Fluorescence tomographic imaging in turbid media using early-arriving photons and Laplace transforms

We present a multichannel tomographic technique to detect fluorescent objects embedded in thick (6.4 cm) tissue-like turbid media using early-arriving photons. The experiments use picosecond laser pulses and a streak camera with single photon counting capability to provide short time resolution and high signal-to-noise ratio. The tomographic algorithm is based on the Laplace transform of an analytical diffusion approximation of the photon migration process and provides excellent agreement between the actual positions of the fluorescent objects and the experimental estimates. Submillimeter localization accuracy and 4- to 5-mm resolution are demonstrated. Moreover, objects can be accurately localized when fluorescence background is present. The results show the feasibility of using early-arriving photons to image fluorescent objects embedded in a turbid medium and its potential in clinical applications such as breast tumor detection.

How photons start vision.

Recent studies have elucidated how the absorption of a photon in a rod or cone cell leads to the generation of the amplified neural signal that is transmitted to higher-order visual neurons. Photoexcited visual pigment activates the GTP-binding protein transducin, which in turn stimulates cGMP phosphodiesterase. This enzyme hydrolyzes cGMP, allowing cGMP-gated cationic channels in the surface membrane to close, hyperpolarize the cell, and modulate transmitter release at the synaptic terminal. The kinetics of reactions in the cGMP cascade limit the temporal resolution of the visual system as a whole, while statistical fluctuations in the reactions limit the reliability of detection of dim light. Much interest now focuses on the processes that terminate the light response and dynamically regulate amplification in the cascade, causing the single photon response to be reproducible and allowing the cell to adapt in background light. A light-induced fall in the internal free Ca2+ concentration coordinates negative feedback control of amplification. The fall in Ca2+ stimulates resynthesis of cGMP, antagonizes rhodopsin's catalytic activity, and increases the affinity of the light-regulated cationic channel for cGMP. We are using physiological methods to study the molecular mechanisms that terminate the flash response and mediate adaptation. One approach is to observe transduction in truncated, dialyzed photoreceptor cells whose internal Ca2+ and nucleotide concentrations are under experimental control and to which exogenous proteins can be added. Another approach is to observe transduction in transgenic mouse rods in which specific proteins within the cascade are altered or deleted.

Photoassembly of the manganese cluster and oxygen evolution from monomeric and dimeric CP47 reaction center photosystem II complexes

Isolated subcomplexes of photosystem II from spinach (CP47RC), composed of D1, D2, cytochrome b559, CP47, and a number of hydrophobic small subunits but devoid of CP43 and the extrinsic proteins of the oxygen-evolving complex, were shown to reconstitute the Mn4Ca1Clx cluster of the water-splitting system and to evolve oxygen. The photoactivation process in CP47RC dimers proceeds by the same two-step mechanism as observed in PSII membranes and exhibits the same stoichiometry for Mn2+, but with a 10-fold lower affinity for Ca2+ and an increased susceptibility to photodamage. After the lower Ca2+ affinity and the 10-fold smaller absorption cross-section for photons in CP47 dimers is taken into account, the intrinsic rate constant for the rate-limiting calcium-dependent dark step is indistinguishable for the two systems. The monomeric form of CP47RC also showed capacity to photoactivate and catalyze water oxidation, but with lower activity than the dimeric form and increased susceptibility to photodamage. After optimization of the various parameters affecting the photoactivation process in dimeric CP47RC subcores, 18% of the complexes were functionally reconstituted and the quantum efficiency for oxygen production by reactivated centers approached 96% of that observed for reconstituted photosystem II-enriched membranes.

Logic gates using high Rydberg states

Connected logic gates can be operated on the levels of one molecule by making use of the special properties of high Rydberg states. Explicit experimental results for the NO molecule are provided as an example. A number of other options, including that of several gates concatenated so as to operate as a full adder, are discussed. Specific properties of high Rydberg states that are used are: their autoionization is delayed so that they can be distinguished from direct multiphoton ionization, during their long life such states also can decay by energy transfer to the molecular core in a way that can be controlled by the judicious application of very weak external electrical fields, and the Rydberg states can be detected by the application of an ionizing electrical field. The combination of two (or three) color photons with and without external weak fields allows the construction of quite elaborate logic circuit diagrams and shows that taking advantage of the different intramolecular dynamics of levels that differ by their excitation enables the compounding of logic operations on one molecular frame.

Direct searches for dark matter: Recent results

There is abundant evidence for large amounts of unseen matter in the universe. This dark matter, by its very nature, couples feebly to ordinary matter and is correspondingly difficult to detect. Nonetheless, several experiments are now underway with the sensitivity required to detect directly galactic halo dark matter through their interactions with matter and radiation. These experiments divide into two broad classes: searches for weakly interacting massive particles (WIMPs) and searches for axions. There exists a very strong theoretical bias for supposing that supersymmetry (SUSY) is a correct description of nature. WIMPs are predicted by this SUSY theory and have the required properties to be dark matter. These WIMPs are detected from the byproducts of their occasional recoil against nucleons. There are efforts around the world to detect these rare recoils. The WIMP part of this overview focuses on the cryogenic dark matter search (CDMS) underway in California. Axions, another favored dark matter candidate, are predicted to arise from a minimal extension of the standard model that explains the absence of the expected large CP violating effects in strong interactions. Axions can, in the presence of a large magnetic field, turn into microwave photons. It is the slight excess of photons above noise that signals the axion. Axion searches are underway in California and Japan. The axion part of this overview focuses on the California effort. Brevity does not allow me to discuss other WIMP and axion searches, likewise for accelerator and satellite based searches; I apologize for their omission.

Increase in the Quantum Yield of Photoinhibition Contributes to Copper Toxicity in Vivo1

The effect of copper on photoinhibition of photosystem II in vivo was studied in bean (Phaseolus vulgaris L. cv Dufrix). The plants were grown hydroponically in the presence of various concentrations of Cu2+ ranging from the optimum 0.3 μm (control) to 15 μm. The copper concentration of leaves varied according to the nutrient medium from a control value of 13 mg kg−1 dry weight to 76 mg kg−1 dry weight. Leaf samples were illuminated in the presence and absence of lincomycin at different light intensities (500–1500 μmol photons m−2 s−1). Lincomycin prevents the concurrent repair of photoinhibitory damage by blocking chloroplast protein synthesis. The photoinhibitory decrease in the light-saturated rate of O2 evolution measured from thylakoids isolated from treated leaves correlated well with the decrease in the ratio of variable to maximum fluorescence measured from the leaf discs; therefore, the fluorescence ratio was used as a routine measurement of photoinhibition in vivo. Excess copper was found to affect the equilibrium between photoinhibition and repair, resulting in a decrease in the steady-state concentration of active photosystem II centers of illuminated leaves. This shift in equilibrium apparently resulted from an increase in the quantum yield of photoinhibition (ΦPI) induced by excess copper. The kinetic pattern of photoinhibition and the independence of ΦPI on photon flux density were not affected by excess copper. An increase in ΦPI may contribute substantially to Cu2+ toxicity in certain plant species.

Identification of a Novel Isoform of the Chloroplast-Coupling Factor α-Subunit 1

Studies were conducted to identify a 64-kD thylakoid membrane protein of unknown function. The protein was extracted from chloroplast thylakoids under low ionic strength conditions and purified to homogeneity by preparative sodium dodecyl sulfate-polyacrylamide gel electrophoresis. Four peptides generated from the proteolytic cleavage of the wheat 64-kD protein were sequenced and found to be identical to internal sequences of the chloroplast-coupling factor (CF1) α-subunit. Antibodies for the 64-kD protein also recognized the α-subunit of CF1. Both the 64-kD protein and the 61-kD CF1 α-subunit were present in the monocots barley (Hordeum vulgare), maize (Zea mays), oat (Avena sativa), and wheat (Triticum aestivum); but the dicots pea (Pisum sativum), soybean (Glycine max Merr.), and spinach (Spinacia oleracea) contained only a single polypeptide corresponding to the CF1 α-subunit. The 64-kD protein accumulated in response to high irradiance (1000 μmol photons m−2 s−1) and declined in response to low irradiance (80 μmol photons m−2 s−1) treatments. Thus, the 64-kD protein was identified as an irradiance-dependent isoform of the CF1 α-subunit found only in monocots. Analysis of purified CF1 complexes showed that the 64-kD protein represented up to 15% of the total CF1 α-subunit.

The rate constant of photoinhibition, measured in lincomycin-treated leaves, is directly proportional to light intensity.

Pumpkin leaves grown under high light (500-700 micromol of photons m-2.s-1) were illuminated under photon flux densities ranging from 6.5 to 1500 micromol.m-2.s-1 in the presence of lincomycin, an inhibitor of chloroplast protein synthesis. The illumination at all light intensities caused photoinhibition, measured as a decrease in the ratio of variable to maximum fluorescence. Loss of photosystem II (PSII) electron transfer activity correlated with the decrease in the fluorescence ratio. The rate constant of photoinhibition, determined from first-order fits, was directly proportional to photon flux density at all light intensities studied. The fluorescence ratio did not decrease if the leaves were illuminated in low light in the absence of lincomycin or incubated in darkness in the presence of lincomycin. The constancy of the quantum yield of photoinhibition under different photon flux densities strongly suggests that photoinhibition in vivo occurs by one dominant mechanism under all light intensities. This mechanism probably is not the acceptor side mechanism characterized in the anaerobic case in vitro. Furthermore, there was an excellent correlation between the loss of PSII activity and the loss of the D1 protein from thylakoid membranes under low light. At low light, photoinhibition occurs so slowly that inactive PSII centers with the D1 protein waiting to be degraded do not accumulate. The kinetic agreement between D1 protein degradation and the inactivation of PSII indicates that the turnover of the D1 protein depends on photoinhibition under both low and high light.