Bolometer operating at the threshold for circuit quantum electrodynamics.

Radiation sensors based on the heating effect of absorbed radiation are typically simple to operate and flexible in terms of input frequency, so they are widely used in gas detection1, security2, terahertz imaging3, astrophysical observations4 and medical applications5. Several important applications are currently emerging from quantum technology and especially from electrical circuits that behave quantum mechanically, that is, circuit quantum electrodynamics6. This field has given rise to single-photon microwave detectors7-9 and a quantum computer that is superior to classical supercomputers for certain tasks10. Thermal sensors hold potential for enhancing such devices because they do not add quantum noise and they are smaller, simpler and consume about six orders of magnitude less power than the frequently used travelling-wave parametric amplifiers11. However, despite great progress in the speed12 and noise levels13 of thermal sensors, no bolometer has previously met the threshold for circuit quantum electrodynamics, which lies at a time constant of a few hundred nanoseconds and a simultaneous energy resolution of the order of 10h gigahertz (where h is the Planck constant). Here we experimentally demonstrate a bolometer that operates at this threshold, with a noise-equivalent power of 30 zeptowatts per square-root hertz, comparable to the lowest value reported so far13, at a thermal time constant two orders of magnitude shorter, at 500 nanoseconds. Both of these values are measured directly on the same device, giving an accurate estimation of 30h gigahertz for the calorimetric energy resolution. These improvements stem from the use of a graphene monolayer with extremely low specific heat14 as the active material. The minimum observed time constant of 200 nanoseconds is well below the dephasing times of roughly 100 microseconds reported for superconducting qubits15 and matches the timescales of currently used readout schemes16,17, thus enabling circuit quantum electrodynamics applications for bolometers.

Cryogenic sensor enabling broad-band and traceable power measurements.

Recently, great progress has been made in the field of ultrasensitive microwave detectors, reaching even the threshold for utilization in circuit quantum electrodynamics. However, cryogenic sensors lack the compatibility with broad-band metrologically traceable power absorption measurements at ultralow powers, which restricts their range of applications. Here, we demonstrate such measurements using an ultralow-noise nanobolometer, which we extend by an additional direct-current (dc) heater input. The tracing of the absorbed power relies on comparing the response of the bolometer between radio frequency and dc-heating powers traced to the Josephson voltage and quantum Hall resistance. To illustrate this technique, we demonstrate two different methods of dc-substitution to calibrate the power that is delivered to the base temperature stage of a dilution refrigerator using our in situ power sensor. As an example, we demonstrate the ability to accurately measure the attenuation of a coaxial input line between the frequencies of 50 MHz and 7 GHz with an uncertainty down to 0.1 dB at a typical input power of -114 dBm.

[Isoenzymes of carbonic anhydrase of an euryhaline fish. Variations related to the osmoregulation]. / Isoenzymes de l'anhydrase carbonique d'un poisson euryhalin. Variations en relation avec l'osmoregulation

A chemical study of carbonic anhydrase (EC 4.2.1.1) from the red blood cells and the gills of an euryhaline fish (Anguilla anguilla) is presented. Animals adapted to fresh water were compared to those adapted to sea water. The physiochemical constants of the various molecular formsisolated by chromatography and isoelectric focusing were determined; isoelectric pH, molecular weight, and the Km and V/E of the enzyme dehydration activity were compared. In both red cells and gills of fish adapted in either media various forms were isolated, characterized by different enzymatic kinetics (high- and low- activity forms) but having the same molecular weight (27 250). Some isoenzymes isolated from the gills differed significantly from those isolated from the red cells. Adaptation to fresh water or sea water is accompanied by modifications in the distribution of the isoenzymes in both red cells and gills: adaptation to sea water is characterized by a shift of the molecular forms towards an isoelectric pH higher than pH equals 6. The role of these enzymes is discussed under both a physiological and biochemical point of view in relation to the electrolyte exchange across fish gill. The origin of the different molecular forms of the carbonic anhydrase is discussed in relation to the prevailing theories on this subject.

Characterization of valine transport in sea urchin eggs.

In unfertilized eggs, the mechanism of valine uptake can be summarized as follows. It is saturable over the external concentration of valine and insensitive to the presence of external sodium, depletion of cellular energy supplies and intracellular acidosis. The activation energy for the transport reaction (16.3 kcal/mol) is within the range of values reported for active transport of small molecules. In fertilized eggs, the total rate of valine uptake can be divided into two components: (i) a Na+-insensitive uptake which accounts for about 7% of total absorption as shown by studies in Na+-free medium seems to possess the same characteristics as in unfertilized eggs, (ii) a Na+-dependent transport of valine which constitutes the main entry is formed about 5 min after fertilization. It follows Michaelis-Menten kinetics characterized by 15-fold increase in Vmax with no change in Km. These two mechanisms have characteristics in common, such as their insensitivity to metabolic energy supply, their energy of activation and their ability to concentrate valine. The relationship between the establishment of the Na+-dependent valine uptake and the ionic events triggered by fertilization is discussed.

Calcium in sea urchin egg during fertilization.

Calcium plays a strikingly important role in two of the major events in developmental biology: cell activation and differentiation. In this review we begin with the location and quantity of intracellular calcium in sea urchin oocytes, and then discuss the changes that occur during fertilization and egg activation, placing special emphasis on the mobilization and redistribution of intracellular calcium. We also discuss the propagation of the calcium wave and the role of the burst of calcium on the process of reorganizing the egg cortex at fertilization.

Calcium pools in sea urchin eggs: roles of endoplasmic reticulum and mitochondria in relation to fertilization.

A preparation of sea urchin eggs permeabilized with digitonin (40 microM for 2.5 min) was used to study the kinetic characteristics of the two cellular compartments suspected to play a key role in cellular calcium transfer during fertilization: an ATP-dependent Ca2+ pool (Km = 0.47 microM; Vm = 0.48 nmol/min.mg protein) probably located in the endoplasmic reticulum and a mitochondrial Ca2+ pool (Km = 1.50 microM; Vm = 0.12 nmol/min.mg protein). Fertilization triggered a decrease in the rate of ATP dependent uptake by the non-mitochondrial pool (Km = 0.59 microM; Vm = 0.15 nmol/min.mg protein) while it transiently increased the Ca2+ uptake into mitochondria (2 min post-fertilization: Km = 2.20 microM; Vm = 0.40 nmol/min.mg protein). Microanalysis studies performed on quickly frozen, freeze substituted and embedded eggs showed a transient Ca2+ enrichment of mitochondria soon after fertilization thus suggesting that mitochondria behave as a Ca2+ sink at fertilization. Results are discussed in relation to the role of endoplasmic reticulum and mitochondria in handling free calcium during the early period following sea urchin egg fertilization.

Caulerpenyne interferes with microtubule-dependent events during the first mitotic cycle of sea urchin eggs.

Caulerpenyne (Cyn), the major secondary metabolite synthesized by the green alga Caulerpa taxifolia proliferating in the Mediterranean Sea, is a cytotoxic sesquiterpene. As this compound has an antiproliferative potency by inhibiting division of many types of cells, we examined the precise effects of Cyn during the early development of the sea urchin Paracentrotus lividus. Whereas Cyn (60 microM) had no effect on fertilization, it blocked the first cell division in the same manner whether added before or after fertilization, provided the drug was added before or during metaphase. Immunofluorescence localization revealed that Cyn had no effect on the microtubular sperm aster formation, pronuclei migration and fusion, chromosome condensation, nuclear envelope breakdown, and bipolar mitotic spindle assembly. However, mitosis was blocked in a metaphase-like stage at which most chromosomes were aligned at the equatorial plate, while a few of them had not even migrated towards the metaphase plate. When added after the metaphase-anaphase transition, the first division occurred normally but the second division was inhibited with the same phenotype as described above. We previously showed that Cyn did not affect protein synthesis or H1 kinase activation or deactivation (Pesando et al., 1996, Aquat. Toxicol. 35, 139), but that it partially inhibited DNA synthesis. Our results establish that Cyn does not affect the microfilament-dependent processes of fertilization and cytokinesis and allows the beginning of mitosis, but prevents normal DNA replication and results in metaphase-like arrest of sea urchin embryos.

Caulerpenyne blocks MBP kinase activation controlling mitosis in sea urchin eggs.

In a previous study, we demonstrated that caulerpenyne (Cyn), a natural sesquiterpene having an antiproliferative potency, blocked the mitotic cycle of sea urchin embryos at metaphase and inhibited the phosphorylation of several proteins, but did not affect histone H1 kinase activation (Pesando et al, 1998, Eur. J. Cell Biol. 77, 19-26). Here, we show that concentrations of Cyn that blocked the first division of the sea urchin Paracentrotus lividus embryos in a metaphase-like stage (45 microM) also inhibited the stimulation of mitogen-activated protein kinase (MAPK) activity in vivo as measured in treated egg extracts using myelin basic protein (MBP) as a substrate (MBPK). However, Cyn had no effect on MBP phosphorylation when added in vitro to an untreated egg extract taken at the time of metaphase, suggesting that Cyn acts on an upstream activation process. PD 98059 (40 microM), a previously characterized specific synthetic inhibitor of MAPK/extracellular signal-regulated kinase-1 (MEK1), also blocked sea urchin eggs at metaphase in a way very similar to Cyn. Both molecules induced similar inhibitory effects on MBP kinase activation in vivo, but had no direct effect on MBP kinase activity in vitro, whereas they did not affect H1 kinase activation neither in vivo nor in vitro. As a comparison, butyrolactone 1 (100 microM), a known inhibitor of H1 kinase activity, did inhibit H1 kinase of sea urchin eggs in vivo and in vitro, and blocked the sea urchin embryo mitotic cycle much before metaphase. Immunoblots of mitotic extracts, treated with anti-active MAP-kinase antibody, showed that both Cyn and PD 98059 reduced the phosphorylation of p42 MAP kinase (Erk2) in vivo. Our overall results suggest that Cyn blocks the sea urchin embryo mitotic cycle at metaphase by inhibiting an upstream phosphorylation event in the MBPK activation pathway. They also show that H1 kinase and MBPK activation can be dissociated from each other in this model system.

Human seminal plasma contains a protein that shares physicochemical, immunochemical, and immunosuppressive properties with pregnancy-associated plasma protein-A.

Pregnancy-associated plasma protein-A (PAPP-A) was detected by RIA in human seminal plasma. This was not due to interference with proteases or binding to seminal plasma proteins, since immunoreactivity was not affected by treatment with protease inhibitors, and the elution of [125I]PAPP-A was not altered by preincubation with seminal plasma. The major component of the seminal plasma PAPP-A coeluted with pure PAPP-A or plasma PAPP-A from pregnant and nonpregnant women. In the RIA, serial dilutions of seminal plasma gave parallel displacement curves to pregnancy plasma. Removing PAPP-A-like material from seminal plasma by adsorbtion to heparin-Sepharose reduced the inhibitory effect of seminal plasma on phytohemagglutinin-induced lymphocyte transformation. The source of seminal plasma PAPP-A-like immunoreactive material remains to be elucidated, but it is unlikely to be the testis, since PAPP-A levels in semen of vasectomized men were similar to those in nonvasectomized men.

Saturation transfer EPR measurements of the rotational motion of a strongly immobilized ouabain spin label on renal Na,K-ATPase.

The rotational motion of an ouabain spin label with sheep kidney Na,K-ATPase has been measured by electron paramagnetic resonance (EPR) and saturation transfer EPR (ST-EPR) measurements. Spin-labelled ouabain binds with high affinity to the Na,K-ATPase with concurrent inhibition of ATPase activity. Enzyme preparations retain 0.61 +/- 0.1 mol of bound ouabain spin label per ATPase alpha beta dimer. The conventional EPR spectrum of the ouabain spin label bound to the ATPase consists almost entirely (greater than 99%) of a broad resonance which is characteristic of a strongly immobilized spin label. ST-EPR measurements of the spin labelled ATPase preparations yield effective correlation times for the bound labels of 209 +/- 11 microseconds at 0 degree C and 44 +/- 4 microseconds at 20 degrees C. These rotational correlation times most likely represent the motion of the protein itself rather than the independent motion of mobile spin probes relative to a slower moving protein. Additional ST-EPR measurements with glutaraldehyde-crosslinked preparations indicated that the observed rotational correlation times predominantly represented the motion of entire Na,K-ATPase-containing membrane fragments, rather than the motion of individual monomeric or dimeric polypeptides within the membrane fragment. The strong immobilization of the ouabain spin label will make it an effective paramagnetic probe of the extracellular surface of the Na,K-ATPase for a variety of NMR and EPR investigations.

Evidence for an acidic compartment in sea urchin eggs (Paracentrotus lividus): role at fertilization.

The characteristics of [14C]methylamine accumulation by isolated cortices were measured in eggs from three species of sea urchins: Paracentrotus lividus, Arbacia lixula and Sphaerechinus granularis. In all cases, the results pointed to the existence of an acidic compartment in the cortical zone. In P. lividus eggs, cortical granules did not participate in proton storage which likely took place in pigment granules. [14C]Methylamine accumulation was dramatically reduced by monovalent cation ionophores (monensin and nigericin) and by NH4Cl, but not by valinomycin. ATP depletion only partially affected the isotope uptake. Simultaneous measurements of intracellular pH and of external titratable acidity during ammonia treatment of eggs, indicate that after fertilization, eggs increased their capacity to concentrate hydrogen ions in an intracellular store. Following insemination, cortices from P. lividus eggs exhibited a 3-fold increase in [14C]methylamine accumulation. It is concluded that the egg cortical area contains acidic organelles sequestering hydrogen ions by means of an electrogenic H+ pump, and that this mechanism, enhanced at fertilization, participates in a local alkalinization. The role of such a mechanism is discussed.

Synthesis and structure-activity relationships of new 1, 3-disubstituted cyclohexanes as structurally rigid leukotriene B(4) receptor antagonists.

A series of 1-hydroxy-3-¿3-hydroxy-7-phenyl-1-hepten-1-yl cyclohexane acetic acid derivatives was designed based on postulated active conformation of leukotriene B(4) (LTB(4)) and evaluated as human cell surface LTB(4) receptor (BLTR) antagonists. Binding was determined through ¿(3)HLTB(4) displacement from human neutrophils and receptor antagonistic assays by in vitro measurements of inhibition of leukocyte chemotaxis induced by LTB(4). On the basis of these assays, a structure-affinity relationship was investigated. Optimization of the acid chain length and omega-substitution of a phenyl group on the lipophilic tail were shown to be critical for binding activity. These modifications led to the discovery of compounds with submicromolar potency and selective BLTR antagonism. The most potent compound 3balpha (IC(50) = 250 nM) was found to significantly inhibit oedema formation in a topical model of phorbolester-induced inflammation. Substantial improvement of in vitro potency was achieved by modification of the carboxylic acid function leading to the identification of the N,N-dimethylamide series. Compound 5balpha, free of agonist activity, displayed higher potency in receptor binding with an IC(50) of 40 nM. These results support the hypothesis that the spatial relationship between the carboxylic acid and allylic hydroxyl functions is crucial for high binding affinity with BLTR.

In vitro production of IgE in humans: kinetic studies and analysis of serum enhancing and inhibitory factors.

By using a sensitive and reliable method for the measurement of IgE produced in vitro by peripheral blood lymphomonocytes, the following observations may be made. Serum obtained from atopic donors shows a significant capacity to enhance the in vitro synthesis of IgE of PBL from atopic as well as of non-atopic donors. On the other hand, serum obtained from non-atopic donors demonstrates a clear-cut inhibitory effect on IgE synthesis of cells from both atopic and non-atopic individuals. The kinetics of the IgE production was studied. As early as 30 min after initiation of culture, PBL of atopic individuals showed the capacity to synthesize large amounts of IgE. The IgE synthesis continued, with variations from case to case, for 7 days. This capacity of the cells is completely inhibited following incubation of the PBL for a short period of time with a protein synthesis inhibitor, cycloheximide. This in vitro procedure represents a valuable tool for the study and identification of the enhancing and suppressor factors of IgE synthesis present in the serum.

Ryanodine Activates Sea Urchin Eggs: (sea urchin egg/activation/ryanodine/inositol phosphate/heparin).

We have studied the effect on sea urchin eggs of ryanodine, a plant alkaloid that causes muscle contraction by opening calcium channels in the sarcoplasmic reticulum terminal cisternae. Ryanodine, although it is less effective that IP3 , produces full or partial activation in 62% of injected sea urchin eggs. In addition ryanodine inhibits in a dose dependant manner 45 Ca pumping in the isolated egg cortex or in eggs permeabilized with digitonin. Efflux experiments show that in fact ryanodine as IP3 stimulates the release of calcium sequestered intracellularly. We further show that these effects of ryanodine are inhibited by Mg++ , ruthenium red and heparin. Our results suggest that ryanodine-sensitive intracellular calcium channels exist in the sea urchin egg.

Photochemical rearrangement of oxaziridines and nitrones in the hexahydroindole series: a convenient synthetic route to 1-azabicyclo[5.2.0]nonan-2-ones as novel RGD mimetics.

[reaction: see text] Photolysis of oxaziridines a or nitrones b provides a convenient synthetic route to fused bicyclic lactams c adequately substituted on both cycles A and B as scaffolds for mimicking conformationally constrained beta-turn peptides as in the tripeptide RGD signaling motif of fibronectin.

Distribution and function of T- and B-cell subpopulations in sarcoidosis.

Some in vitro functions of purified T and B cells from PPD- and Candida albicans-negative sarcoidosis patients were analyzed. It appears that in those patients there is: 1) A decrease in the absolute number of T lymphocytes and an increase in the number of B cells; 2) a rather normal response of T lymphocytes to PHA and Con A; 3) a rather normal capacity of T and B cells to produce MIF in vitro; and 4) an ability of T cells from sarcoid patients (but not B cells) to produce LMF. These results suggest that the frequent deficit in cell-mediated immunity observed in sarcoidosis seems to correlate with a quantitative deficit in T cells. The cause of this T-cell deficit is unknown.

Effect of antibacterial plant flavanones on the intracellular calcium compartment involved in the first cleavage of sea urchin eggs.

Two plant flavanones, (Sigmoidin A and B) having noticeable antibacterial activity, were assayed using a preparation for the study of sea urchin egg cleavage. When added after insemination, both toxins inhibit egg division with a half maximal dose of 7.5 microM for Sigmoidin A and 12 microM for Sigmoidin B. The first Ca2+ signal following fertilization was not modified by the molecules, however, the intracellular storage of calcium in isolated non-mitochondrial compartments was reduced in a dose-dependent manner by Sigmoidin A and Sigmoidin B. Both trigger a complete discharge of the sequestered calcium. In vivo the flavanones dramatically reduced the capacity of storage of non-mitochondrial intracellular calcium compartments necessary to the cyclical elevation of cytosolic free calcium during the cell cycle.

HgCl2-induced cell injury. Differential effects on membrane-located transport systems in unfertilized and fertilized sea urchin eggs.

The inhibitory potency of HgCl2 on amino acid and Na+ transport and the mechanism of action of this heavy metal are studied in unfertilized and fertilized sea urchin eggs, in which amino acid transport systems comparable to that described in mammalian somatic cells have been characterized. These transport systems called "L" for leucine, "ASC" for alanine, serine, cysteine, and "A" for alanine are differentiated mainly by their Na+-dependency and by the amino acids transported. The carrier-mediated amino acid uptake is reduced in a dose- and time-dependent manner by HgCl2, with a 50% inhibitory concentration (IC50) in the micromolar range. The mechanisms of inhibition of the Na+-independent (L system) and the Na+-dependent (A or ASC system) transport components are different: in unfertilized eggs, HgCl2 directly interferes with the L amino acid carrier leading to a decrease of its affinity for amino-acids, whereas in fertilized eggs the inhibition of the Na+-dependent uptake of amino acids may result from an elevation of Na+ content induced both by an inhibition of the Na+ pump and by an increase in Na+ permeability. It is also shown that the action of HgCl2 on amino acid diffusion differs between unfertilized and fertilized eggs. Our findings are discussed in the context of the role of membrane in xenobiotic toxicity.

The nitroxide stable radical tempo prevents metal-induced inhibition of CYP1A1 expression and induction.

Heavy metals are known to provoke oxidative stress in fish liver cells. Because H2O2, OH*- and intracellular superoxide are involved in this oxidation, we investigated the effect of nitroxide radical, 2,2,6,6-tetramethylpiperidinyl-N-oxyl (abbreviated as TEMPO), a cell-permeable agent possessing antioxidant properties, on CYP1A expression in trout (Oncorhynchus mykiss) hepatocytes. 3-methylcholanthrene (3-MC) induced the CYP1A-related EROD activity. This induction was inhibited by concomitant exposure to Cd (II), Cu (II), Pb (II) or Zn (II). CYP1A mRNA levels were also reduced. Simultaneous treatment with 3-MC, a heavy metal and TEMPO suppressed both the inhibition of EROD activity and the decrease of CYP1A mRNA expression. These results suggest a working hypothesis that heavy metals produce multiple oxidative effects, including generation of hydroxyl radicals, which could down-regulate CYP1A1 expression. This metal-induced inhibition was prevented by TEMPO, which might protect trout hepatocytes by scavenging free radicals and thus preventing their inhibitory effects on CYP1A induction and expression.

Embryotoxic potency of 2,4,5-trichlorophenoxyacetic acid on sea urchin eggs: Association with calcium homoeostasis.

The effect of 2,4,5-trichlorophenoxyacetic acid (2,4,5-T), a polychlorinated herbicide, on the early development of sea urchin eggs and on Ca(2+) permeability was investigated. Concentrations lower than 5 x 10(-4)m delayed the first cleavages and produced a teratogenic effect characterized by a large spectrum of structural malformations at the pluteus stage. The cleaving stage (pre-hatching) was the period most sensitive to the compound. Upper concentrations caused a stepwise dose-dependent lethality associated with arrest of cleavage. 2,4,5-T increased plasmalemma Ca(2+) permeability of unfertilized eggs by opening voltage-dependent Ca(2+) channels; verapamil (10(-4)m) and nifedipine (10(-4)m) abolished this effect. Ca(2+) permeability was also increased by 2,4,5-T after fertilization of the eggs. ATP-dependent intracellular sequestration of Ca(2+), measured in isolated cortices, was inhibited by 2,4,5-T. Ca(2+) movement was affected over a range of concentrations similar to those producing embryonic abnormalities and lethality. The results suggest that the teratogenic potency of 2,4,5-T is associated with delay in first cleavages and alterations in Ca(2+) homoeostasis.