BackgroundIn humans, ethanol exposure during pregnancy causes a spectrum of developmental defects (fetal alcohol syndrome or FAS). Individuals vary in phenotypic expression. Zebrafish embryos develop FAS-like features after ethanol exposure. In this study, we ask whether stage-specific effects of ethanol can be identified in the zebrafish, and if so, whether they allow the pinpointing of sensitive developmental mechanisms. We have therefore conducted the first large-scale (>1500 embryos) analysis of acute, stage-specific drug effects on zebrafish development, with a large panel of readouts.Methodology/Principal FindingsZebrafish embryos were raised in 96-well plates. Range-finding indicated that 10% ethanol for 1 h was suitable for an acute exposure regime. High-resolution magic-angle spinning proton magnetic resonance spectroscopy showed that this produced a transient pulse of 0.86% concentration of ethanol in the embryo within the chorion. Survivors at 5 days postfertilisation were analysed. Phenotypes ranged from normal (resilient) to severely malformed. Ethanol exposure at early stages caused high mortality (≥88%). At later stages of exposure, mortality declined and malformations developed. Pharyngeal arch hypoplasia and behavioral impairment were most common after prim-6 and prim-16 exposure. By contrast, microphthalmia and growth retardation were stage-independent.ConclusionsOur findings show that some ethanol effects are strongly stage-dependent. The phenotypes mimic key aspects of FAS including craniofacial abnormality, microphthalmia, growth retardation and behavioral impairment. We also identify a critical time window (prim-6 and prim-16) for ethanol sensitivity. Finally, our identification of a wide phenotypic spectrum is reminiscent of human FAS, and may provide a useful model for studying disease resilience.

Photochemically induced dynamic nuclear polarisation (photoCIDNP) in intact bacterial reaction centres has been observed by 13Csolid state NMR under continuous illumination with white light. Strong intensity enhancement of 13C NMR signals of the aromatic rings allows probing the electronic ground state of the two BChl cofactors of the special pair at the molecular scale with atomic selectivity. Differences between the two BChl cofactors are discussed. Several aliphatic 13C atoms of cofactors, as well as 13C atoms of the imidazole ring of histidine residue(s), show nuclearspin polarisation to the same extent as the aromatic nuclei of the cofactors. Mechanisms and applications of polarisation transfer are discussed.

ABSTRACT. A sample of solubilized and quinone-depleted reaction centers (RC) from the purple bacterium Rhodobacter (R.) sphaeroides wild-type (WT) has been prepared entirely 13 C and 15 N isotope labelled at all positions of the protein as well as of the cofactors. In this sample, the occurrence of the solid-state photo-CIDNP (photochemically induced dynamic nuclear polarization) effect has been probed by 13 C solid-state magic-angle spinning (

Abstract. The electron donor in photosystem I, the chlorophyll dimer P700, is studied by photo-CIDNP (photochemically induced dynamic nuclear polarization) MAS (magic-angle spinning) NMR on selectively 13C and uniformly 15N labeled PSI core preparations (PSI-100) obtained from the aquatic plant duckweed (Spirodela oligorrhiza). Light-induced signals originate from the isotope labelled nuclei of the cofactors involved in the spin-correlated radical pair forming upon light excitation. Signals are assigned to the two donor cofactors (Chl a and Chl a') and the two acceptor cofactors (both Chl a). Light induced signals originating from both donor and acceptor cofactors demonstrate that electron transfer occurs through both branches of cofactors in the pseudo-C2 symmetric Reaction Center (RC). The experimental results supported by quantum chemical calculations indicate that this functional symmetry occurs in PSI despite similarly sized chemical-shift differences between cofactors of PSI and the functionally asymmetric special-pair donor of the bacterial RC of Rhodobacter sphaeroides. This contributes to converging evidence that local differences in time-averaged electronic ground-state properties, over the donor are of little importance for functional symmetry breaking across photosynthetic RC species.