Physical parameters, modeling, and methodological details in using IR laser pulses to warm frozen or vitrified cells ultra-rapidly

We report additional details of the thermal modeling, selection of the laser, and construction of the Cryo Jig used for our ultra-rapid warming studies of mouse oocytes (Jin et al., 2014). A Nd:YAG laser operating at 1064 nm was selected to deliver short 1ms pulses of sufficient power to produce a warming rate of 1×10(7)°C/min from -190°C to 0°C. A special Cryo Jig was designed and built to rapidly remove the sample from LN2 and expose it to the laser pulse. India ink carbon black particles were required to increase the laser energy absorption of the sample. The thermal model reported here is more general than that previously reported. The modeling reveals that the maximum warming rate achievable via external warming across the cell membrane is proportional to (1/R(2)) where R is the cell radius

Trehalose Is A Chemical Attractant In The Establishment Of Coral Symbiosis

Coral reefs have evolved with a crucial symbiosis between photosynthetic dinoflagellates (genus Symbiodinium) and their cnidarian hosts (Scleractinians). Most coral larvae take up Symbiodinium from their environment; however, the earliest steps in this process have been elusive. Here we demonstrate that the disaccharide trehalose may be an important signal from the symbiont to potential larval hosts. Symbiodinium freshly isolated from Fungia scutaria corals constantly released trehalose (but not sucrose, maltose or glucose) into seawater, and released glycerol only in the presence of coral tissue. Spawning Fungia adults increased symbiont number in their immediate area by excreting pellets of Symbiodinium, and when these naturally discharged Symbiodinium were cultured, they also released trehalose. In Y-maze experiments, coral larvae demonstrated chemoattractant and feeding behaviors only towards a chamber with trehalose or glycerol. Concomitantly, coral larvae and adult tissue, but not symbionts, had significant trehalase enzymatic activities, suggesting the capacity to utilize trehalose. Trehalase activity was developmentally regulated in F. scutaria larvae, rising as the time for symbiont uptake occurs. Consistent with the enzymatic assays, gene finding demonstrated the presence of a trehalase enzyme in the genome of a related coral, Acropora digitifera, and a likely trehalase in the transcriptome of F. scutaria. Taken together, these data suggest that adult F. scutaria seed the reef with Symbiodinium during spawning and the exuded Symbiodinium release trehalose into the environment, which acts as a chemoattractant for F. scutaria larvae and as an initiator of feeding behavior- the first stages toward establishing the coral-Symbiodinium relationship. Because trehalose is a fixed carbon compound, this cue would accurately demonstrate to the cnidarian larvae the photosynthetic ability of the potential symbiont in the ambient environment. To our knowledge, this is the first report of a chemical cue attracting the motile coral larvae to the symbiont

Scour holes and ripples occur below the hydraulic smooth to rough transition of movable beds

© 2017 The Authors. Scour holes often form in shallow flows over sand on the beach and in morphodynamic scale experiments of river reaches, deltas and estuarine landscapes. The scour holes are on average 2cm deep and 5cm long, regardless of the flow depth and appear to occur under similar conditions as current ripples: at low boundary Reynolds numbers, in fine sand and under relatively low sediment mobility. In landscape experiments, where the flow is only about 1cm deep, such scours may be unrealistically large and have unnatural effects on channel formation, bar pattern and stratigraphy. This study tests the hypotheses that both scours and ripples occur in the same conditions and that the roughness added by sediment saltation explains the difference between the ripple–dune transition and the clear-water hydraulic smooth to rough transition. About 500 experiments are presented with a range of sediment types, sediment mobility and obstructions to provoke scour holes, or removal thereof to assess scour hole persistence. Most experiments confirm that ripples and scour holes both form in the ripple stability field in two different bedform stability diagrams. The experiments also show that scours can be provoked by perturbations even below generalized sediment motion. Moreover, the hydraulic smooth to rough transition modified with saltation roughness depending on sediment mobility was similar in magnitude and in slope to ripple–dune transitions. Given uncertainties in saltation relations, the smooth to rough transitions modified for movable beds are empirically equivalent to the ripple–dune transitions. These results are in agreement with the hypothesis that scours form by turbulence caused by localized flow separation under low boundary Reynolds numbers, and do not form under generalized flow separation over coarser particles and intense sediment saltation. Furthermore, this suggests that ripples are a superposition of two independent forms: periodic bedforms occurring in smooth and rough conditions plus aperiodic scours occurring only in hydraulic smooth conditions

Improved estimation of Fokker-Planck equations through optimisation

An improved method for the description of hierarchical complex systems by means of a Fokker-Planck equation is presented. In particular the limited-memory Broyden-Fletcher-Goldfarb-Shanno algorithm for constraint problems (L-BFGS-B) is used to minimize the distance between the numerical solutions of the Fokker-Planck equation and the empirical probability density functions and thus to estimate properly the drift and diffusion term of the Fokker-Planck equation. The optimisation routine is applied to a time series of velocity measurements obtained from a turbulent helium gas jet in order to demonstrate the benefits and to quantify the improvements of this new optimisation routine

Morphological effects of vegetation on the tidal-fluvial transition in Holocene estuaries

Vegetation enhances bank stability and sedimentation to such an extent that it can modify river patterns, but how these processes manifest themselves in full-scale estuarine settings is poorly understood. On the one hand, tidal flats accrete faster in the presence of vegetation, reducing the flood storage and ebb dominance over time. On the other hand flow-focusing effects of a tidal floodplain elevated by mud and vegetation could lead to channel concentration and incision. Here we study isolated and combined effects of mud and tidal marsh vegetation on estuary dimensions. A 2-D hydromorphodynamic estuary model was developed, which was coupled to a vegetation model and used to simulate 100 years of morphological development. Vegetation settlement, growth and mortality were determined by the hydromorphodynamics. Eco-engineering effects of vegetation on the physical system are here limited to hydraulic resistance, which affects erosion and sedimentation pattern through the flow field. We investigated how vegetation, combined with mud, affects the average elevation of tidal flats and controls the system-scale planform. Modelling with vegetation only results in a pattern with the largest vegetation extent in the mixed-energy zone of the estuary, which is generally shallower. Here vegetation can cover more than 50 % of the estuary width while it remains below 10 %–20 % in the outer, tide-dominated zone. This modelled distribution of vegetation along the estuary shows general agreement with trends in natural estuaries observed by aerial image analysis. Without mud, the modelled vegetation has a limited effect on morphology, again peaking in the mixed-energy zone. Numerical modelling with mud only shows that the presence of mud leads to stabilisation and accretion of the intertidal area and a slight infill of the mixed-energy zone. Combined modelling of mud and vegetation leads to mutual enhancement with mud causing new colonisation areas and vegetation stabilising the mud. This occurs in particular in a zone previously described as the bedload convergence zone. While vegetation focusses the flow into the channels such that mud sedimentation in intertidal side channels is prevented on a timescale of decades, the filling of intertidal area and the resulting reduction in tidal prism may cause the infilling of estuaries over centuries

Effects of mud supply on large-scale estuary morphology and development over centuries to millennia

Alluvial river estuaries consist largely of sand but are typically flanked by mudflats and salt marshes. The analogy with meandering rivers that are kept narrower than braided rivers by cohesive floodplain formation raises the question of how large-scale estuarine morphology and the late Holocene development of estuaries are affected by cohesive sediment. In this study we combine sand and mud transport processes and study their interaction effects on morphologically modelled estuaries on centennial to millennial timescales. The numerical modelling package Delft3D was applied in 2-DH starting from an idealised convergent estuary. The mixed sediment was modelled with an active layer and storage module with fluxes predicted by the Partheniades–Krone relations for mud and Engelund–Hansen for sand. The model was subjected to a range of idealised boundary conditions of tidal range, river discharge, waves and mud input. The model results show that mud is predominantly stored in mudflats on the side of the estuary. Marine mud supply only influences the mouth of the estuary, whereas fluvial mud is distributed along the whole estuary. Coastal waves stir up mud and remove the tendency to form muddy coastlines and the formation of mudflats in the downstream part of the estuary. Widening continues in estuaries with only sand, while mud supply leads to a narrower constant width and reduced channel and bar dynamics. This self-confinement eventually leads to a dynamic equilibrium in which lateral channel migration and mudflat expansion are balanced on average. However, for higher mud concentrations, higher discharge and low tidal amplitude, the estuary narrows and fills to become a tidal delta

Sediment Transport of Fine Sand to Fine Gravel on Transverse Bed Slopes in Rotating Annular Flume Experiments

Large‐scale morphology, in particular meander bend depth, bar dimensions, and bifurcation dynamics, are greatly affected by the deflection of sediment transport on transverse bed slopes due to gravity and by secondary flows. Overestimating the transverse bed slope effect in morphodynamic models leads to flattening of the morphology, while underestimating leads to unrealistically steep bars and banks and a higher braiding index downstream. However, existing transverse bed slope predictors are based on a small set of experiments with a minor range of flow conditions and sediment sizes, and in practice models are calibrated on measured morphology. The objective of this research is to experimentally quantify the transverse bed slope effect for a large range of near‐bed flow conditions with varying secondary flow intensity, sediment sizes (0.17–4 mm), sediment transport mode, and bed state to test existing predictors. We conducted over 200 experiments in a rotating annular flume with counterrotating floor, which allows control of the secondary flow intensity separate from the streamwise flow velocity. Flow velocity vectors were determined with a calibrated analytical model accounting for rough bed conditions. We isolated separate effects of all important parameters on the transverse slope. Resulting equilibrium transverse slopes show a clear trend with varying sediment mobilities and secondary flow intensities that deviate from known predictors depending on Shields number, and strongly depend on bed state and sediment transport mode. Fitted functions are provided for application in morphodynamic modelin

Rectification of the Water Permeability in COS-7 Cells at 22, 10 and 0°C

The osmotic and permeability parameters of a cell membrane are essential physico-chemical properties of a cell and particularly important with respect to cell volume changes and the regulation thereof. Here, we report the hydraulic conductivity, Lp, the non-osmotic volume, Vb, and the Arrhenius activation energy, Ea, of mammalian COS-7 cells. The ratio of Vb to the isotonic cell volume, Vc iso, was 0.29. Ea, the activation energy required for the permeation of water through the cell membrane, was 10,700, and 12,000 cal/mol under hyper- and hypotonic conditions, respectively. Average values for Lp were calculated from swell/shrink curves by using an integrated equation for Lp. The curves represented the volume changes of 358 individually measured cells, placed into solutions of nonpermeating solutes of 157 or 602 mOsm/kg (at 0, 10 or 22°C) and imaged over time. Lp estimates for all six combinations of osmolality and temperature were calculated, resulting in values of 0.11, 0.21, and 0.10 µm/min/atm for exosmotic flow and 0.79, 1.73 and 1.87 µm/min/atm for endosmotic flow (at 0, 10 and 22°C, respectively). The unexpected finding of several fold higher Lp values for endosmotic flow indicates highly asymmetric membrane permeability for water in COS-7. This phenomenon is known as rectification and has mainly been reported for plant cell, but only rarely for animal cells. Although the mechanism underlying the strong rectification found in COS-7 cells is yet unknown, it is a phenomenon of biological interest and has important practical consequences, for instance, in the development of optimal cryopreservation

Ultra-Rapid Warming Yields High Survival of Mouse Oocytes Cooled to −196°C in Dilutions of a Standard Vitrification Solution

Intracellular ice is generally lethal. One way to avoid it is to vitrify cells; that is, to convert cell water to a glass rather than to ice. The belief has been that this requires both the cooling rate and the concentration of glass-inducing solutes be very high. But high solute concentrations can themselves be damaging. However, the findings we report here on the vitrification of mouse oocytes are not in accord with the first belief that cooling needs to be extremely rapid. The important requirement is that the warming rate be extremely high. We subjected mouse oocytes in the vitrification solution EAFS 10/10 to vitrification procedures using a broad range of cooling and warming rates. Morphological survivals exceeded 80% when they were warmed at the highest rate (117,000°C/min) even when the prior cooling rate was as low as 880°C/min. Functional survival was >81% and 54% with the highest warming rate after cooling at 69,000 and 880°C/min, respectively. Our findings are also contrary to the second belief. We show that a high percentage of mouse oocytes survive vitrification in media that contain only half the usual concentration of solutes, provided they are warmed extremely rapidly; that is, >100,000°C/min. Again, the cooling rate is of less consequence