Coherent optical transitions in implanted nitrogen vacancy centers.

We report the observation of stable optical transitions in nitrogen-vacancy (NV) centers created by ion implantation. Using a combination of high temperature annealing and subsequent surface treatment, we reproducibly create NV centers with zero-phonon lines (ZPL) exhibiting spectral diffusion that is close to the lifetime-limited optical line width. The residual spectral diffusion is further reduced by using resonant optical pumping to maintain the NV(-) charge state. This approach allows for placement of NV centers with excellent optical coherence in a well-defined device layer, which is a crucial step in the development of diamond-based devices for quantum optics, nanophotonics, and quantum information science.

Integrated high-quality factor optical resonators in diamond.

The realization of an integrated diamond photonic platform, based on a thin single crystal diamond film on top of a silicon dioxide/silicon substrate, is reported. Using this approach, we demonstrate high-quality factor single crystal diamond race-track resonators, operating at near-infrared wavelengths (1550 nm). The devices are integrated with low-loss diamond waveguides terminated with polymer pads (spot size converters) to facilitate in- (out-) coupling of light from (to) an optical fiber. Optical characterization of these resonators reveal quality factors as high as ~250,000 and overall insertion losses as low as 1 dB/facet. Scattering induced mode splitting as well as signatures of nonlinear effects such as optical bistability are observed at an input pump power of ~100 mW in the waveguides.

Coupling of NV centers to photonic crystal nanobeams in diamond.

The realization of efficient optical interfaces for solid-state atom-like systems is an important problem in quantum science with potential applications in quantum communications and quantum information processing. We describe and demonstrate a technique for coupling single nitrogen vacancy (NV) centers to suspended diamond photonic crystal cavities with quality factors up to 6000. Specifically, we present an enhancement of the NV center's zero-phonon line fluorescence by a factor of ~ 7 in low-temperature measurements.

Chemoautotrophy, symbiosis and sedimented diatoms support high biomass of benthic molluscs in the Namibian shelf.

The molluscs Lucinoma capensis, Lembulus bicuspidatus and Nassarius vinctus are highly abundant in Namibian oxygen minimum zone sediments. To understand which nutritional strategies allow them to reach such impressive abundances in this extreme habitat we investigated their trophic diversity, including a chemosymbiosis in L. capensis, focussing on nitrogen biochemical pathways of the symbionts. We combined results of bulk nitrogen and carbon (δ13C and δ15N) and of compound-specific isotope analyses of amino acid nitrogen (AAs-δ15NPhe and δ15NGlu), with 16S rRNA gene sequencing of L. capensis tissues and also with exploratory results of ammonium, nitrate and nitrite turnover. The trophic position (TP) of the bivalve L. capensis is placed between autotrophy and mixotrophy, consistent with its proposed symbiosis with sulfur-oxidizing Candidatus Thiodiazotropha sp. symbionts. The symbionts are here revealed to perform nitrate reduction and ammonium uptake, with clear indications of ammonium host-symbionts recycling, but surprisingly unable to fix nitrogen. The TP of the bivalve L. bicuspidatus is placed in between mixotrophy and herbivory. The TP of the gastropod N. vinctus reflected omnivory. Multiple lines of evidences in combination with current ecosystem knowledge point to sedimented diatoms as important components of L. bicuspidatus and N. vinctus' diet, likely supplemented at times with chemoautotrophic bacteria. This study highlights the importance of benthic-pelagic coupling that fosters the dietary base for macrozoobenthos in the OMZ. It further unveils that, in contrast to all shallow water lucinid symbionts, deeper water lucinid symbionts rely on ammonium assimilation rather than dinitrogen fixation to obtain nitrogen for growth.

B lymphocytes in vivo fail to prime naive T cells but can stimulate antigen-experienced T lymphocytes.

The ability of B cells or macrophages and dendritic cells (DC) to elicit class II-restricted T cell responses in vivo was compared using a mouse chimera model. Severe combined immunodeficient (SCID) mice (H-2d), reconstituted either with T or T+B lymphocytes from (H-2d x H-2b) donors, were immunized subcutaneously with protein antigen (Ag) to induce a class II-restricted T cell response. The frequency and major histocompatibility complex restriction of the resulting Ag-specific T cells were analyzed to establish whether B cells were necessary for the induction of class II-restricted T cell responses, and to determine the cell type on which priming had occurred. The results indicated that: (a) B cells are not necessary for the induction of a class II-restricted T cell response in vivo, as the frequencies of interleukin 2 (IL-2)- or IL-3-secreting T cells induced in the presence or absence of B cells were comparable. (b) Activation of naive T cells requires presentation of Ag on DC; Ag presented only on B cells is not sufficient to elicit a response. No H-2b-restricted, IL-3-secreting cells could in fact be detected in SCID mice reconstituted with naive (H-2d x H-2b) T cells and nonimmune or antigen-primed (H-2d x H-2b) B cells. (c) Previously primed T cells are able to be stimulated by Ag presented by both B cells and DC. H-2b-restricted, IL-3-secreting cells could in fact be readily demonstrated in SCID mice reconstituted with antigen-primed (H-2d x H-2b) T and B cells. Irrespective of whether the T cells were naive or previously activated, B cells were able to respond with an Ag-specific immunoglobulin G response, indicating that B cells were functional and able to present Ag in order to receive specific T cell help. Therefore, it appears that B cells are not necessary and do not participate in the initial priming of T cells; however, Ag presented by B cells can reactivate previously primed T cells. Taken together, these data indicate that during the course of an immune response Ag is first presented to naive T cells via DC, and only subsequently primed T cells can be stimulated by Ag presented by B cells.

Mice transgenic for a soluble form of murine CTLA-4 show enhanced expansion of antigen-specific CD4+ T cells and defective antibody production in vivo.

CD4+ T cell responses were analyzed in transgenic mice expressing a soluble form of murine CTLA-4, mCTLA4-H gamma 1, which blocks the interaction of the T cell activation molecules CD28 and CTLA-4 with their costimulatory ligands. Consistent with previous reports (Linsley, P. S., P. M. Wallace, J. Johnson, M. G. Gibson, J. L. Greene, J. A. Ledbetter, C. Singh, and M. A. Tepper. 1992. Science (Wash. DC). 257:792), T cell-dependent antibody production was profoundly inhibited in mCTLA4-H gamma 1 transgenic mice immunized with a protein antigen. Surprisingly, however, transgenic mice could generate quantitatively and qualitatively normal primary T cell responses, as measured by limiting dilution assays and lymphokine production. In addition, in vivo expansion of antigen-specific T cells after secondary or tertiary immunization was enhanced in mCTLA4-H gamma 1 transgenics as compared with normal mice. Although unable to deliver cognate help to B cells in vivo, T cells from mCTLA4-H gamma 1 transgenic mice were not anergic as they could help B cells to produce specific antibodies when adoptively transferred into nude hosts. Taken together, these data suggest that the engagement of CD28 and/or CTLA-4 may not be required for the induction of T cell responses, as is currently understood, but rather for the expression of T cell effector function such as the delivery of T cell help to B cells.

Signaling efficiency of the T cell receptor controlled by a single amino acid in the beta chain constant region.

A single amino acid residue, Gln136, located within the connecting peptide domain of Cbeta controls the ability of the alpha/beta TCR to transmit a full signal. TCRs in which this Cbeta residue is mutated to Phe, the residue found in TCR-gamma, are unresponsive to antigenic ligands. Interestingly, this Cbeta residue is either polar or charged in every species studied thus far, including the trout and the skate. In contrast, the analogous residue in Cgamma is always hydrophobic. In spite of their compromised antigen responsiveness, the mutant TCR complex contains the CD3-gamma, -delta, -epsilon, and -zeta chains, and undergoes zeta chain phosphorylation and ZAP-70 recruitment. However, the biological response of the mutant TCR could be rescued with a calcium ionophore, implying that mutant TCRs are defective in generating a calcium-mediated signal. The implications of the differences between Cbeta and Cgamma are considered.

Mta, the maternally transmitted antigen, is determined jointly by the chromosomal Hmt and the extrachromosomal Mtf genes.

Mus spretus from four stocks, originating in Spain, Portugal, and Morocco, were tested for the maternally transmitted antigen, Mta. All expressed a variant form not found in other species of mice. Analysis of appropriate crosses with inbred mice showed that the spretus form of Mta is determined by a new allele, c, of the Hmt gene. The Hmtc allele has been isolated in coupling with four different H-2 haplotypes. It is possible to raise CTL specific for the spretus form of Mta. The maternally transmitted factor, Mtf alpha s, of spretus mice determines, in conjunction with the Hmta allele of C57BL/6, an Mta that is indistinguishable from the common form found in C57BL/6 and most other inbred mice. Our experiments show that the specificity of the cell surface antigen Mta is governed jointly by the cytoplasmic gene Mtf and the chromosomal gene Hmt. We propose that Hmt encodes a class I histocompatibility antigen that acts as a restricting element for the Mtf gene product, thus meeting the requirements of T killer cell recognition.

Positive selection through a motif in the alphabeta T cell receptor.

The two lineages of T cells, alphabeta and gammadelta, differ in their developmental requirements: only alphabeta T cells require major histocompatibility complex recognition, a process known as positive selection. The alphabeta T cell receptor (TCR), but not its gammadelta counterpart, contains a motif within the alpha-chain connecting peptide domain (alpha-CPM) that has been conserved over the last 500 million years. In transgenic mice expressing an alphabeta TCR lacking the alpha-CPM, thymocytes were blocked in positive selection but could undergo negative selection. Thus, the alpha-CPM seems to participate in the generation of signals required for positive selection.

Cytoplasmic inheritance of a cell surface antigen in the mouse.

Mta is a cell surface antigen of the mouse and serves as a target for specific T killer lymphocytes. Using a killer cell assay, the antigen has been found in 72 strains of laboratory mice and, with one exception, in all tested samples of mice caught in the wild or bred from such, including Mus molossinus, Mus castaneus and Mus spretus. Five strains of rats, non-inbred NMRI mice, most substrains of NZB mice and the closely related strain NZO are negative for Mta. In reciprocal F1 crosses between several Mta+ and two Mta- strains, the antigen is maternally transmitted; that is, Mta+ females bear only positive offspring, whereas Mta- females bear only negative offspring, regardless of the genotype of the male. Since 34 foster-nursed mice had the Mta type of their genetic mothers, the factor that determines expression of Mta must be transmitted before birth and not via the milk. The cytoplasmic genes of Mta+ strains have been combined with the chromosomal genes of Mta- strains, and vice versa, by repeated backcrossing. All progeny retained the Mta type of their maternal lines. Thus, the Mta type is determined solely by maternal inheritance and is not influenced by chromosomal genes. We found no evidence of incompatibility between the cytoplasmic factors and nuclear genes of Mta- and Mta+ strains.

Cerebellar external granule cells are attached to the basal lamina from the onset of migration up to the end of their proliferative activity.

The development of cerebellar external granule cells in rats was studied from the time of demarcation of the cerebellar anlage on embryonal day 12 up to the time of their disappearance on postnatal day 20. Two types of cells were found. The first was orientated tangentially to the cerebellar surface and was characterized by a persistent contact to the basal lamina via an external process, with a lamellopodial tip and a cytoskeleton characteristic for migratory cells, and a retracting internal process featuring a single cilium. This cell type was the first to appear on embryonal day 14 in the caudolateral angle of the cerebellar anlage and, later, spread over the whole cerebellar surface. It disappeared after the external granular layer was completely expanded over the cerebellum. The second cell type appeared for the first time on embryonal day 16 in the caudal part of the cerebellar anlage and disappeared on postnatal day 20. It was orientated radially and also had contact with the basal lamina either with its cell body or with one or two short, radial processes, whose morphology differed from that of the external process of tangential cells by the absence of a lamellopodium and a prominent cytoskeleton. After postnatal day 17 contacts of external granule cells with the basal lamina decreased rapidly in length and number and were absent on postnatal day 20. We interpret these findings to indicate that tangential external granule cells are migrating before taking on a radial orientation characteristic for the mitotic cycle of proliferating external granule cells. In the light of increasing evidence implicating extracellular matrix in various developmental events of the nervous system we propose that the basal lamina of the cerebellum may be used as substrate and guidance structure by migrating external granule cells, and, furthermore, that the persistent contact with the basal lamina may mediate stimuli maintaining external granule cells in a proliferative state.

Fetal brain grafts rescue adult retinal ganglion cells from axotomy-induced cell death.

After intraorbital transection of the optic nerve of adult rats, 90% of the retinal ganglion cells die within 30 days. Since fetal brain extracts and cocultured fetal target regions support the survival of retinal ganglion cells in vitro (Nurcombe and Bennett: Exp. Brain Res. 44: 249-258, '81; McCaffery et al.: Exp. Brain Res. 48: 377-386, '82; Armson and Bennett: Neurosci. Lett. 38: 181-186, '83) we investigated whether cell death in the adult retina could be prevented by transplanting fetal (E16) thalamus and tectum to the proximal stump of the optic nerve of adult rats that was completely transected 2-3 mm behind the optic disc. Unoperated eyes contained 119,973 (+/- 939, SEM) retinal ganglion cells, estimated from axon counts of the intact optic nerve. Of these, 11,601 (+/- 1,857) remained in control operated eyes at 30 days postoperation while in the eyes of grafted rats, 35,086 (+/- 2,278) retinal ganglion cells were counted. Thus, 23,485 (= 22% of those normally dying after transection of the optic nerve) ganglion cells were rescued by the fetal grafts from cell death normally following axotomy. These results indicate that fetal target regions of retinal ganglion cells contain and/or produce neurotrophic molecules that promote the survival of adult axotomized retinal ganglion cells.

Regeneration of axons from the adult rat optic nerve: influence of fetal brain grafts, laminin, and artificial basement membrane.

After transection of the optic nerve of adult rats, most of the axons in the proximal stump die and the surviving ones are unable to regenerate into the distal optic nerve. Since the fetal brain has an inherent capacity to regenerate axons, we investigated whether fetal (E16) target regions of optic axons (thalamus and tectum) transplanted to the completely transected optic nerve of adult rats would promote axon regeneration. In control operated rats, axon growth beyond the site of transection was restricted to a few fibers that grew irregularly within the connective tissue scar. By contrast, in grafted animals directed outgrowth of optic axons toward the transplant started at 6 days postoperation (p.o.) and reached its maximum 15 days p.o. and later, when numerous single optic fibers and small axon fascicles had grown toward and into the graft, where they formed arborizations and terminal varicosities. Regenerating optic axons were further advanced than GFAP-positive strands of astroglia that emanated from the proximal optic nerve stump. Laminin immunoreactivity appeared at 6 days p.o. in the zone of reactive astroglia in the terminal part of the optic nerve stump. Later it showed a distribution complementary to the pattern of GFAP immunoreactivity, which it seemd to circumscribe. There was no unequivocal codistribution of laminin immunoreactivity with regenerating axons. In further experiments, target regions from different ontogenetic stages (E14 to neonate and adult) and nontarget regions (E16, cerebral cortex or spinal cord) were grafted to the optic nerve stump. With the exception of the adult grafts, all transplants had effects on axon regeneration comparable to those of E16 target regions. In order to test the effects of extracellular matrix molecules on axon regeneration, a basement membrane gel reconstituted from individual components of the Engelbreth-Holm-Sarcoma (EHS) sarcoma was implanted between proximal and distal optic nerve stumps. No axons were induced to regenerate by this matrix. Likewise, laminin adsorbed to nitrocellulose paper and implanted at the lesion site did not stimulate axon growth from the proximal optic nerve stump. These results indicate that fetal brain is able to induce and direct regrowth of axons from the optic nerve toward the graft across a substrate that is not composed of astroglia or basement membrane components like laminin. The directed growth of axons in the absence of a preformed substrate implies a chemotactic growth response along a concentration gradient mediated by neurotropic molecules released from the graft.

Derivation of cerebellar Golgi neurons from the external granular layer: evidence from explantation of external granule cells in vivo.

The present report provides evidence to challenge the traditional view that cerebellar Golgi cells are derived from the ventricular neuroepithelium, postulating instead that they originate from external granule cells. Supporting evidence for this assertion comes from three sources: 1) Typical Golgi cells are found in ectopic granule cell colonies, both outside the cerebellum (in the subarachnoid space) and also within the cerebellar cortex between fused folia. Because ectopic granule cell colonies are derived from external granule cells, which become displaced after treatment with 6-hydroxydopamine (6-OHDA), it was assumed that the ectopic Golgi cells also stem from such displaced external granule cells. 2) In order to demonstrate that Golgi cell precursors migrate from the external granular layer into the Purkinje cell plate, the development of the cerebellar cortex was studied over the period of Golgi cell genesis. On E19 the external granular layer in the rat is subdivided into an outer proliferative and an inner subproliferative zone. At the inner margin of the external granular layer, and in the marginal zone, radially oriented, darkly staining cells are present that exhibit all the characteristics of migrating neurons possessing a leading process oriented toward the Purkinje cell plate, a somatic cilium, and a close association with radial glia fibers. In later stages, these cells are also found deep to the Purkinje cell plate. Because Golgi cells arise during the period between E19 and postnatal day 2 in the rat (Altman and Bayer, '77, '78) and as the basket cells, the first neurons of proven origin from the external granular layer, are not produced before the second postnatal day (Altman, '72), the earlier migrating neurons are presumed to be Golgi cells. 3) Available data from cell kinetic 3H-thymidine studies show that there is no unequivocal evidence for Golgi cell genesis from the ventricular neuroepithelium, because, at the time of Golgi cell birth, ventricular and external granular stem cell populations are proliferating, and with the present methods it is not possible to decide which of these are the precursors of Golgi cells. Thus, taken together, the findings of this study show that Golgi cells are more likely to arise from the external granular layer than from the ventricular neuroepithelium. This concept would unify cerebellar histogenesis by proposing that projection neurons arise from the ventricular neuroepithelium, whereas all interneurons of the cerebellar cortex are descendants of the external granular layer.

Fibroblast growth factors promote the survival of adult rat retinal ganglion cells after transection of the optic nerve.

Basic and acidic fibroblast growth factors (FGF) were implanted next to the proximal stump of the transected optic nerve of adult rats, in order to assess whether these molecules have neurotrophic activity in vivo. Of the 119,973 +/- 2484 (S.E.M.) retinal ganglion cells present in retinae of unoperated control rats, 11,375 +/- 2413 (S.E.M.) remained at 30 days after transection of the optic nerve in control operated rats. After implantation of gel foam soaked in basic FGF, the number of retinal ganglion cells surviving at 30 days after axotomy tripled (36,387 +/- 3270 (S.E.M.], after acidic FGF, it increased almost 4-fold (40,916 +/- 5405 (S.E.M.]. These results indicate that FGF has neurotrophic activity in the adult central nervous system, and that this molecule is able to rescue adult retinal ganglion cells from axotomy induced cell death. It remains to be shown whether FGF acts directly on retinal ganglion cells or indirectly via glial cells or other cells.

Prognostic value of Doppler transmitral filling patterns in patients with chronic heart failure.

BACKGROUND: Chronic heart failure is a significant cause of cardiovascular morbidity and mortality. This study tested the hypothesis that restrictive filling pattern may provide useful prognostic data for identifying patients with chronic heart failure at high risk of all-cause cardiac death. METHODS: Ninety patients with chronic heart failure [70 men and 20 women, mean age (58.1 +/- 11.6) years] were investigated and followed for (18.8 +/- 7.9) months. During this period, 14 patients died of progressive pump failure, 12 patients underwent heart transplantation, 5 patients died suddenly, and 2 patients died of acute myocardial infarction. A new criterion, the restrictive filling index (RFI), was designed to subgroup patients into a restrictive and a nonrestrictive group. RESULTS: Patients with restrictive filling pattern had a more severe left ventricular dysfunction and a higher cardiac mortality. Analysis by the Kaplan-Meier method revealed that patients in the RFI > or = 1 and RFI < 1 groups had a cardiac events-free survival rate of 52% versus 94% at 1 year, and 27.5% versus 92% at 2 years, respectively. The multivariate Cox proportional hazard model selected RFI as the most powerful prognostic factor (chi(2) = 8.8017, P = 0.0030) for all-cause cardiac death. CONCLUSION: These results indicate that RFI is a simple, noninvasive, and specific clinical predictor for adult chronic heart failure patients who are at a high risk for all-cause cardiac death.

A robust scanning diamond sensor for nanoscale imaging with single nitrogen-vacancy centres.

The nitrogen-vacancy defect centre in diamond has potential applications in nanoscale electric and magnetic-field sensing, single-photon microscopy, quantum information processing and bioimaging. These applications rely on the ability to position a single nitrogen-vacancy centre within a few nanometres of a sample, and then scan it across the sample surface, while preserving the centre's spin coherence and readout fidelity. However, existing scanning techniques, which use a single diamond nanocrystal grafted onto the tip of a scanning probe microscope, suffer from short spin coherence times due to poor crystal quality, and from inefficient far-field collection of the fluorescence from the nitrogen-vacancy centre. Here, we demonstrate a robust method for scanning a single nitrogen-vacancy centre within tens of nanometres from a sample surface that addresses both of these concerns. This is achieved by positioning a single nitrogen-vacancy centre at the end of a high-purity diamond nanopillar, which we use as the tip of an atomic force microscope. Our approach ensures long nitrogen-vacancy spin coherence times (∼75 µs), enhanced nitrogen-vacancy collection efficiencies due to waveguiding, and mechanical robustness of the device (several weeks of scanning time). We are able to image magnetic domains with widths of 25 nm, and demonstrate a magnetic field sensitivity of 56 nT Hz(-1/2) at a frequency of 33 kHz, which is unprecedented for scanning nitrogen-vacancy centres.