Ocular effects of exposure to low-humidity environment with contact lens wear: A pilot study.

PURPOSE: To compare the ocular effects of exposure to a low-humidity environment with and without contact lens (CL) wear using various non-invasive tests. METHODS: Fourteen habitual soft CL wearers were exposed to controlled low humidity (5% relative humidity [RH]) in an environmental chamber for 90 min on two separate occasions. First, when wearing their habitual spectacles and then, on a separate visit, when wearing silicone hydrogel CLs that were fitted specifically for this purpose. All participants had adapted to the new CL prior to data collection. Three non-invasive objective measurements were taken at each visit: blinking rate, objective ocular scatter (measured using the objective scatter index) and ocular surface cooling rate (measured using a long-wave infrared thermal camera). At each visit, measurements were taken before the exposure in comfortable environmental conditions (RH: 45%), and after exposure to environmental stress (low humidity, RH: 5%). RESULTS: CL wearers showed increased blinking rate (p < 0.005) and ocular scatter (p = 0.03) but similar cooling rate of the ocular surface (p = 0.08) when compared with spectacle wear in comfortable environmental conditions. The exposure to low humidity increased the blinking rate significantly with both types of corrections (p = 0.01). Interestingly, ocular scatter (p = 0.96) and cooling rate (p = 0.73) were not significantly different before and after exposure to low humidity. There were no significant two-way interactions between correction and exposure in any of the measurements. CONCLUSIONS: CLs significantly increased the blinking rate, which prevented a quick degradation of the tear film integrity as it was refreshed more regularly. It is hypothesised that the increased blinking rate in CL wearers aids in maintaining ocular scatter quality and cooling rate when exposed to a low-humidity environment. These results highlight the importance of blinking in maintaining tear film stability.

Exceedances of Secondary Aerosol Formation from In-Use Natural Gas Heavy-Duty Vehicles Compared to Diesel Heavy-Duty Vehicles.

This work, for the first time, assessed the secondary aerosol formation from both in-use diesel and natural gas heavy-duty vehicles of different vocations when they were operated on a chassis dynamometer while the vehicles were exercised on different driving cycles. Testing was performed on natural gas vehicles equipped with three-way catalysts (TWCs) and diesel trucks equipped with diesel oxidation catalysts, diesel particulate filters, and selective catalytic reduction systems. Secondary aerosol was measured after introducing dilute exhaust into a 30 m3 environmental chamber. Particulate matter ranged from 0.18 to 0.53 mg/mile for the diesel vehicles vs 1.4-85 mg/mile for the natural gas vehicles, total particle number ranged from 4.01 × 1012 to 3.61 × 1013 for the diesel vehicles vs 5.68 × 1012-2.75 × 1015 for the natural gas vehicles, and nonmethane organic gas emissions ranged from 0.032 to 0.05 mg/mile for the diesel vehicles vs 0.012-1.35 mg/mile for the natural gas vehicles. Ammonia formation was favored in the TWC and was found in higher concentrations for the natural gas vehicles (ranged from ∼0 to 1.75 g/mile) than diesel vehicles (ranged from ∼0 to 0.4 g/mile), leading to substantial secondary ammonium nitrate formation (ranging from 8.5 to 98.8 mg/mile for the natural gas vehicles). For the diesel vehicles, one had a secondary ammonium nitrate of 18.5 mg/mile, while the other showed essentially no secondary ammonium nitrate formation. The advanced aftertreatment controls in diesel vehicles resulted in almost negligible secondary organic aerosol (SOA) formation (ranging from 0.046 to 2.04 mg/mile), while the natural gas vehicles led to elevated SOA formation that was likely sourced from the engine lubricating oil (ranging from 3.11 to 39.7 mg/mile). For two natural gas vehicles, the contribution of lightly oxidized lubricating oil in the primary organic aerosol was dominant (as shown in the mass spectra analysis), leading to enhanced SOA mass. Heavily oxidized lubricating oil was also observed to contribute to the SOA formation for other natural gas vehicles.

Process-Level Modeling Can Simultaneously Explain Secondary Organic Aerosol Evolution in Chambers and Flow Reactors.

Secondary organic aerosol (SOA) data gathered in environmental chambers (ECs) have been used extensively to develop parameters to represent SOA formation and evolution. The EC-based parameters are usually constrained to less than one day of photochemical aging but extrapolated to predict SOA aging over much longer timescales in atmospheric models. Recently, SOA has been increasingly studied in oxidation flow reactors (OFRs) over aging timescales of one to multiple days. However, these OFR data have been rarely used to validate or update the EC-based parameters. The simultaneous use of EC and OFR data is challenging because the processes relevant to SOA formation and evolution proceed over very different timescales, and both reactor types exhibit distinct experimental artifacts. In this work, we show that a kinetic SOA chemistry and microphysics model that accounts for various processes, including wall losses, aerosol phase state, heterogeneous oxidation, oligomerization, and new particle formation, can simultaneously explain SOA evolution in EC and OFR experiments, using a single consistent set of SOA parameters. With α-pinene as an example, we first developed parameters by fitting the model output to the measured SOA mass concentration and oxygen-to-carbon (O:C) ratio from an EC experiment (<1 day of aging). We then used these parameters to simulate SOA formation in OFR experiments and found that the model overestimated SOA formation (by a factor of 3-16) over photochemical ages ranging from 0.4 to 13 days, when excluding the abovementioned processes. By comprehensively accounting for these processes, the model was able to explain the observed evolution in SOA mass, composition (i.e., O:C), and size distribution in the OFR experiments. This work suggests that EC and OFR SOA data can be modeled consistently, and a synergistic use of EC and OFR data can aid in developing more refined SOA parameters for use in atmospheric models.

Cold air exposure at - 15 °C induces more airway symptoms and epithelial stress during heavy exercise than rest without aggravated airway constriction.

PURPOSE: Exposure to cold air may harm the airways. It is unclear to what extent heavy exercise adds to the cold-induced effects on peripheral airways, airway epithelium, and systemic immunity among healthy individuals. We investigated acute effects of heavy exercise in sub-zero temperatures on the healthy airways. METHODS: Twenty-nine healthy individuals underwent whole body exposures to cold air in an environmental chamber at - 15 °C for 50 min on two occasions; a 35-min exercise protocol consisting of a 5-min warm-up followed by 2 × 15 min of running at 85% of VO2max vs. 50 min at rest. Lung function was measured by impulse oscillometry (IOS) and spirometry before and immediately after exposures. CC16 in plasma and urine, and cytokines in plasma were measured before and 60 min after exposures. Symptoms were surveyed pre-, during and post-trials. RESULTS: FEV1 decreased after rest (- 0.10 ± 0.03 L, p < 0.001) and after exercise (- 0.06 ± 0.02 L, p = 0.012), with no difference between trials. Exercise in - 15 °C induced greater increases in lung reactance (X5; p = 0.023), plasma CC16 (p < 0.001) as well as plasma IL-8 (p < 0.001), compared to rest. Exercise induced more intense symptoms from the lower airways, whereas rest gave rise to more general symptoms. CONCLUSION: Heavy exercise during cold air exposure at - 15 °C induced signs of an airway constriction to a similar extent as rest in the same environment. However, biochemical signs of airway epithelial stress, cytokine responses, and symptoms from the lower airways were more pronounced after the exercise trial.

Emission of methylene diphenyl diisocyanate and methylene dianiline during use of cure-in-place methylene diphenyl diisocyanate-based consumer products.

The aim of this study was to provide realistic isocyanate and amine emission data when using different methylene diphenyl diisocyanate (MDI)-based polyurethane consumer products. Emission testing (air sampling) of diisocyanates and corresponding diamines was performed in a full-scale controlled-environment chamber during different work operations, such as gluing, mixing and foaming. The polyurethane products used were construction glue, one-component foam and two different two-component adhesives used in parquet flooring. Air sampling for isocyanates and amines was performed in the breathing zone of the worker and at different positions inside the controlled-environment chamber while the work operations were performed. Air sampling was also performed after the application, at different positions inside the chamber, to cover the post curing phase. Low air concentrations (0.1-0.7 µg MDI/m3, 0.03-0.2 µg isocyanate group (NCO)/m3) were found in the breathing zone and close to the work operation for some of the gluing applications. No methylene diphenyl diamine (MDA) concentrations above the limit of quantification were found for any of the applications in the breathing zone air. These results indicated that inhalation exposure to MDA or MDI would be expected to be minimal during application of do-it-yourself consumer products containing MDI.

Space Environmental Chamber for Planetary Studies.

We describe a versatile simulation chamber that operates under representative space conditions (pressures from < 10-5 mbar to ambient and temperatures from 163 to 423 K), the SpaceQ chamber. This chamber allows to test instrumentation, procedures, and materials and evaluate their performance when exposed to outgassing, thermal vacuum, low temperatures, baking, dry heat microbial reduction (DHMR) sterilization protocols, and water. The SpaceQ is a cubical stainless-steel chamber of 27,000 cm3 with a door of aluminum. The chamber has a table which can be cooled using liquid nitrogen. The chamber walls can be heated (for outgassing, thermal vacuum, or dry heat applications) using an outer jacket. The chamber walls include two viewports and 12 utility ports (KF, CF, and Swagelok connectors). It has sensors for temperature, relative humidity, and pressure, a UV-VIS-NIR spectrometer, a UV irradiation lamp that operates within the chamber as well as a stainless-steel syringe for water vapor injection, and USB, DB-25 ports to read the data from the instruments while being tested inside. This facility has been specifically designed for investigating the effect of water on the Martian surface. The core novelties of this chamber are: (1) its ability to simulate the Martian near-surface water cycle by injecting water multiple times into the chamber through a syringe which allows to control and monitor precisely the initial relative humidity inside with a sensor that can operate from vacuum to Martian pressures and (2) the availability of a high-intensity UV lamp, operating from vacuum to Martian pressures, within the chamber, which can be used to test material curation, the role of the production of atmospheric radicals, and the degradation of certain products like polymers and organics. For illustration, here we present some applications of the SpaceQ chamber at simulated Martian conditions with and without atmospheric water to (i) calibrate the ground temperature sensor of the Engineering Qualification Model of HABIT (HabitAbility: Brines, Irradiation and Temperature) instrument, which is a part of ExoMars 2022 mission. These tests demonstrate that the overall accuracy of the temperature retrieval at a temperature between -50 and 10 °C is within 1.3 °C and (ii) investigate the curation of composite materials of Martian soil simulant and binders, with added water, under Martian surface conditions under dry and humid conditions. Our studies have demonstrated that the regolith, when mixed with super absorbent polymer (SAP), water, and binders exposed to Martian conditions, can form a solid block and retain more than 80% of the added water, which may be of interest to screen radiation while maintaining a low weight.

Electron Microscopic Recording of the Power and Recovery Strokes of Individual Myosin Heads Coupled with ATP Hydrolysis: Facts and Implications.

The most straightforward way to get information on the performance of individual myosin heads producing muscle contraction may be to record their movement, coupled with ATP hydrolysis, electron-microscopically using the gas environmental chamber (EC). The EC enables us to visualize and record ATP-induced myosin head movement in hydrated skeletal muscle myosin filaments. When actin filaments are absent, myosin heads fluctuate around a definite neutral position, so that their time-averaged mean position remains unchanged. On application of ATP, myosin heads are found to move away from, but not towards, the bare region, indicating that myosin heads perform a recovery stroke (average amplitude, 6 nm). After exhaustion of ATP, myosin heads return to their neutral position. In the actin⁻myosin filament mixture, myosin heads form rigor actin myosin linkages, and on application of ATP, they perform a power stroke by stretching adjacent elastic structures because of a limited amount of applied ATP ≤ 10 µM. The average amplitude of the power stroke is 3.3 nm and 2.5 nm at the distal and the proximal regions of the myosin head catalytic domain (CAD), respectively. The power stroke amplitude increases appreciably at low ionic strength, which is known to enhance Ca2+-activated force in muscle. In both the power and recovery strokes, myosin heads return to their neutral position after exhaustion of ATP.

Ocular response to environmental variations in contact lens wearers.

PURPOSE: To assess the influence of different indoor environments simulated in an environmental chamber on soft contact lens (CL) wearers. METHODS: Fifty-four CL wearers were grouped based on their symptoms while wearing their CLs. Subjects were fitted with two different CL types, conventional hydrogel (Omafilcon A) and silicone hydrogel (comfilcon A), and exposed to two controlled indoor environmental conditions, standard [50% relative humidity (RH), 23°C, 930 mb of atmospheric pressure] and adverse (in-flight air cabin environment: 5% RH, localised air flow, 23°C, 750 mb atmospheric pressure), for 90 min in an environmental chamber, making a total of four visits. Symptoms, tear osmolarity, pre-lens tear breakup time (PLBUT), phenol red thread test, visual acuity, bulbar and limbal hyperaemia, corneal and limbal staining, and CL dehydration were assessed using repeated measures analysis of variance. A linear mixed model was used to analyse the effect of environment, CL type, discomfort-based grouping, and time on blink rate. RESULTS: Environment was found to significantly (p ≤ 0.018) affect limbal and bulbar hyperaemia, PLBUT, tear osmolarity, and CL dehydration. Likewise, CL type significantly (p ≤ 0.04) affected nasal and total corneal staining, limbal conjunctival staining, CL dehydration, comfort, and blurred vision. The environment, CL type, and time had significant effects (p ≤ 0.0001) on the blink rate. CONCLUSIONS: Ocular surface integrity and blink rate in CL wearers depend on the environment CL users are exposed to, as well as on the soft CL type that they are wearing. Tight control of environmental conditions can contribute to a better understanding of CL-related discomfort.

Budget-limited thermal biology: Design, construction and performance of a large, walk-in style temperature-controlled chamber.

We describe a partial redesign of the conventional air-conditioning system and apply it to the construction of a relatively large (1.87m(3) air mass), walk-in style temperature-controlled chamber (TCC) using parts easily obtained in most countries. We conducted several tests to demonstrate the performance of the TCC. Across the physiologically relevant range of 5-37°C, the TCC took 26.5-50.0min to reach the desired set point temperature. Once at set point, temperature inside the chamber was controlled with an accuracy of ±1.0°C. User-entry effects on deviations from and return times to set point temperature were minimal. Overall, performance of the TCC was sufficient to make precise physiological measurements of insect metabolic rate while controlling assay temperature. Major advantages of the TCC include its simplicity, flexibility, and low cost.

Trehalose phosphate synthase overexpression in Parachlorella kessleri improves growth and photosynthetic performance under high light conditions.

Parachlorella kessleri is a promising oil-bearing marine alga which shows decreased growth under high light stress. Osmolytes are known to relieve stress by protecting the cell membrane, proteins, and enzymes. Enhanced production of osmolyte (trehalose) was thus used to relieve stress in P. kessleri by overexpression of trehalose phosphate synthase (TPS) gene. Transformed P. kessleri was grown under different light regimes to study the effect of trehalose overproduction on growth. Study of one of the TPS transformants showed increased trehalose as well as increased biomass and decreased pigments, reactive oxygen species, and lipid peroxidation of cell membrane. The improved photosynthetic performance of the transformant was also signified by pulse-amplitude-modulated fluorometric analysis. All of these factors reveal improved stress tolerance under high light conditions by increased trehalose accumulation due to TPS overexpression in P. kessleri.

Insights into the formation of secondary organic aerosols from agricultural residue burning emissions: A review of chamber-based studies.

Organic aerosols (OA) are a significant component of fine particulate matter in the ambient air and are formed through primary and secondary processes. Primary organic aerosols (POA) are directly released from sources, while secondary organic aerosols (SOA) are formed through the oligomerization and/or oxidation of volatile organic compounds (VOCs) in the atmosphere. Recently, there has an increasing attention on the SOA budgets, their formation pathways, and photochemical evolution due to their impacts on climate and human health. Biomass burning (BB) is a significant source of OA, contributing around 5-30 % to the SOA burden globally. Agricultural residue burning (ARB) is a type of BB that contributes ~10 % of total atmospheric OA mass worldwide, whereas it contributes higher in Asian regions like China and India. ARB emits a significant amount of air pollutants, including VOCs, into the atmosphere. However, there is inadequate information on the transformation of ARB emissions to SOA due to limited laboratory studies. The present review focuses on the formation mechanism of SOA from ARB emissions, summarizing the current state of the art about ARB precursors and their oxidation products from chamber-based studies, including measurement methods and analytical instrumentation. The review also discusses the role of different types of oxidants in OA mass enhancement, factors affecting the overall SOA yield, and the uncertainties involved in the process.

A multisource mass transfer model for simulating VOC emissions from paints.

Indoor decoration generates a large number of volatile organic compounds (VOCs), which are simultaneously released from different paints. Nevertheless, the interaction mechanism of pollutant diffusion from multisource building materials (such as primer and finish) needs to be examined. In this paper, a multisource mass transfer model for VOC emissions from different combinations of paints is established, and the analytical solution is derived. The finite difference method is used to simulate the experimental results of VOC release in the environmental chambers, and its convergence and stability are verified. Using the optimization parameters of the single-source model and the law of conservation of mass, the key parameters of the multisource mass transfer model are obtained. The results show that the established model is in excellent agreement with both experimental data and literature data. In addition, the Little number Lt is used to analyse the change trend from the initial released concentration in the single-source and multisource models.

Effect of temperature on embryonation of Ascaris suum eggs in an environmental chamber.

The influence of temperature on the development and embryonation of Ascaris suum eggs was studied using coarse sand medium in an environmental chamber with 50% humidity. The time required for development and embryonation of eggs was examined under 3 different temperature conditions, 5°C, 25°C, and 35°C. A. suum eggs did not develop over 1 month at the temperature of 5°C. However, other temperature conditions, 25°C and 35°C, induced egg development to the 8-cell-stage at days 5-6 after incubation. All eggs examined developed to the 8-cell stage at day 6 after incubation in the sand medium at 25°C. The higher temperature, 35°C, slightly accelerated the A. suum egg development compared to 25°C, and the development to the 8-cell stage occurred within day 5 after incubation. The formation of larvae in A. suum eggs at temperatures of 35° and 25°C appeared at days 17 and 19 after incubation, respectively. These findings show that 35° condition shortens the time for the development of A. suum eggs to the 8-cell-stage in comparison to 25°C, and suggest the possibility of accelerated transmission of this parasite, resulting from global warming and ecosystem changes.

Suspension of pheromone microcapsules on vine leaves acting as passive dispensers against pests.

Pheromones are increasingly used as alternatives to pesticides to protect vineyards against L. botrana, a key grape pest. To diffuse (7E,9Z)-7,9-dodecadien-1-ylacetate, the L. botrana pheromone, passive, or aerosol dispensers are commonly applied. This paper deals with another method based on spraying an aqueous formulation, Lobesia Pro Spray, containing the pheromone encapsulated in a resin. The objectives were to assess the ability of vine leaves to act as pheromone dispensers and to check that encapsulation protects the plant from pheromone penetration. Laboratory testing based on an emission cell combined with airborne pheromone measurements by active sampling on sorbent tubes followed by ATD-GC-MS analysis was developed to accurately characterise the release of the pheromone into the air. Release kinetics analysis performed on the vine leaves showed a high pheromone release (about 30% of the sprayed quantity) the first day of the test. The release rate then decreased rapidly to reach about 650 mg/day/ha after 4 days. Kinetic modelling showed that it would be possible to maintain an effective airborne concentration of pheromone for approximately 12 days. Release tests were also carried out on glass, PVC and blotting paper. The results obtained showed that the vine leaves behaved as a non-absorbent material, implying that the pheromone used in the Lobesia Pro Spray formulation did not penetrate the plant. These first results prove the feasibility of using vine leaves as pheromone dispensers for a sprayed formulation and the ability to optimise the treatment conditions (dose and frequency) through laboratory testing.

A natural environmental chamber study on the emissions and fate of organophosphate esters in the indoor environment.

In this study, we investigated the emission and fate of 9 organophosphate esters (OPEs) from a natural environment chamber, in which three environment matrices (i.e., air, dust, and window film samples) as well as three decoration materials (i.e., laminate flooring, latex paint, and nonwoven paper) were collected within gradient variation of room temperature and relative humidity. ΣAlkyl-OPEs and ΣCl-OPEs were the predominant classes in the three environment matrices, accounting - on average - for 98.7%, 99.8% and 99.3% of ΣOPEs in indoor dust, air and window film, respectively. TBOEP was the most abundant OPE in air, dust, and laminate flooring, respectively, while tris (2-chloro-isopropyl) phosphate (TCIPP) and tris (1,3-dichloro-2-propyl) phosphate (TDCIPP) in nonwoven paper and latex paint, respectively. The results showed that higher room temperature expedited the emission of OPEs to indoor air. However, the room temperature and relative humidity had no effect on the levels of OPEs in dust. The OPEs equilibrium time in indoor environment may be dependent on room temperature and relative humidity. The area specific emission rates (SERs) of the three materials were calculated, and an optimal expression based on the concept of mass balance model was constructed, preliminarily revealing a general relationship between OPEs source and sink effects in indoor environment.

An experimental exposure study revealing composite airway effects of physical exercise in a subzero environment.

Exposure to a cold climate is associated with an increased morbidity and mortality, but the specific mechanisms are largely unknown. People with cardiopulmonary disease and winter endurance athletes are particularly vulnerable. This study aimed to map multiple domains of airway responses to exercise in subzero temperature in healthy individuals.Thirty-one healthy subjects underwent whole-body exposures for 50 minutes on two occasions in an environmental chamber with intermittent moderate-intensity exercise in +10 °C and -10 °C. Lung function, plasma/urine CC16 , and symptoms were investigated before and after exposures.Compared to baseline, exercise in -10 °C decreased FEV1 (p=0.002), FEV1/FVC (p<0.001), and increased R20Hz (p=0.016), with no differences between exposures. Reactance increased after +10 °C (p=0.005), which differed (p=0.042) from a blunted response after exercise in -10 °C. Plasma CC16 increased significantly within exposures, without differences between exposures. Exercise in -10 °C elicited more intense symptoms from the upper airways, compared to +10 °C. Symptoms from the lower airways were few and mild. Short-duration moderate-intensity exercise in -10 °C induces mild symptoms from the lower airways, no lung function decrements or enhanced leakage of biomarkers of airway epithelial injury, and no peripheral bronchodilatation, compared to exercise in +10 °C.

Qualitative identification and characterisation of self-reported symptoms arising in humans during experimental exposure to cold air.

BACKGROUND: Exposure to cold air is associated with increased morbidity and mortality in the general population. It is difficult to study the effects of whole-body exposure to cold air under controlled conditions in real life. OBJECTIVES: The aim of this study was to (1) explore and describe the experience of symptoms in humans during experimental and controlled exposures to cold air, by using controlled environmental chamber exposures and qualitative methodology, and to (2) categorise the symptoms. METHOD: The study used a randomised, double blind design, in which 34 subjects undertook rest and moderate-intensity exercise in an environmental chamber set to two or three different temperatures (0, -10, and -17°C) on separate occasions. During the chamber exposures, subjects were interviewed. Qualitative content analysis was selected as the method of analysis. FINDINGS: Subjects reported 50 distinct symptoms during the exposures. The symptoms were grouped into ten sub-categories and two major categories; airway versus whole-body symptoms. CONCLUSION: We have identified a broad range of symptoms in humans undertaking rest and moderate-intensity exercise at sub-zero temperatures. The symptoms and their categories may well be used to more extensively and quantitatively map cold-induced morbidity.

Constraining Aging Processes of Black Carbon in the Community Atmosphere Model Using Environmental Chamber Measurements.

The direct radiative forcing of black carbon aerosol (BC) on the Earth system remains unsettled, largely due to the uncertainty with physical properties of BC throughout their lifecycle. Here we show that ambient chamber measurements of BC properties provide a novel constraint on the crude BC aging representation in climate models. Observational evidence for significant absorption enhancement of BC can be reproduced when the aging processes in the four-mode version of the Modal Aerosol Module (MAM4) aerosol scheme in the Community Atmosphere Model version 5 are calibrated by the recent in situ chamber measurements. An observation-based scaling method is developed in the aging timescale calculation to alleviate the influence of biases in the simulated model chemical composition. Model sensitivity simulations suggest that the different monolayer settings in the BC aging parameterization of MAM4 can cause as large as 26% and 24% differences in BC burden and radiative forcing, respectively. We also find that an increase in coating materials (e.g., sulfate and secondary organic aerosols) reduces BC lifetime by increasing the hygroscopicity of the mixture but enhances its absorption, resulting in a net increase in BC direct radiative forcing. Our results suggest that accurate simulations of BC aging processes as well as other aerosol species are equally important in reducing the uncertainty of BC forcing estimation.

Basic properties of ATP-induced myosin head movement in hydrated myosin filaments, studied using the gas environmental chamber.

Although more than 50 years have passed since the monumental discovery of Huxley and Hanson that muscle contraction results from relative sliding between actin and myosin filaments, coupled with ATP hydrolysis, the mechanism underlying the filament sliding still remains to be a mystery. It is generally believed that the myofilament sliding is caused by cyclic attachment-detachment between myosin heads in myosin filaments and myosin-binding sites in actin filaments. Attempts to prove the myosin head movement using techniques of X-ray diffraction and chemical probes attached to myosin heads have failed to obtain clear results because of the asynchronous nature of myosin head movement. Using the gas environmental chamber (EC) attached to an electron microscope, we succeeded in recording myosin head movement in hydrated myosin filaments, coupled with ATP hydrolysis with the following results: (1)In the absence of actin filaments, myosin heads fluctuate around a definite neutral position, so that their time-averaged position remains unchanged; (2) On ATP application, myosin heads bind with ATP to be in the charged-up state, M-ADP-Pi, and perform a recovery stroke in the direction away from the myosin filament central bare zone and stay in the post-recovery stroke position; (3) In the actin-myosin filament mixture, myosin heads form rigor linkages with actin, and bind with applied ATP to be in the charged-up state, M-ADP-Pi, and perform a power stroke in the direction towards the myosin filament bare zone, while releasing ADP and Pi to stay in the post-power stroke position; (4) In both recovery and power strokes, myosin heads in the non charged-up state return to the neutral position. These results indicate that the charged-up myosin heads decide their direction of movement without being guided by actin filaments.

Automated Multichamber Time-lapse Videography for Long-term In Vivo Observation of Migrating Cells.

AIM: To observe and document the migration of living cells by time-lapse videography, we constructed a low-budget system based on a common inverted microscope. MATERIALS AND METHODS: Long-term observation of six-well plates is enabled through maintenance of cell culture conditions (5% CO2 in air at 37°C). Points of interest can be revisited in definable intervals with <1 µm repositioning error. Digital photographs from each programmed time point are paired with environmental data and combined into a record. RESULTS: We used this new chamber to observe the migration of various cell lines. The design represents a good compromise between low cost and good precision. Detailed analyses verified that the environmental conditions were appropriately maintained, enabling long-term observation of viable cells. The stimulating influence of irradiation with photons (radiotherapy) on cellular motility of glioblastoma cells is presented. CONCLUSION: This study demonstrates that useful videographic systems can be constructed at low cost.