hexamers, the 21-membered cyclopeptoid cP1 did not complex monovalent cations (Na+, K+, Ag+) but showed selectivity for divalent cations (Ca2+, Ba2+, Sr2+ and Mg2+). Hexacoordinated C-3 symmetrical complexes were demonstrated for divalent cations with ionic radii around 1 Å (Ca2+ and Ba2+), while 5-coordination is preferred for divalent cations with larger (Ba2+) or smaller ionic radii (Mg2+)

A Markov chain model for changes in users’ assessment of search results

Previous research shows that users tend to change their assessment of search results over time. This is a first study that investigates the factors and reasons for these changes, and describes a stochastic model of user behaviour that may explain these changes. In particular, we hypothesise that most of the changes are local, i.e. between results with similar or close relevance to the query, and thus belong to the same ”coarse” relevance category. According to the theory of coarse beliefs and categorical thinking, humans tend to divide the range of values under consideration into coarse categories, and are thus able to distinguish only between cross-category values but not within them. To test this hypothesis we conducted five experiments with about 120 subjects divided into 3 groups. Each student in every group was asked to rank and assign relevance scores to the same set of search results over two or three rounds, with a period of three to nine weeks between each round. The subjects of the last three-round experiment were then exposed to the differences in their judgements and were asked to explain them. We make use of a Markov chain model to measure change in users’ judgments between the different rounds. The Markov chain demonstrates that the changes converge, and that a majority of the changes are local to a neighbouring relevance category. We found that most of the subjects were satisfied with their changes, and did not perceive them as mistakes but rather as a legitimate phenomenon, since they believe that time has influenced their relevance assessment. Both our quantitative analysis and user comments support the hypothesis of the existence of coarse relevance categories resulting from categorical thinking in the context of user evaluation of search results