Water permeability of chloroplast envelope membranes. In vivo measurement by saturation-transfer NMR

In tulip tree (Liriodendron tulipifera) leaves, the proton NMR signal from chloroplast water is resolved from that of water in other leaf compartments. We used the saturation-transfer NMR method to measure the mean water molecule residence time within a chloroplast, (88 +/- 17) ms at 20 degrees C. From the measured chloroplast dimensions, we calculate an effective permeability coefficient of (9 +/- 2) X 10(-4) cm/s for the chloroplast envelope membrane. This is the first in vivo measurement of chloroplast water permeability.     

Thermal damage to chloroplast envelope membranes

Nuclear magnetic resonance was used to detect thermal injury to chloroplasts in vivo. A lesion occurs in the chloroplast envelope membrane at temperatures between 53°C and 57°C, depending on species, leaf condition, and heating rate. The injury is associated with a sudden loss of water from the chloroplast.     

Water is allocated differently to chloroplasts in sun and shade leaves

Hydrogen-1 nuclear magnetic resonance spectroscopy was used to study water allocation in cell compartments of sun and shade leaves. NMR spectra of Acer platanoides were resolved into two peaks that were assigned to chloroplast and nonchloroplast water. Sun leaves contained 1.7 times more water per unit area of surface than shade leaves, and the water was allocated differently. Chloroplasts in sun leaves contained 17% of the total leaf water versus 47% in shade leaves. Comparing equal leaf surface areas, the chloroplasts in shade leaves contained 60% more water than those in sun leaves.     

Hepatic 6-phosphofructo-2-kinase/fructose-2,6-bisphosphatase: phosphate dependence and effects of other oxyanions

The effects of various oxyanions on the activities of rat liver 6-phosphofructo-2-kinase/fructose-2,6-bisphosphatase (EC 2.7.1.105/3.1.3.46) were examined. No evidence could be found for an absolute dependence of the kinase activity on inorganic phosphate as was recently reported by M. Laloux, E. Van Schaftingen, and H.-G. Hers ((1985) Eur. J. Biochem. 148, 155-159). Rather, phosphate and arsenate activated the kinase by decreasing the enzyme's Km for fructose 6-phosphate without affecting its Km for ATP or Vmax. The Km of the kinase for fructose 6-phosphate in the presence of inorganic phosphate was found to be significantly lower (6 microM) than previously reported (30 microM) when the hydrolysis of fructose 2,6-bisphosphate by the concomitant bisphosphatase activity at low Fru 6-P concentrations was taken into account. The KA's for phosphate and arsenate activation of the kinase were 0.2 and 0.3 mM, respectively. A number of other oxyanions, including pyrophosphate, sulfate, tungstate, selenate, and molybdate all inhibited the kinase by increasing the Km for fructose 6-phosphate. The apparent Ki's for inhibition of the kinase were in the 0.5-1 mM range. In contrast, all of these oxyanions activated the bisphosphatase, with half-maximal effects requiring millimolar concentrations. Inorganic phosphate was the most potent activator with a KA of 1 mM. In contrast to the other oxyanions, vanadate and meta-periodate inhibited the kinase but had no effect on the bisphosphatase. Vanadate appeared to be a noncompetitive inhibitor since its effects were not overcome by Pi, ATP, or fructose 6-phosphate, and the species responsible was shown to be decavanadate. Like vanadate, meta-periodate had no effect on the bisphosphatase, though it was a potent inhibitor (I0.5 = 30 microM) of the kinase. Its effects were shown to be time-dependent and reversed by dithiothreitol, suggesting that it acted by an oxidative mechanism. These results augment the mounting body of evidence that the enzyme's two reactions are catalyzed at discrete active sites.     

Combined effects of light and water stress on chloroplast volume regulation.

A nuclear magnetic resonance technique was used to measure changes in the water content of Acer platanoides chloroplasts in leaf discs that had reached osmotic equilibrium with external solutions either in the dark or under exposure to light. Results showed that chloroplast volume regulation (CVR) maintained constant water content in the chloroplasts over a range of water potentials in the dark, but CVR failed when the water potential fell below a critical value. The critical potential was lower in the dark in sun leaves than in shade leaves. Upon exposure to intense light, CVR remained effective in sun leaves over the same range as in the dark, but it failed in shade leaves at all water potentials. Osmolytes are necessary for CVR, but KCl is relatively ineffective; increased concentrations of intracellular KCl did not fully support an increase in the range of CVR. The results indicate that leaves need reserve supplies of cytosolic osmolytes to maintain CVR at low water potentials, and a larger reserve supply is needed in leaves that are exposed to intense light.     

Circulating 1α,25-dihydroxyvitamin D in the chicken: Enhancement by injection of prolactin and during egg laying

In order to investigate possible modulation of vitamin D metabolism by prolactin, circulating 1α,25-dihydroxyvitamin D (1α,25-(OH)₂D) was measured by radioreceptor assay in chicks given injections of prolactin for five days. At a dose of 100 μg/day, the lactogenic hormone elicited a two-fold increase in plasma 1α,25-(OH)₂D. This effect may explain the known action of prolactin in producing hypercalcemia and could be physiologically important in birds. The laying hen represents a physiologic state in which calcuim absorption is known to be stimulated and prolactin has been reported to be elevated. Assay of serum 1α,25-(OH)₂D in the laying hen demonstrates a nine-fold enhancement over non-laying controls. Since this marked increase during egg laying is at least partially mimicked by injecting prolactin, a possible causative relationship between elevated prolactin and 1α,25-(OH)₂D is suggested.     

Tools,techniques, and future opportunities for characterizing the mechanobiology of uterine myometrium

The myometrium is the smooth muscle layer of the uterus that generates the contractions that drive processes such as menstruation and childbirth. Aberrant contractions of the myometrium can result in preterm birth, insufficient progression of labor, or other difficulties that can lead to maternal or fetal complications or even death. To investigate the underlying mechanisms of these conditions, the most common model systems have conventionally been animal models and human tissue strips, which have limitations mostly related to relevance and scalability, respectively. Myometrial smooth muscle cells have also been isolated from patient biopsies and cultured in vitro as a more controlled experimental system. However, in vitro approaches have focused primarily on measuring the effects of biochemical stimuli and neglected biomechanical stimuli, despite the extensive evidence indicating that remodeling of tissue rigidity or excessive strain is associated with uterine disorders. In this review, we first describe the existing approaches for modeling human myometrium with animal models and human tissue strips and compare their advantages and disadvantages. Next, we introduce existing in vitro techniques and assays for assessing contractility and summarize their applications in elucidating the role of biochemical or biomechanical stimuli on human myometrium. Finally, we conclude by proposing the translation of “organ on chip” approaches to myometrial smooth muscle cells as new paradigms for establishing their fundamental mechanobiology and to serve as next-generation platforms for drug development.     

Midpoint attractors and species richness: Modelling the interaction between environmental drivers and geometric constraints

We introduce a novel framework for conceptualising, quantifying and unifying discordant patterns of species richness along geographical gradients. While not itself explicitly mechanistic, this approach offers a path towards understanding mechanisms. In this study, we focused on the diverse patterns of species richness on mountainsides. We conjectured that elevational range midpoints of species may be drawn towards a single midpoint attractor – a unimodal gradient of environmental favourability. The midpoint attractor interacts with geometric constraints imposed by sea level and the mountaintop to produce taxon‐specific patterns of species richness. We developed a Bayesian simulation model to estimate the location and strength of the midpoint attractor from species occurrence data sampled along mountainsides. We also constructed midpoint predictor models to test whether environmental variables could directly account for the observed patterns of species range midpoints. We challenged these models with 16 elevational data sets, comprising 4500 species of insects, vertebrates and plants. The midpoint predictor models generally failed to predict the pattern of species midpoints. In contrast, the midpoint attractor model closely reproduced empirical spatial patterns of species richness and range midpoints. Gradients of environmental favourability, subject to geometric constraints, may parsimoniously account for elevational and other patterns of species richness.     

A Randomized Cross-over Exploratory Study of the Effect of Visiting Therapy Dogs on College Student Stress Before Final Exams

This exploratory study investigated the effect of visiting therapy dogs on college-student perceived and physiological stress the week prior to final exams. Students (n = 78) were randomly assigned to order of a therapydog intervention and attention-control condition, each 15 minutes long. Students completed the Perceived Stress Scale (PSS), a stress visual analog scale (SVAS), and provided saliva for measuring nerve growth factor (sNGF) and alpha amylase (sAA), prior to randomization. Saliva samples and SVAS were again collected after each condition. There was no effect of group order on demographics, PSS, or initial SVAS. Repeated measures models were used to analyze the complete data sets of 57 students. There were no significant differences in sAA between or within students completing the intervention and control conditions. sNGF was not subjected to analysis as most levels were undetectable. Significant differences in SVAS scores were found between the intervention and control condition, with large effect sizes. SVAS scores were lower following the intervention, regardless of condition order (intervention first, p = 0.0001, d = 1.87; intervention second, p = 0.0004, d = 1.63). No SVAS differences were found for the control condition. Based on these findings, campus events with visiting therapy dogs represent a costeffective, easily accessible activity to reduce perceived, but not physiological, stress for college students prior to final exams.