Effects of calcium channel blockers and DCMU on motility and the photophobic response of Gyrodinium dorsum

The effects of the calcium channel blockers, verapamil, diltiazem and lanthanum ions and the Ca²⁺ dependency on motility as well as the photophobic response (stop-response) of Gyrodinium dorsum were studied. At Ca²⁺ concentrations below 10^-3 M, motility was inhibited. La³⁺ inhibits the stop-response, in contrast to verapamil and diltiazem. The only calcium channel blocker that increased the amount of non-motile cells was verapamil. The results indicate that motility are Ca²⁺ dependent and that the stop-responses of G. dorsum could be affected by extracellular Ca²⁺. Effects of the photosythesis inhibitor (DCMU) on the stop-response was also determined. With background light of different wavelength (614, 658 and 686 nm) the stop-response increased. DCMU inhibited this effect of background light. Negative results with the monoclonal antibody Pea-25 directed to phytochrome and the results with DCMU, indicate that the stop-response of G. dorsum is coupled to photosynthesis rather than to a phytochrome-like pigment. Oxygen evolution, but not cell movement, was completely inhibited by 10^-6 M DCMU.Abbreviations DCMU 3-(3,4-dichlorophenyl)-1,1-methylurea - DILT diltiazem - DMSO dimethylsulfoxide - SDS-PAGE sodium dodecyl sulphate-polyacrylamide gel electrophoresis - VER verapamil     

Huge increase in bacterivores on freshly killed barley roots

Abstract Adding fresh roots to intact soil cores resulted in marked increases in microbial and microfaunal activity at the resource islands. Microbial activity increased in two phases following root addition. Respiratory activity and concentration of respiratory enzyme (dehydrogenase) in soil adhering to the roots was very high during the first three weeks resulting in anaerobic conditions in the soil. After a period of low respiratory activity and enzyme content, these quantities increased from 6 to 20 weeks, but not enough to maintain anaerobic conditions. Numbers of protozoa peaked earlier than the nematodes. Based on yield coefficients of microbes and bacterivores, the increase in bacterivores was in accordance with root-induced respiration activity. In soil adhering to roots, numbers of bacterial grazers (protozoa and nematodes) were up to 80 and 30 times higher, respectively, than in the surrounding soil. This effect is up to 20 times higher than observed around live root systems, which may suggest that the rhizosphere effect on microbivores could for the major part result from the decomposition of dead segments of the root system.     

PACAP, a VIP-like peptide, in neurons of the esophagus.

The lower esophagus of guinea-pig, cat, sheep and man was analyzed for pituitary adenylate cyclase activating peptide (PACAP), a novel vasoactive intestinal peptide (VIP)-like peptide, using immunocytochemistry and radioimmunoassay. PACAP-immunoreactive nerve fibers were numerous in the longitudinal and circular muscle layers of sheep and man, moderate in numbers in cat, while being few in the esophagus of guinea-pig. A few PACAP-immunoreactive nerve cell bodies and numerous nerve fibers were seen in the myenteric ganglia of the esophagus of cat, sheep and man. In the lower esophagus of cat, sheep and man all PACAP-containing nerve cell bodies and nerve fibers stored VIP. The results of radioimmunoassay of PACAP in extracts of specimens from man were in good agreement with the immunocytochemical findings. High performance liquid chromatography revealed one major peak of PACAP-like immunoreactivity in extracts of human esophagus. We suggest that neuronal PACAP may serve to modulate motor activity and secretion in the lower esophageal sphincter region.     

Effects of UV-B radiation and nitrogen starvation on enzyme activities in isolated plasma membranes of Euglena gracilis

Circadian rhythms are characteristic of many physiological and biochemical processes in the freshwater flagellate Euglena gracilis. Earlier, we found that the rhythms of photosynthesis, phototaxis and cell shape followed the same pattern in control organisms, but were differently affected by stress such as UV-B irradiation and nitrogen deficiency. Here we extend our studies to use isolated plasma membranes to characterize the rhythms of some plasma membrane-bound enzymes. Also, we wanted to see whether stress-induced changes of these rhythms could be detected at the subcellular level and possibly be coupled to the changes seen in photosynthesis, phototaxis and cell shape. The isolation of plasma membranes using aqueous polymer two-phase partitioning was successful, as judged by the large enrichment of the plasma membrane-marker 5′-nucleotidase, and the difference in the polypeptide pattern compared with the microsomal fraction from which it was prepared. Two other enzymes were analyzed, K⁺, Mg²⁺-ATPase, and adenylyl cyclase. The specific activities of all three enzymes were decreased by UV-B radiation by ca 30–50%, compared with the control cultures. On the other hand, nitrogen deficiency not only reduced the activity of the K⁺.Mg²⁺-ATPase but also increased the activities of the 5′-nucleotidase and adenylyl cyclase. The different treatments also resulted in differences in polypeptide pattern, e.g., a polypeptide around 30 kDa seemed to be specific to plasma membranes of nitrogen-deficient cultures and one at 39 kDa for the UV-B radiated ones. All three enzymes showed diurnal rhythms that were affected by UV-B radiation. The peak in the rhythm of the ATPase was shifted by UV-B radiation, the rhythm of the 5′-nucleotidase nearly eliminated. The first peak of adenylyl cyclase activity was delayed, so that it looked more like a broad peak between 2 and 11 h after the onset of light. The rhythm of ATPase activity could be correlated with that of photosynthesis in both control and UV-B irradiated cultures. Also, the rhythms of adenylyl cyclase activity and cell shape changes showed some similarities.     

Energy Expenditure Compared to Physical Activity Measured by Accelerometry and Self-Report in Adolescents: A Validation Study

Background

Physical inactivity is responsible for 5.3 million deaths annually worldwide. To measure physical activity energy expenditure, the doubly labeled water (DLW) method is the gold standard. However, questionnaires and accelerometry are more widely used. We compared physical activity measured by accelerometer and questionnaire against total (TEE) and physical activity energy expenditure (PAEE) estimated by DLW.

Methods

TEE, PAEE (TEE minus resting energy expenditure) and body composition were measured using the DLW technique in 25 adolescents (16 girls) aged 13 years living in Pelotas, Brazil. Physical activity was assessed using the Actigraph accelerometer and by self-report. Physical activity data from accelerometry and self-report were tested against energy expenditure data derived from the DLW method. Further, tests were done to assess the ability of moderate-to-vigorous intensity physical activity (MVPA) to predict variability in TEE and to what extent adjustment for fat and fat-free mass predicted the variability in TEE.

Results

TEE varied from 1,265 to 4,143 kcal/day. It was positively correlated with physical activity (counts) estimated by accelerometry (rho  = 0.57; p = 0.003) and with minutes per week of physical activity by questionnaire (rho  = 0.41; p = 0.04). An increase of 10 minutes per day in moderate-to-vigorous intensity physical activity (MVPA) relates to an increase in TEE of 141 kcal/day. PAEE was positively correlated with accelerometry (rho  = 0.64; p = 0.007), but not with minutes per week of physical activity estimated by questionnaire (rho  = 0.30; p = 0.15). Physical activity by accelerometry explained 31% of the vssariability in TEE. By incorporating fat and fat-free mass in the model, we were able to explain 58% of the variability in TEE.

Conclusion

Objectively measured physical activity significantly contributes to the explained variance in both TEE and PAEE in Brazilian youth. Independently, body composition also explains variance in TEE, and should ideally be taken into account when using accelerometry to predict energy expenditure values.     

Determinants of Change in Children’s Sedentary Time

Background

Understanding the determinants of sedentary time during childhood contributes to the development of effective intervention programmes.

Purpose

To examine family and home-environmental determinants of 1-year change in objectively measured sedentary time after-school and at the weekend.

Methods

Participants wore accelerometers at baseline and 1 year later. Longitudinal data for after-school and weekend analyses were available for 854 (41.5%male, mean±SD age 10.2±0.3years) and 718 (41.8%male, age 10.2±0.3years) participants. Information on 26 candidate determinants, including socioeconomic status (SES), availability of electronic media and parental rules for sedentary behaviours was self-reported by children or their parents at baseline. Change in the proportion of registered time spent sedentary was used as the outcome variable in multi-level linear regression models, adjusted for age, sex, body mass index and baseline sedentary time. Simple and multiple models were run and interactions with sex explored.

Results

Children from higher socioeconomic status families exhibited greater increases in after-school (beta; 95% CI for change in % time spent sedentary 1.02; 0.37, 1.66) and weekend (1.42; 0.65, 2.18) sedentary time. Smaller increases in after-school sedentary time were observed in children with more siblings (−1.00; −1.69, −0.30), greater availability of electronic media (−0.81; −1.29, −0.33) and, for boys, more frequent family visits to the park (−1.89; −3.28, −0.51) and family participation in sport (−1.28; −2.54, −0.02). Greater maternal weekend screen-time (0.45; 0.08, 0.83) and, in girls, greater parental restriction on playing outside (0.91; 0.08, 1.74) were associated with larger increases in weekend sedentary time. The analytical sample was younger, more likely to be female, had lower BMI and was of higher SES than the original baseline sample.

Conclusions

Intervention strategies aimed at reducing parents’ weekend screen-time, increasing family participation in sports or recreation (boys) and promoting freedom to play outside (girls) may contribute towards preventing the age-related increase in sedentary time.     

Blood oxygen transport in common map turtles during simulated hibernation

We assessed the effects of cold and submergence on blood oxygen transport in common map turtles (Graptemys geographica). Winter animals were acclimated for 6-7 wk to one of three conditions at 3 degrees C: air breathing (AB-3 degrees C), normoxic submergence (NS-3 degrees C), and hypoxic (PO2=49 Torr) submergence (HS-3 degrees C). NS-3 degrees C turtles exhibited a respiratory alkalosis (pH 8.07; PCO2=7.9 Torr; [lactate]=2.2 mM) relative to AB-3 degrees C animals (pH 7.89; PCO2=13.4 Torr; [lactate]=1.1 mM). HS-3 degrees C animals experienced a profound metabolic acidosis (pH 7.30; PCO2=7.9 Torr; [lactate]=81 mM). NS-3 degrees C turtles exhibited an increased blood O2 capacity; however, isoelectric focusing revealed no seasonal changes in the isohemoglobin (isoHb) profile. Blood O2 affinity was significantly increased by cold acclimation; half-saturation pressures (P50's) for air-breathing turtles at 3 degrees and 22 degrees C were 6.5 and 18.8 Torr, respectively. P50's for winter animals submerged in normoxic and hypoxic water were 5.2 and 6.5 Torr, respectively. CO2 Bohr slopes (Delta logP50/Delta pH) were -0.15, -0.16, and -0.07 for AB-3 degrees C, NS-3 degrees C, and HS-3 degrees C turtles, respectively; the corresponding value for AB-22 degrees C was -0.37. The O2 equilibrium curve (O2EC) shape was similar for AB-3 degrees C and NS-3 degrees C turtles; Hill plot n coefficients ranged from 1.8 to 2.0. The O2EC shape for HS-3 degrees C turtles was anomalous, exhibiting high O2 affinity below P50 and a right-shifted segment above half-saturation. We suggest that increases in Hb-O2 affinity and O2 capacity enhance extrapulmonary O2 uptake by turtles overwintering in normoxic water. The anomalous O2EC shape and reduced CO2 Bohr effect of HS-3 degrees C turtles may also promote some aerobic metabolism in hypoxic water.     

Method for spiking soil samples with organic compounds

We examined the harmful side effects on indigenous soil microorganisms of two organic solvents, acetone and dichloromethane, that are normally used for spiking of soil with polycyclic aromatic hydrocarbons for experimental purposes. The solvents were applied in two contamination protocols to either the whole soil sample or 25% of the soil volume, which was subsequently mixed with 75% untreated soil. For dichloromethane, we included a third protocol, which involved application to 80% of the soil volume with or without phenanthrene and introduction of Pseudomonas fluorescens VKI171 SJ132 genetically tagged with luxAB::Tn5. For both solvents, application to the whole sample resulted in severe side effects on both indigenous protozoa and bacteria. Application of dichloromethane to the whole soil volume immediately reduced the number of protozoa to below the detection limit. In one of the soils, the protozoan population was able to recover to the initial level within 2 weeks, in terms of numbers of protozoa; protozoan diversity, however, remained low. In soil spiked with dichloromethane with or without phenanthrene, the introduced P. fluorescens VKI171 SJ132 was able to grow to a density 1,000-fold higher than in control soil, probably due mainly to release of predation from indigenous protozoa. In order to minimize solvent effects on indigenous soil microorganisms when spiking native soil samples with compounds having a low water solubility, we propose a common protocol in which the contaminant dissolved in acetone is added to 25% of the soil sample, followed by evaporation of the solvent and mixing with the remaining 75% of the soil sample.     

TPN-evoked dysfunction of islet lysosomal activity mediates impairment of glucose-stimulated insulin release

We examined the relation between nutrient-stimulated insulin secretion and the islet lysosome acid glucan-1,4-alpha-glucosidase system in rats undergoing total parenteral nutrition (TPN). During TPN treatment, serum glucose was normal, but free fatty acids, triglycerides, and cholesterol were elevated. Islets from TPN-infused rats showed increased basal insulin release, a normal insulin response to cholinergic stimulation but a greatly impaired response when stimulated by glucose or alpha-ketoisocaproic acid. This impairment of glucose-stimulated insulin release was only slightly ameliorated by the carnitine palmitoyltransferase 1 inhibitor etomoxir. However, in parallel with the impaired insulin response to glucose, islets from TPN-infused animals displayed reduced activities of islet lysosomal enzymes including the acid glucan-1,4-alpha-glucosidase, a putative key enzyme in nutrient-stimulated insulin release. By comparison, the same lysosomal enzymes were increased in liver tissue. Furthermore, in intact control islets, the pseudotetrasaccharide acarbose, a selective inhibitor of acid alpha-glucosidehydrolases, dose dependently suppressed islet acid glucan-1,4-alpha-glucosidase and acid alpha-glucosidase activities in parallel with an inhibitory action on glucose-stimulated insulin secretion. By contrast, when incubated with intact TPN islets, acarbose had no effect on either enzyme activity or glucose-induced insulin release. Moreover, when acarbose was added directly to TPN islet homogenates, the dose-response effect on the catalytic activity of the acid alpha-glucosidehydrolases was shifted to the right compared with control homogenates. We suggest that a general dysfunction of the islet lysosomal/vacuolar system and reduced catalytic activities of acid glucan-1,4-alpha-glucosidase and acid alpha-glucosidase may be important defects behind the impairment of the transduction mechanisms for nutrient-stimulated insulin release in islets from TPN-infused rats.     

Sk?ne Emergency Department Assessment of Patient Load (SEAL)—A Model to Estimate Crowding Based on Workload in Swedish Emergency Departments

Objectives

Emergency department (ED) crowding is an increasing problem in many countries. The purpose of this study was to develop a quantitative model that estimates the degree of crowding based on workload in Swedish EDs.

Methods

At five different EDs, the head nurse and physician assessed the workload on a scale from 1 to 6 at randomized time points during a three week period in 2013. Based on these assessments, a regression model was created using data from the computerized patient log system to estimate the level of crowding based on workload. The final model was prospectively validated at the two EDs with the largest census.

Results

Workload assessments and data on 14 variables in the patient log system were collected at 233 time points. The variables Patient hours, Occupancy, Time waiting for the physician and Fraction of high priority (acuity) patients all correlated significantly with the workload assessments. A regression model based on these four variables correlated well with the assessed workload in the initial dataset (r2 = 0.509, p < 0.001) and with the assessments in both EDs during validation (r2 = 0.641; p < 0.001 and r2 = 0.624; p < 0.001).

Conclusions

It is possible to estimate the level of crowding based on workload in Swedish EDs using data from the patient log system. Our model may be applicable to EDs with different sizes and characteristics, and may be used for continuous monitoring of ED workload. Before widespread use, additional validation of the model is needed.