Intracellular pH of bovine sperm increases during capacitation

The effect of heparin-induced capacitation on the intracellular pH (pH_i) of individual bovine sperm was determined with image analysis. Sperm were loaded with the acetoxymethyl ester of the pH sensitive fluorescent indicator, 2′,7′-bis(carboxyethyl)-5(6)-carboxy-fluorescein (BCECF). The pH_i of 5303 sperm was evaluated from a total of five bulls at .5, 2, 3, 4, and 5 h of incubation. The pH_i did not differ between the sperm head and mid-piece (P > 0.05). An increase in sperm head pH_i was seen in heparin-treated sperm at 3, 4, and 5 h of incubation relative to sperm incubated without heparin (control, P < 0.05). At 5 h of incubation, the pH_i in heparin-treated sperm was 6.92 ± 0.07, while control-treated sperm pH_i was 6.70 ± 0.03. Initially a normal frequency distribution was seen for sperm pH_i in both heparin- and control-treated sperm. As the incubation progressed, the frequency distribution began to skew towards higher pH_i in both samples but was more dispersed for the heparin-treated sperm. Following an NH₄Cl-induced alkaline load, the pH_i of both control- and heparin-treated sperm recovered toward the resting pH_i with a half-time of recovery of 1.5–1.7 min. The recovery of sperm pH_i was not due to leakage of NH⁴⁺ into sperm because recovery also occurred with trimethylamine. The instantaneous velocity of the pH_i recovery (v_i) was dependent on pH_i and decreased as pH_i decreased. Capacitation by heparin was associated with an 81% decrease in v_i at a pH_i of 7.00, but there was no effect of capacitation on the proton buffering power of the sperm, which was 87 ± 8 mM/pH unit. Results demonstrate that both the regulation of pH_i and resting pH_i were altered during capacitation of bovine sperm by heparin. © 1995 Wiley-Liss, Inc.     

Applying climate change refugia to forest management and old-growth restoration

Recent studies highlight the potential of climate change refugia (CCR) to support the persistence of biodiversity in regions that may otherwise become unsuitable with climate change. However, a key challenge in using CCR for climate resilient management lies in how CCR may intersect with existing forest management strategies, and subsequently influence how landscapes buffer species from negative impacts of warming climate. We address this challenge in temperate coastal forests of the Pacific Northwestern United States, where declines in the extent of late-successional forests have prompted efforts to restore old-growth forest structure. One common approach for doing so involves selectively thinning forest stands to enhance structural complexity. However, dense canopy is a key forest feature moderating understory microclimate and potentially buffering organisms from climate change impacts, raising the possibility that approaches for managing forests for old-growth structure may reduce the extent and number of CCR. We used remotely sensed vegetation indices to identify CCR in an experimental forest with control and thinned (restoration) treatments, and explored the influence of biophysical variables on buffering capacity. We found that remotely sensed vegetation indices commonly used to identify CCR were associated with understory temperature and plant community composition, and thus captured aspects of landscape buffering that might instill climate resilience and be of interest to management. We then examined the interaction between current restoration strategies and CCR, and found that selective thinning for promoting old-growth structure had only very minor, if any, effects on climatic buffering. In all, our study demonstrates that forest management approaches aimed at restoring old-growth structure through targeted thinning do not greatly decrease buffering capacity, despite a known link between dense canopy and CCR. More broadly, this study illustrates the value of using remote sensing approaches to identify CCR, facilitating the integration of climate change adaptation with other forest management approaches.     

Cerebral Acid Buffering Capacity at Different Ages Measured In Vivo by 31P and 1H Nuclear Magnetic Resonance Spectroscopy

Cerebral acidosis occurring during ischemia has been proposed as one determinant of tissue damage. Newborn animals appear to be less susceptible to ischemic tissue damage than adults. One possible component of ischemic tolerance could derive from maturational differences in the extent of acid production and buffering in newborns compared to adults. The purpose of this study was to measure the dependency of acid production on the blood plasma glucose concentrations and acid buffering capacity of piglets at different stages of development. Complete ischemia was induced in 29 piglets ranging in postconceptual age from 111 to 156 days (normal term conception, 115 days). Brain buffering capacity during the first 30 min of ischemia was quantified in vivo, via 31P and 1H nuclear magnetic resonance (NMR) spectroscopy, by measuring the change in intracellular brain pH for a given change in the concentration of compounds that contribute to the production of hydrogen ions. Animals from all four age groups showed a similar linear correlation between preischemia blood glucose concentration and intracellular pH after 30 min of ischemia. For each animal the slope of the plot of intracellular pH versus cerebral buffer base deficit was used to calculate the buffer capacity. Using data obtained over the entire 30 min of ischemia, there was no difference in the mean buffer capacity of the different age groups, nor was there a significant correlation between buffer capacity and age. However, there was a significant increase in buffer capacity for the intracellular pH range 6.6-6.0, compared to 7.0-6.6, for all age groups. No significant differences in buffer capacity for these two pH ranges were observed between any of the age groups. Acid buffering capacity was also measured by performing pH titrations on brain tissue homogenized in the presence of inhibitors of glycolysis and creatine kinase. Plots of homogenate pH versus buffer base deficit showed a nonlinear trend similar to that seen in vivo, indicating an increase in buffer capacity as intracellular pH decreases. A comparison of newborn and 1-month-old brain tissue frozen under control conditions or after 45 min of ischemia revealed no differences that could be attributed to age and a slight decrease in buffer capacity of ischemic brain compared to control brain tissue homogenates. There was no difference between the brain buffering capacity measured in vivo using 31P and 1H NMR and that measured in vitro using brain homogenates.     

Responses of exchangeable base cations to continuously increasing nitrogen addition in alpine steppe: A case study of Stipa purpurea steppe

土壤交换性盐基离子(Ca ²⁺、Mg ²⁺、K ⁺、Na ⁺)在维持土壤养分与缓冲土壤酸化中起着重要作用, 了解其对氮添加的响应有助于准确评估氮沉降背景下生态系统结构与功能的动态变化。然而, 目前关于土壤交换性盐基离子对氮添加响应的相关研究主要集中在酸性土中。鉴于目前在碱性土中研究相对较少的现状, 该研究以青藏高原高寒草原为研究对象, 依托氮添加控制实验平台, 通过连续3年(2014-2016)的测定, 考察了8个不同施氮水平(0、1、2、4、8、16、24、32 g·m ^-2·a ^-1)下土壤交换性盐基离子含量变化及其可能原因。结果显示: 随着施氮量的增加, 土壤交换性盐基离子, 尤其是Mg ²⁺与Na ⁺含量显著降低。并且, 盐基离子含量与植物地上生物量显著负相关(p < 0.05), 说明氮添加通过促进植物生长, 加速植物对盐基离子的吸收, 进而导致土壤中盐基离子含量降低。此外, 盐基离子含量也与土壤无机氮含量呈显著负相关(p < 0.05)关系, 说明施氮还通过提高土壤中无机氮含量进而导致更多NH₄ ⁺与土壤吸附的盐基离子交换, 同时加剧NO₃ ^-淋溶, 带走等电荷阳离子。需要指出的是, 虽然连续施氮导致土壤pH值下降, 但该土壤目前仍处于碳酸盐缓冲阶段, 说明通常在酸性土中报道的“因缓冲土壤酸化引起的盐基离子损失机制”在碱性土中并不成立。这些结果意味着持续的氮输入会造成碱性土中盐基离子损失, 进而影响土壤缓冲能力与植被生产力, 未来草原生态系统管理中应重视这一问题。     

Buffering effect of a prophylactic gel on dental plaque in institutionalised elderly

Objectives: The effect of multiple daily applications of a prophylactic gel, with buffering substances, on plaque acidogenicity in elderly institutionalised individuals was evaluated. Background: Many elderly suffer from reduced salivary flow, poor oral hygiene and increased levels of cariogenic bacteria and are considered to be at an increased risk for coronal and root caries. Reinforcing the buffering capacity of dental plaque by the addition of substances such as bicarbonate and phosphates may decrease their caries activity. Materials and methods: Fourteen elderly, with subjective dry mouth, were treated for 16‐day‐periods at random with: (i) Profylin fluoride gel with buffering components; (ii) Profylin fluoride gel without buffering components and (iii) rinsing with water. Applications were made four times a day and each period was followed by a 2‐week wash‐out period. The plaque pH was registered after a carbohydrate challenge and the following were recorded before and after each test period: stimulated salivary secretion rate, buffer capacity, number Colony Farming Units (CFU) mutans streptococci, lactobacilli and a sample of Candida albicans on oral mucosa. Results: Eleven participants (mean age 76.6 years) fulfilled the study. Changes in plaque pH measurements, when calculated as area under the curve (AUC_6.2 and AUC_5.7) values (pH × min), before and after each of the three treatments, showed no significant differences. A tendency to a higher plaque acidogenicity and amount of cariogenic microorganisms was found after the gel treatments. C. albicans was found in low levels. Conclusion: Frequent applications of the gel did not result in an improved neutralising effect in the elderly. This may be caused by a combination of several factors, such as the level of oral dryness of the individuals and low solubility, release and retention of the gel substances in plaque. Instead, an increased plaque acidogenicity was noted.     

A comparative study of four rumen buffering agents on productive performance,rumen fermentation and meat quality in growing lambs fed a total mixed ration

Controlling rumen fermentation using buffering agents could contribute to enhancing ruminant productivity and performance. This study was realized to investigate the effect of dietary supplementation of AcidBuf, sodium bicarbonate, calseapowder and WMC seaweed (Utva Lactuca extra) on the animal performance, volatile fatty acids, rumen pH, rumen histology and carcass characteristics of growing male Awassi lambs. A total of 60 lambs was divided into five groups. One group served as a control and fed only on a concentrate diet without any buffering, whereas the other four groups were fed the concentrate diet supplemented with 0.4% AcidBuf (Buf1), 0.4% AcidBuf plus sodium bicarbonate, 50 : 50 (Buf2), 0.4% calseapowder (Buf3) or 0.4% WMC Seaweed (Buf4) for 98 days. The feed conversion ratio was (P<0.05) improved in Buf2 compared to the control and other treatment groups. The propionic acid decreased, whereas butyric acid was increased in the treatment groups (P<0.05) compared to the control. The pH of the rumen fluid and the length of submucosa were (P<0.05) higher in Buf4 and Buf1, respectively, compared to the control. Hot and cold carcass weights were (P<0.05) higher in Buf4 compared to Buf1. Lean meat percentage and rib eye area were (P<0.05) higher in Buf4; while the fat percentage was (P<0.05) lower in Buf2 and Buf4 groups compared to the control. The lightness and yellowness of meat were (P<0.05) higher in Buf1 and Buf4 compared to the control. The meat pH was (P<0.05) higher in Buf3 and Buf4 compared to Buf2 (at 1 h) and control (at 24 h). The visceral depot fat (%) was reduced with Buf3 and Buf4 compared to the control. The results indicated that dietary supplementation of different buffering agents improved feed efficiency, rumen pH, carcass characteristics and decreased the body fat in growing Awassi lambs.     

Evolutionary and ecological aspects of early brain malnutrition in humans

This article reviews the effects of malnutrition on early brain development using data generated from animal experiments and human clinical studies. Three related processes, each with their own functional consequences, are implicated in the alteration of brain development. (1) Maternal undernutrition at the start of pregnancy results in reduced transfer of nutrients across the placenta, allowing the conservation of effort for future reproductive episodes. (2) Differential allocation to growing organs by the fetus in response to nutritional stress spares the brain to a large though still limited degree, reflecting the organ’s relative contribution to survival and reproductive success. (3) Prenatal malnutrition disrupts developing neurotransmitter systems, which results in the expression of specific cognitive and affective traits. It is argued that the increasing size and therefore cost of the brain, in conjunction with increasing ecological instability and marginality, reinforced selection for maternally controlled growth suppression of offspring, reallocation of organ growth rates by offspring, and behavioral changes related to development of neurotransmitter systems. William D. Lukas is a graduate student in the Department of Anthropology at Boston University. He is currently studying the relationship of hormones to immune function and body composition in different populations of African men. Benjamin C. Campbell is Assistant Professor of Anthropology at Boston University. He studies human reproductive ecology, the endocrinology of male fertility and health over the life span, and biocultural interactions in African populations.     

Factors controlling equilibrium boron (B) concentration in nutrient solution buffered with B-specific resin (Amberlite IRA-743)

In conventional solution culture where boron (B) is added as boric acid, fluctuating external B supply often produces confounding and ill-defined physiological and biochemical responses in plants, especially when grown at deficient and marginal B supply. Our previous studies proposed the use of the B-specific resin – Amberlite IRA-743 to develop a B-buffered solution culture. The present study aims to evaluate crucial factors determining equilibrium B concentrations in nutrient solution buffered with the B-loaded resin, including the B loading of the resin, pH in the nutrient solution and B removal from the solution. The equilibrium B concentrations in nutrient solution were determined by both the amount of B sorbed by the resin and the solution pH. At pH 6.05±0.05, the relationship between the resin B content and equilibrium B concentration in the nutrient solution is closely described by the equation: Y = 18.8 X^1.457 [ where, Y = equilibrium B concentration (μM) in nutrient solution and X = B content of the resin (mg B g⁻¹ moist resin)]. However, at a given resin B content, lowering solution pH from 7 to 4 significantly increased B concentrations in solution through the release of B from the solid phase of the resin beads. The B-loaded resin was capable of maintaining stable B concentrations in the nutrient solutions, ranging from deficient to marginally adequate B concentrations for dicot species. In conclusion, B concentrations ranging from 0.05 to 11 μM, were buffered for 5 days with the resin loaded with 0.004 – 0.691 mg B g⁻¹ moist resin in the nutrient solution. Precise pH control in the nutrient solution is critical for the success of a B-buffered solution culture study. This revised version was published online in June 2006 with corrections to the Cover Date.     

Electrophysiological evidence suggests a defective Ca2+ control mechanism in a newParamecium mutant

Summary A new mutant ofParamecium tetraurelia, k-shyA, was characterized behaviorally and electrophysiologically. The mutant cell exhibited prolonged backward swimming episodes in response to depolarizing conditions. Electrophysiological comparison of k-shyA with wild type cells under voltage clamp revealed that the properties of three Ca²⁺-regulated currents were altered in the mutant. (i) The voltage-dependent Ca²⁺ current recovered from Ca²⁺-dependent inactivation two- to 10-fold more slowly than wild type. Ca²⁺ current amplitudes were also reduced in the mutant, but could be restored by EGTA injection. (ii) The decay of the Ca²⁺-dependent K⁺ tail current was slower in the mutant. (iii) The decay of the Ca²⁺-dependent Na⁺ tail current was also slower in the mutant. All other membrane properties studied, including the resting membrane potential and resistance and the voltage-sensitive K⁺ currents, were normal in k-shyA. Considered together, these observations are consistent with a defect in the ability of k-shyA to reduce the free intracellular Ca²⁺ concentration following stimulation. The possible targets of the genetic lesion and alternative explanations are discussed. The k-shy mutants may provide a useful tool for molecular and physiological analyses of the regulation of Ca²⁺ metabolism inParamecium.     

Crystallization and liquid‐liquid phase separation of monoclonal antibodies and fc‐fusion proteins: Screening results

Crystallization holds the potential to be used for protein purification and low‐viscosity drug substance and drug product formulations. Twenty‐two different proteins (20 monoclonal antibodies and two Fc‐fusions) were examined to determine the breadth of applicability of crystallization to these therapeutic proteins. Vapor diffusion technique and an evaporative screening method were used to identify crystallization conditions using around a 100 initial conditions based on reagents that are generally regarded as safe (GRAS). Of 16 IgG₂s examined, at least four formed diffraction‐quality crystals and four others formed crystal‐like particles. At least three of the IgG₂s that crystallized well were also crystallized under the same set of operating conditions using inexpensive GRAS reagents. The crystals were formed to high‐yields in a few hours and were dissolved quickly without impacting product quality. Although only a fraction of the proteins examined crystallized, all exhibited liquid‐liquid phase separation (LLPS), which could be used for their concentration or possibly purification. One of the Fc‐fusions, for example, was concentrated by LLPS to a self‐buffering solution at 150 g/L. Crystallization and LLPS in the salting‐in region were shown to be feasible. © 2011 American Institute of Chemical Engineers Biotechnol. Prog., 2011