Co-localization of ecdysteroid receptors and c-fos-like protein in the brain of Manduca sexta larvae

Summary The presence of c-fos, a marker for cell activation, was investigated in cerebral neurons actively expressing ecdysteroid receptors during larval-pupal development in the tobacco hornworm, Manduca sexta. Colocalization was accomplished by ecdysteroid autoradiography using the tritiated high affinity 20-hydroxyecdysone agonist ponasterone A and immunocytochemistry with an antibody to a peptide sequence which is highly conserved in both human and murine c-fos. Immunoreactivity to a c-fos-like protein(s) was present in nuclei of many neurons of all the developmental stages examined. However, with the exception of the optic lobe, cells expressing nuclear ecdysteroid receptors were more immunoreactive than non-ecdysteroid-binding neurons. These data suggest that ecdysteroid-induced gene activation and translation may involve c-fos expression. Offprint requests to: H.-J. Bidmon     

Quantitative structure-activity relationships (QSAR) in sweet aspartyl dipeptide methyl esters

In drug design the pharmacochemists frequently use physicochemicalconstants to correlate the structure with the observed potency.Curiously this approach has hardly been followed by psychophysiciststo indicate the increase of taste potency in a series of structurallyrelated compounds of the same stimulus. In the present experiments we correlated the relative sweetness(S) of 40 aspartyl dipeptide methyl esters [general formulaCH₂(COO^–). CH(NH₃⁺ ).CO.NH.CH(R).COOCH₃] with 8 physicochemicalparameters. Among the compounds we had 7 non-sweet stimuli whilethe potency of the remaining 33 peptide esters varied from 1to 27,000 (1 = sucrose). We calculated for the side chain Rthe values of the parachor parameter P, the hydrophobic fragmentalconstant f and 5 STERIMOL parameters (L, B₁ up to B₅). A multipleregression analysis programme selected by stages the most relevantparameters and tested their significance. We observed that the criterion whether a dipeptide ester issweet or not, is among others defined by the L and B₅ parametersof the side chain R. Compounds are sweet provided L is confinedto certain limits (0.50 nm<L<0.62 nm), or otherwise whenL exceeds these limits, the B₅ parameter has to be greater than0.45 nm (when L<0.50 nm) or smaller than 0.72 nm (when L>0.62nm). The sweet potency defined as log S correlated very significantlywith the parameters P and B₄ (n=33, r=0.812, s=0.60, F=29.06).When two compounds, which were shown to be situated at the borderlineof the length and volume parameters, were omitted in the analysis,the correlation improved (n=31, r=0.909, s=0.40, F=42.60). Inthe latter situation we found the following equation when theintercept was set at zero: log S=0.194f + 1.472.10^–2P—3.357B₅ A previously proposed conformation of aspartame (R=CH₂-Øat the receptor site was computed in detail. We calculated thedistances of the AH-B moieties to the third binding site (thecentre of Ø) and indicated the width of the receptoraccess for this series of sweet, structurally related, dipeptidemethyl esters.     

The morphology of the Limulus visual system

An ocellus of the horseshoe crab, Limulus polyphemus, has been serially sectioned for light and electron microscopy, its sensory cells have been indexed, and the interconnections of a third of these traced. The ocellus contains 155 retinula cells and 26 arhabdomeric cells, which are secondary sensory neurons. Of these, 55 retinula cells constitute 7 quasi-ommatidial assemblages, each innervated by at least one and a total of 9 arhabdomeric cells. When known electrotonic coupling patterns are compared with gap-junctional connections, retinula cells sensitive to visible or ultraviolet light can be tentatively identified. Retinula cell axons contribute collaterals to a synaptic plexus, in which the arhabdomeric cells apparently do not participate.     

P2 pyridine N-oxide thrombin inhibitors: a novel peptidomimetic scaffold

In this study, we have demonstrated that the critical hydrogen bonding motif of the established 3-aminopyrazinone thrombin inhibitors can be effectively mimicked by a 2-aminopyridine N-oxide. As this peptidomimetic core is more resistant toward oxidative metabolism, it also overcomes the metabolic liability associated with the pyrazinones. An optimization study of the P(1) benzylamide delivered the potent thrombin inhibitor 21 (K(i) = 3.2 nM, 2xaPTT = 360 nM), which exhibited good plasma levels and half-life after oral dosing in the dog (C(max) = 2.6 microM, t(1/2) = 4.5 h).     

Diversity surveys and evolutionary relationships of aoxB genes in aerobic arsenite-oxidizing bacteria

A new primer set was designed to specifically amplify ca. 1,100 bp of aoxB genes encoding the As(III) oxidase catalytic subunit from taxonomically diverse aerobic As(III)-oxidizing bacteria. Comparative analysis of AoxB protein sequences showed variable conservation levels and highlighted the conservation of essential amino acids and structural motifs. AoxB phylogeny of pure strains showed well-discriminated taxonomic groups and was similar to 16S rRNA phylogeny. Alphaproteobacteria-, Betaproteobacteria-, and Gammaproteobacteria-related sequences were retrieved from environmental surveys, demonstrating their prevalence in mesophilic As-contaminated soils. Our study underlines the usefulness of the aoxB gene as a functional marker of aerobic As(III) oxidizers.     

Immunological characteristics associated with the protective efficacy of antibodies to ricin

A/B toxins, produced by bacteria and plants, are among the deadliest molecules known. The B chain binds the cell, whereas the A chain exerts the toxic effect. Both anti-A chain and anti-B chain Abs can neutralize toxins in vivo and in vitro. B chain Abs block binding of the toxin to the cell. It is not known how anti-A chain Abs function. Working with ricin toxin, we demonstrate that immunization with A chain induces greater protection than immunization with B chain. A panel of mAbs, binding to A chain, B chain, or both chains, has been produced and characterized. Immunologic characteristics evaluated include isotype, relative avidity, and epitope specificity. The ability to inhibit ricin enzymatic or cell binding activity was studied, as was the ability to block ricin-mediated cellular cytotoxicity on human and murine cell lines. Finally, the in vivo protective efficacy of the Abs in mice was studied. The Ab providing the greatest in vivo protective efficacy was directed against the A chain. It had the greatest relative avidity and the greatest ability to block enzymatic function and neutralize cytotoxicity. Interestingly, we also obtained an anti-A chain Ab that bound with high avidity, blocked enzymatic activity, did not neutralize cytotoxicity, and actually enhanced the in vivo toxicity of ricin. Anti-A chain Abs with moderate avidity had no in vivo effect, nor did any anti-B chain Abs.