Gray matter alterations in parosmia.

Parosmia is a common olfactory disorder. In this condition, odors are perceived in a different quality than usual. This distorted olfactory percept is typically reported to be unpleasant. Little is known about the pathophysiology of this phenomenon. Previous studies demonstrated smaller volumes of the olfactory bulbs in patients with parosmia compared to subjects without parosmia. In order to investigate structural brain alterations in areas beyond the olfactory bulb, in the current study voxel-based morphometry was applied. A group of 22 parosmic patients was compared with control subjects matched for age- and sex, who exhibited a similar performance in olfactory tests. Performing a whole brain analysis, we found profound gray matter volume loss in the left anterior insula in parosmic patients. In an additional volume of interest analysis including primary and secondary olfactory areas, we also found volume loss in the right anterior insula, the anterior cingulate cortex, the hippocampus bilaterally, and the left medial orbitofrontal cortex. Many of these areas are critically involved in olfactory quality discrimination and odor memory. The present results indicate that reduced gray matter volume in brain regions supporting odor discrimination and memory is related to disturbed olfactory sensation in parosmia.

Monitoring the effect of restoration measures in Indonesian peatlands by radar satellite imagery.

In the context of the ongoing climate change discussions the importance of peatlands as carbon stores is increasingly recognised in the public. Drainage, deforestation and peat fires are the main reasons for the release of huge amounts of carbon from peatlands. Successful restoration of degraded tropical peatlands is of high interest due to their huge carbon store and sequestration potential. The blocking of drainage canals by dam building has become one of the most important measures to restore the hydrology and the ecological function of the peat domes. This study investigates the capability of using multitemporal radar remote sensing imagery for monitoring the hydrological effects of these measures. The study area is the former Mega Rice Project area in Central Kalimantan, Indonesia, where peat drainage and forest degradation is especially intense. Restoration measures started in July 2004 by building 30 large dams until June 2008. We applied change detection analysis with more than 80 ENVISAT ASAR and ALOS PALSAR images, acquired between 2004 and 2009. Radar signal increases of up to 1.36 dB show that high frequency multitemporal radar satellite imagery can be used to detect an increase in peat soil moisture after dam construction, especially in deforested areas with a high density of dams. Furthermore, a strong correlation between cross-polarised radar backscatter coefficients and groundwater levels above -50 cm was found. Monitoring peatland rewetting and quantifying groundwater level variations is important information for vegetation re-establishment, fire hazard warning and making carbon emission mitigation tradable under the voluntary carbon market or REDD (Reducing Emissions from Deforestation and Degradation) mechanism.

Increased damage from fires in logged forests during droughts caused by El Niño.

In 1997-98, fires associated with an exceptional drought caused by the El Niño/Southern Oscillation (ENSO) devastated large areas of tropical rain forests worldwide. Evidence suggests that in tropical rainforest environments selective logging may lead to an increased susceptibility of forests to fire. We investigated whether this was true in the Indonesian fires, the largest fire disaster ever observed. We performed a multiscale analysis using coarse- and high-resolution optical and radar satellite imagery assisted by ground and aerial surveys to assess the extent of the fire-damaged area and the effect on vegetation in East Kalimantan on the island of Borneo. A total of 5.2 +/- 0.3 million hectares including 2.6 million hectares of forest was burned with varying degrees of damage. Forest fires primarily affected recently logged forests; primary forests or those logged long ago were less affected. These results support the hypothesis of positive feedback between logging and fire occurrence. The fires severely damaged the remaining forests and significantly increased the risk of recurrent fire disasters by leaving huge amounts of dead flammable wood.

Light affects cAMP signaling and cell movement activity in Dictyostelium discoideum.

The multicellular, slug stage of the slime mould Dictyostelium discoideum lacks specific sensory cells and organs but can nevertheless respond in a very sensitive manner to external stimuli such as temperature and light. Within the migrating slug, the behavior of up to 100,000 individual amoebae is coordinated by cAMP mediated cell-cell signaling and chemotaxis. We report here the striking result that light directly modulates the cAMP cell-cell signaling system. Light-induced secretion of cAMP from the slug tips decreased the period length of optical density waves and speeded up cell movement. A local effect of light on cAMP release within the slug tip could modulate cell movement within the slug and thus control its phototactic turning and orientation toward a light source.

4D confocal microscopy of Dictyostelium discoideum morphogenesis and its presentation on the Internet.

Methods to present three-dimensional (3D) and time series of 3D datasets (4D) are demonstrated using the recent advances in confocal microscopy and computer visualization. The process of cell sorting during tip formation in the slime mould Dictyostelium discoideum is examined as an example by in vivo confocal microscopy of spectrally different green fluorescent protein (GFP) variants as reporters of cell-type specific gene expression. Also, cell sorting of the co-aggregating slime mould species D. discoideum and D. mucoroides is observed using a GFP variant and a spectrally distinguishable fluorescent vital stain. The confocal data are handled as 3D and 4D datasets, their processing and the advantages of different methods of visualization are discussed step by step. Selected sequences of the experiments can be viewed on the Internet, giving a much better impression of the complex cellular movements during Dictyostelium morphogenesis than printed photographs.

Simultaneous detection of two GFP spectral mutants during in vivo confocal microscopy of migrating Dictyostelium cells.

A method is described that allows simultaneous measurement of two spectrally distinguishable green fluorescent protein (GFP) mutants with a confocal microscope. In contrast to previously described methods, neither UV excitation nor repetition of scans is required. Therefore the method is well-suited to the long-time observation of living cells in three-dimensional microscopy and time series recording, as demonstrated with GFP-expressing Dictyostelium discoideum cells.

Induction of optical density waves and chemotactic cell movement in Dictyostelium discoideum by microinjection of cAMP pulses.

The development of most multicellular organisms involves coordinated cell movement. The early aggregation of Dictyostelium cells has been shown to be mediated by chemotactic movement to propagating waves of cAMP. We have proposed that propagating waves of a chemoattractant, most likely cAMP, also control the movement of cells in mounds and slugs. We have now used periodic pressure injection of pulses of cAMP in the extracellular space of aggregation streams, mounds, and slugs to investigate whether these signals can be relayed and control cell movement, using quantitative digital time-lapse microscopy. Our major findings are (1) short (0.1 s) pulses of cAMP (10(7) molecules) were able to elicit optical density (OD) waves in fields of aggregating amoebae. They propagate from the micropipet outward and interact with endogenous OD waves. (2) Periodic injection of cAMP pulses into aggregation streams blocked the pulses coming from the center and led to the rapid accumulation of cells downstream of the pipet around the pipet. (3) Injection of pulses of cAMP into mounds elicited OD waves, which propagated from the pipet outward and interacted with the endogenous waves, indicating that the same propagator carries them. (4) Periodic microinjection of cAMP in the prespore zone of slugs led to accumulation of anterior-like cells around the micropipet followed by tip formation. Furthermore, the cAMP signal could control the spacing of the endogenous sorting pattern. These results strongly support the hypothesis that the optical density waves observed during early development up to the mound stage represent propagating cAMP waves. They suggest furthermore that cAMP is the morphogen that controls cell movements in slugs.

Twisted scroll waves organize Dictyostelium mucoroides slugs.

Cellular slime moulds (Dictyosteloids) are characterised by at least two different modes of slug migration. Most species, e.g. Dictyostelium mucoroides, produce a stalk continuously during slug migration, while a few species, e.g. Dictyostelium discoideum are characterised by stalk-less slug migration and only produce a stalk upon culmination. Experiments on D. discoideum and theoretical model calculations have shown that D. discoideum slugs are organized by a cAMP scroll wave in the tip which produces planar waves in the back. These waves guide cell movement in slugs: spiralling in the tip and forward movement parallel to the slug axis in the back. Simple changes in model parameters can lead to the formation of a twisted scroll wave which extends throughout the slug. In order to investigate whether such twisted scroll waves occur naturally we have analysed the movement of fluorescently labelled single cells in migrating D. mucoroides slugs. The results show that cells in the prespore zone of D. mucoroides slugs move in a spiral path. Although the velocity of single cells in D. mucoroides is faster than in D. discoideum, the net forward component of their movement is less due to their spiral trajectories. As a result D. mucoroides slugs move more slowly than D. discoideum slugs. The entire D. mucoroides slug also describes a spiralling path leaving corkscrew shaped stalks behind. Based on these observations we propose that cell movement in D. mucoroides slugs is controlled by a propagating twisted scroll wave of cAMP which extends throughout the length of the slug.

Analysis of cell movement and signalling during ring formation in an activated G alpha1 mutant of Dictyostelium discoideum that is defective in prestalk zone formation.

Mound formation in the cellular slime mould Dictyostelium results from the chemotactic aggregation of competent cells. Periodic cAMP signals propagate as multiarmed spiral waves and coordinate the movement of the cells. In the late aggregate stage the cells differentiate into prespore and several prestalk cell types. Prestalk cells sort out chemotactically to form the tip, which then controls all further development. The tip organises cell movement via a scroll wave that converts to planar waves in the prespore zone leading to rotational cell movement in the tip and periodic forward movement in the prespore zone. Expression of an activated G alpha1 protein under its own promoter leads to a severely altered morphogenesis from the mound stage onwards. Instead of forming a tipped mound, the cells form a ring-shaped structure without tip. Wave propagation pattern and dynamics during aggregation and mound formation in the mutant are indistinguishable from the parental strain AX3. However, at the time of tip formation the spiral waves that organise the late aggregate do not evolve in a scroll-organising centre in the tip but transform into a circularly closed (twisted) scroll ring wave. This leads to the formation of a doughnut-shaped aggregate. During further development, the doughnut increases in diameter and the twisted scroll wave converts into a train of planar waves, resulting in periodic rotational cell movement. Although biochemical consequences resulting from this mutation are still unclear, it must affect prestalk cell differentiation. The mutant produces the normal proportion of prespore cells but is unable to form functional prestalk cells, i.e., prestalk cells with an ability to sort out from the prespore cells and form a prestalk zone. Failure of sorting leads to an altered signal geometry, ring-shaped scroll waves, that then directs ring formation. This mutant demonstrates the importance of prestalk cell sorting for the stabilisation of the scroll wave that organises the tip.

Null mutations of the Dictyostelium cyclic nucleotide phosphodiesterase gene block chemotactic cell movement in developing aggregates.

Extracellular cAMP is a critical messenger in the multicellular development of the cellular slime mold Dictyostelium discoideum. The levels of cAMP are controlled by a cyclic nucleotide phosphodiesterase (PDE) that is secreted by the cells. The PDE gene (pdsA) is controlled by three promoters that permit expression during vegetative growth, during aggregation, and in prestalk cells of the older structures. Targeted disruption of the gene aborts development, and complementation with a modified pdsA restores development. Two distinct promoters must be used for full complementation, and an inhibitory domain of the PDE must be removed. We took advantage of newly isolated PDE-null cells and the natural chimerism of the organism to ask whether the absence of PDE affected individual cell behavior. PDE-null cells aggregated with isogenic wild-type cells in chimeric mixtures, but could not move in a coordinated manner in mounds. The wild-type cells move inward toward the center of the mound, leaving many of the PDE-null cells at the periphery of the aggregate. During the later stages of development, PDE-null cells in the chimera segregate to regions which correspond to the prestalk region and the rear of the slug. Participation in the prespore/spore population returns with the restoration of a modified pdsA to the null cells.

Periodic phenomena in Proteus mirabilis swarm colony development.

Proteus mirabilis colonies exhibit striking geometric regularity. Basic microbiological methods and imaging techniques were used to measure periodic macroscopic events in swarm colony morphogenesis. We distinguished three initial phases (lag phase, first swarming phase, and first consolidation phase) followed by repeating cycles of subsequent swarming plus consolidation phases. Each Proteus swarm colony terrace corresponds to one swarming-plus-consolidation cycle. The duration of the lag phase was dependent upon inoculation density in a way that indicated the operation of both cooperative and inhibitory multicellular effects. On our standard medium, the second and subsequent swarm phases displayed structure in the form of internal waves visible with reflected and dark-field illumination. These internal waves resulted from organization of the migrating bacteria into successively thicker cohorts of swarmer cells. Bacterial growth and motility were independently modified by altering the composition of the growth medium. By varying the glucose concentration in the substrate, it was possible to alter biomass production without greatly affecting the kinetics of colony surface area expansion. By varying the agar concentration in the substrate, initial bacterial biomass production was unaffected but colony expansion dynamics were significantly altered. Higher agar concentrations led to slower, shorter swarm phases and longer consolidation phases. Thus, colony growth was restricted by higher agar concentrations but the overall timing of the swarming-plus-consolidation cycles remained constant. None of a variety of factors which had significant effects on colony expansion altered terracing frequencies at 32 degrees C, but the length of the swarming-plus-consolidation cycle was affected by temperature and medium enrichment. Some clinical isolates displayed significant differences in terracing frequencies at 32 degrees C. Our results defined a number of readily quantifiable parameters in swarm colony development. The data showed no connection between nutrient (glucose) depletion and the onset of different phases in swarm colony morphogenesis. Several observations point to the operation of density-dependent thresholds in controlling the transitions between distinct phases.

The role of the cortical cytoskeleton: F-actin crosslinking proteins protect against osmotic stress, ensure cell size, cell shape and motility, and contribute to phagocytosis and development.

We generated Dictyostelium double mutants lacking the two F-actin crosslinking proteins alpha-actinin and gelation factor by inactivating the corresponding genes via homologous recombination. Here we investigated the consequences of these deficiencies both at the single cell level and at the multicellular stage. We found that loss of both proteins severely affected growth of the mutant cells in shaking suspension, and led to a reduction of cell size from 12 microns in wild-type cells to 9 microns in mutant cells. Moreover the cells did not exhibit the typical polarized morphology of aggregating Dictyostelium cells but had a more rounded cell shape, and also exhibited an increased sensitivity towards osmotic shock and a reduced rate of phagocytosis. Development was heavily impaired and never resulted in the formation of fruiting bodies. Expression of developmentally regulated genes and the final developmental stages that were reached varied, however, with the substrata on which the cells were deposited. On phosphate buffered agar plates the cells were able to form tight aggregates and mounds and to express prespore and prestalk cell specific genes. Under these conditions the cells could perform chemotactic signalling and cell behavior was normal at the onset of multicellular development as revealed by time-lapse video microscopy. Double mutant cells were motile but speed was reduced by approximately 30% as compared to wild type. These changes were reversed by expressing the gelation factor in the mutant cells. We conclude that the actin assemblies that are formed and/or stabilized by both F-actin crosslinking proteins have a protective function during osmotic stress and are essential for proper cell shape and motility.

Analysis of optical density wave propagation and cell movement during mound formation in Dictyostelium discoideum.

Aggregation fields of Dictyostelium amoebae are organized by propagating concentric or spiral waves of cAMP. These waves coordinate cell movement directed toward the aggregation center. We now systematically investigated dark-field wave propagation and chemotactic cell movement during late aggregation and mound formation. The period and the signal propagation velocity decreased continuously during aggregation leading to a 15-fold decrease of the chemical wavelength. By analyzing the behavior of single GFP-labeled cells in aggregates and mounds we measured cell movement velocity, changes in cell shape, periodicity of cell movement, and cell trajectories. In early mounds of strain AX-3 dark-field waves propagated frequently as multiarmed (high-frequency) spirals. During the high-frequency waves observed in the early mound stage, cell movement speed is low and cell movement rather undirected. During tip formation the wave period decreased again and the cells started to rotate in the mound at unusually high average speeds of 40 microns/min. The rotation was almost monotonic with no clear periodicity. Since at this time the majority of the cells had already differentiated into prespore cells, this implies that prespore cells moved faster than aggregation stage cells. At 12 hr of development cell movement velocity dropped again and became highly periodic. These measurements show that the relay system is characterized by a specific temporal evolution, which is closely correlated with cellular differentiation. The remarkable changes in cell movement speed and period indicate a qualitative change in signal and movement parameters which might well be caused by the observed switch from high- to low-affinity cAMP receptors during mound formation. This switch might be required to copy with the increase in cell density and most likely plays a crucial role in the process of cell sorting.

Spatial pattern formation during aggregation of the slime mould Dictyostelium discoideum.

Stream formation and spiral wave behaviour during the aggregation of Dictyostelium discoideum (Dd) are studied in a model based on the Martiel-Goldbeter equations for cAMP relay, combined with chemotactic motion of Dd cells. The results show that stream formation occurs if the turnover rate of intracellular cAMP is increased. This increase in the turnover rate of cAMP[in] leads to a dependence of the speed of the cAMP wave on the cell density. We propose that this dependence of wave speed on cell density is the underlying mechanism for stream formation. Besides stream formation, increasing the turnover rate of cAMP[in] also results in a spiral wave period that decreases during aggregation, a phenomenon that is commonly observed in situ. Furthermore, the dependence of wave speed on cell density is measured empirically. The speed of the cAMP wave is found to decrease as the wave travels from high to low cell density. This indicates that in situ, wave speed does depend on cell density.

Analysis of cell movement during the culmination phase of Dictyostelium development.

Co-ordinated cell movement of tens of thousands of cells and periodic signals characterise the multicellular development of the cellular slime mould Dictyostelium discoideum. We investigated cell movement by analysing time-lapse video recordings made during the slug stage and the culmination phase of Dictyostelium development. Slugs viewed from the side showed an even, straight forward movement with the tip slightly raised in the air. Slugs that had migrated for a prolonged period of time either culminated or showed a behaviour best described as abortive culmination. Culmination is initiated by a local aggregation of anterior-like cells at the base of the slug at the prestalk-prespore boundary, where they form a stationary mass of cells. Prespore cells continue to move forward over this stationary pile and, as a result, are lifted into the air. The stationary group of anterior-like cells thereby end up to the back of the slug. At this point the slug either falls back on the agar surface or continues culmination. If the slug continues to migrate these cells regain motility, move forward to the prespore-prestalk boundary and form a new pile again. In the case of culmination the neutral red stained cells in the pile move to the back of the slug and form a second signalling centre beside the tip. Both centres are characterised by vigorous rotational cell movement. The cells belonging to the basal centre will form the basal disc and the lower cup in the fruiting body. The upper cup will be formed by the prestalk cells rotating most vigorously at the prestalk-prespore boundary. The remaining neutral red stained anterior-like cells in the prespore zone sort either to the upper or lower organising centre in the fruiting body.

Spiral and concentric waves organize multicellular Dictyostelium mounds.

BACKGROUND: It has been known for more than 20 years that the early aggregation of the slime mould Dictyostelium is driven by periodic waves of cAMP, which instruct the cells to collect at the aggregation centre. Although it has been hypothesized that cAMP waves are also involved in the organization of multicellular morphogenesis, wave propagation in the later stages of Dictyostelium development has not previously been demonstrated. RESULTS: We have developed special optical and digital-image-processing techniques that allow propagating waves of chemotactic activity to be visualized in multicellular aggregates. Using this technology, we have observed signal propagation in the multicellular, 'mound' stage of Dictyostelium discoideum. Within mounds, these waves were propagated as concentric rings, single armed spirals or multi-armed spirals. The spontaneous appearance of the latter structures was new and unexpected. The geometry of wave propagation was strain specific: strain XP55 predominantly showed concentric ring waves, whereas spiral waves were typical of a derivative of XP55, streamer F mutant NP377, and of the widely used axenic strain AX-3. The different geometry of the signals was reflected by distinct cell-movement patterns and different cell-movement speeds--cells in AX-3 mounds, organized by spiral waves, moved faster than cells in XP55 mounds, and spiral waves were always accompanied by rotational cell movement, whereas cells in XP55 mounds moved towards the aggregation centre. CONCLUSIONS: The same principles--wave propagation and chemotaxis--that control Dictyostelium aggregation also govern the morphogenesis of the mound stage. Mounds behave as a highly excitable system in which a diverse range of signal-propagation geometries create the same biological structure--a migrating slug.

Three-dimensional scroll waves of cAMP could direct cell movement and gene expression in Dictyostelium slugs.

Complex three-dimensional waves of excitation can explain the observed cell movement pattern in Dictyostelium slugs. Here we show that these three-dimensional waves can be produced by a realistic model for the cAMP relay system [Martiel, J. L. & Goldbeter, A. (1987) Biophys J. 52, 807-828]. The conversion of scroll waves in the prestalk zone of the slug into planar wave fronts in the prespore zone can result from a smaller fraction of relaying cells in the prespore zone. Further, we show that the cAMP concentrations to which cells in a slug are exposed over time display a simple pattern, despite the complex spatial geometry of the waves. This cAMP distribution agrees well with observed patterns of cAMP-regulated cell type-specific gene expression. The core of the spiral, which is a region of low cAMP concentration, might direct expression of stalk-specific genes during culmination.

Patterns of cell movement within the Dictyostelium slug revealed by cell type-specific, surface labeling of living cells.

There are cells scattered in the rear, prespore region of the Dictyostelium slug that share many of the properties of the prestalk cells and that are therefore called anterior-like cells (ALCs). By placing the gene encoding a cell surface protein under the control of an ALC-specific promoter and immunologically labeling the living cells, we analyze the movement of ALCs within the slug. There is a posterior to anterior cellular flow, and the ALCs change their movement pattern as they enter the prestalk zone. Prestalk cells are periodically shed from the migrating slug. They must be replaced if the correct ratio of prestalk to prespore cells is to be maintained, and we present evidence for the transdifferentiation of prespore into prestalk cells, with ALCs functioning as intermediates in the transition. The slug has, therefore, a surprisingly dynamic structure, both with respect to cellular differentiation and cell movement.

A gradient method for the quantitative analysis of cell movement and tissue flow and its application to the analysis of multicellular Dictyostelium development.

We describe the application of a novel image processing method, which allows quantitative analysis of cell and tissue movement in a series of digitized video images. The result is a vector velocity field showing average direction and velocity of movement for every pixel in the frame. We apply this method to the analysis of cell movement during different stages of the Dictyostelium developmental cycle. We analysed time-lapse video recordings of cell movement in single cells, mounds and slugs. The program can correctly assess the speed and direction of movement of either unlabelled or labelled cells in a time series of video images depending on the illumination conditions. Our analysis of cell movement during multicellular development shows that the entire morphogenesis of Dictyostelium is characterized by rotational cell movement. The analysis of cell and tissue movement by the velocity field method should be applicable to the analysis of morphogenetic processes in other systems such as gastrulation and neurulation in vertebrate embryos.

Three-dimensional waves of excitation during Dictyostelium morphogenesis.

Cells in Dictyostelium slugs follow well-defined patterns of motion. We found that the chemotactic cell response is controlled by a scroll wave of messenger concentration in the highly excitable prestalk zone of the slug that decays in the less-excitable prespore region into planar wave fronts. This phenomenon is investigated by numerical solutions of partial differential equations that couple local nonlinear kinetics and diffusive transport of the chemotactic signal. In the interface of both regions a complex twisted scroll wave is formed that reduces the wave frequency in the prespore zone. The spatio-temporal dynamics of waves and filaments are followed over 33 periods of rotation. These results yield an explanation of collective self-organized cell motion in a multicellular organism.