An in vivo experimental system to study sugar phloem unloading in ripening grape berries during water deficiency stress

An in vivo experimental system-called the 'berry-cup' technique-was developed to study sugar phloem unloading and the accumulation of sugar in ripening grape berries. The berry-cup system consists of a single peeled grape berry immersed in a buffer solution in a cup prepared from a polypropylene syringe. A small cross-incision (2 mm in length) is made on the stylar remnant of a berry during its ripening phase, the skin of the berry then being easily peeled off, exposing the dorsal vascular bundles without damaging either these or the pulp tissue of the berry. The sites of sugar phloem unloading are thus made directly accessible and may be regulated by the buffer solution. In addition, the unloaded photoassimilates are easily transported into the buffer solution in the berry-cup. With the berry-cup technique, it takes 60 min to purge the sugar already present in the apoplast, after which the amount of sugar in the buffer solution is a direct measure of the sugar unloading from the grape berry phloem. The optimum times for sampling were 20 or 30 min, depending on the type of experiment. Sugar phloem unloading was significantly inhibited by the inclusion of either 7.5 mm NaF or 2.5 mm PCMB in the buffer solution. This study indicates that sugar phloem unloading in ripening grape berries is via the apoplastic network and that the process requires the input of energy. The system was shown to be an appropriate experimental system with which to study sugar phloem unloading in ripening grape berries, and was applied successfully to the study of berry sugar unloaded from grapevines subjected to water stress. The results showed that water deficiency inhibits sugar unloading in grape berries.     

Two types of HKT transporters with different properties of Na+ and K+ transport in Oryza sativa

It is thought that Na+ and K+ homeostasis is crucial for salt-tolerance in plants. To better understand the Na+ and K+ homeostasis in important crop rice (Oryza sativa L.), a cDNA homologous to the wheat HKT1 encoding K+-Na+ symporter was isolated from japonica rice, cv Nipponbare (Ni-OsHKT1). We also isolated two cDNAs homologous to Ni-OsHKT1 from salt-tolerant indica rice, cv Pokkali (Po-OsHKT1, Po-OsHKT2). The predicted amino acid sequence of Ni-OsHKT1 shares 100% identity with Po-OsHKT1 and 91% identity with Po-OsHKT2, and they are 66-67% identical to wheat HKT1. Low-K+ conditions (less than 3 mM) induced the expression of all three OsHKT genes in roots, but mRNA accumulation was inhibited by the presence of 30 mM Na+. We further characterized the ion-transport properties of OsHKT1 and OsHKT2 using an expression system in the heterologous cells, yeast and Xenopus oocytes. OsHKT2 was capable of completely rescuing a K+-uptake deficiency mutation in yeast, whereas OsHKT1 was not under K+-limiting conditions. When OsHKTs were expressed in Na+-sensitive yeast, OsHKT1 rendered the cells more Na+-sensitive than did OsHKT2 in high NaCl conditions. The electrophysiological experiments for OsHKT1 expressed in Xenopus oocytes revealed that external Na+, but not K+, shifted the reversal potential toward depolarization. In contrast, for OsHKT2 either Na+ or K+ in the external solution shifted the reversal potential toward depolarization under the mixed Na+ and K+ containing solutions. These results suggest that two isoforms of HKT transporters, a Na+ transporter (OsHKT1) and a Na+- and K+-coupled transporter (OsHKT2), may act harmoniously in the salt tolerant indica rice.     

硫酸镁对大鼠海马CA1区神经元钠电流的抑制作用

利用全细胞膜片钳技术研究了硫酸镁 (MgSO4 )对大鼠海马CA1区神经元钠电流的影响。结果表明 ,MgSO4 可浓度依赖和电压依赖地抑制钠电流 ,半数抑制浓度为 4 0 5mmol/L。这一抑制作用与刺激频率无关。结果还表明 ,4mmol/LMgSO4 不影响钠电流的失活过程 ,却使半数激活电压由 - 5 5 8± 6 8mV变为 - 3 4 2± 6 2mV (n =8,P <0 0 1) ,而激活曲线的斜率因子不变。结果提示 ,MgSO4 抑制大鼠海马CA1区神经元的钠电流可能是其抗缺血缺氧造成的中枢神经系统损伤的机制之一     

Prospects for using genetic transformation for improved SIT and new biocontrol methods

The genetic manipulation of non-drosophilid insect species is possible by the creation of recombinant DNA constructs that can be integrated into host genomes by several transposon-based vector systems. This technology will allow the development and testing of a variety of systems that can improve existing biological control methods, and the development of new highly efficient methods. For programs such as sterile insect technique (SIT), transgenic strains may include fluorescent protein marker genes for detection of released insects, and conditional gene expression systems that will result in male sterility and female lethality for genetic sexing. Conditional expression systems include the yeast GAL4 system and the bacterial Tet-off and Tet-on systems that can, respectively, negatively or positively regulate expression of genes for lethality or sterility depending on a dietary source of tetracycline. Importantly, strains for male sterility must also incorporate an effective system for genetic sexing, since typically, surviving females would remain fertile. Models for the use of these expression systems and associated genetic material come from studies in Drosophila and, while many of these systems should be transferable to other insects, continued research will be necessary in insects of interest to clone genes, optimize germ-line transformation, and perform vector stability studies and risk assessment for their release as transgenic strains.     

Preparation and characterization of flurbiprofen beads by melt solidification technique

Amelt solidification technique has been developed to obtain sustained-release waxy beads of flurbiprofen. Low glass transition temperature (t _g) and shear-induced crystallization of flurbiprofen made it a suitable candidate for melt solidification technique. The process involved emulsification and solidification of flurbiprofen-cetyl alcohol melt at significantly low temperature (5°C). The effect of variables, namely, the amount of cetyl alcohol and the speed of agitation, was studied using 3² factorial design. The technique and the beads were evaluated on the basis of process and desired yield, surface topography, Fourier-transform infrared (FT-IR), differential scanning calorimetry (DSC), particle size distribution, crushing strength, and drug release. Average values for process and desired yields were 97% wt/wt and 26% wt/wt, respectively. No interaction was observed between drug and excipient. Multiple regression analysis was carried out, and response surfaces were obtained. A curvilinear relationship was observed between percentage of desired yield and the amount of cetyl alcohol. Linear decrease in crushing strength was observed with increase in the amount of cetyl alcohol. Drug released from the beads followed zero order kinetics. Burst release was shown to a greater extent in beads containing a lower amount of cetyl alcohol. Response surfaces of time required for certain percentage of drug (t _D%) showed that after critical concentration of about 20% of cetyl alcohol (400 mg/batch), no significant release retardant effect was observed.     

Effects of mixtures of lauric and myristic acid on rumen methanogens and methanogenesis in vitro

AIMS: To identify the most effective mixture of non-esterified lauric (C12) and myristic (C14) acid in suppressing ruminal methanogenesis, and to investigate their effects on the methanogenic population. METHODS AND RESULTS: C12/C14 mixtures were incubated with rumen fluid using the Hohenheim gas test apparatus. Methane production and the numbers of Archaea declined with an increasing proportion of C12. With a 2 : 1 proportion of C12/C14, the maximum methane-suppressing effect (96%) was achieved similar to that with C12 alone. The proportions of the individual methanogenic orders of total methanogens were altered by varying the C12/C14 ratio. CONCLUSIONS: Although C14 alone had no effect on methanogenesis, C14 enhanced the methane-suppressing effect of C12 in certain mixtures. SIGNIFICANCE AND IMPACT OF THE STUDY: The results support strategies for an environment-friendly ruminant nutrition as it was demonstrated that part of the less palatable C12 could be replaced by C14 without losing its methane-suppressing potential.     

Comparison of sample collection methods for the PCR detection of oral anaerobic pathogens

AIMS: To provide evidence that DNA-PCR diagnostics of oral pathogens based on standard sample collection by paper point insertion from the depth of the periodontal pocket can be replaced by a novel non-invasive collection method based on swab technique from the gingiva. METHODS AND RESULTS: In this study we compared the results from two collection methods performed in 35 patients with chronic adult periodontitis. Statistical analysis showed a highly significant association of diagnostic results between both collection techniques. CONCLUSIONS: The Pocket-out method represents a reliable alternative to the standard collection technique for PCR diagnosis of oral pathogens. SIGNIFICANCE AND IMPACT OF THE STUDY: Due to its simplicity and non-invasiveness, the Pocket-out collection could be performed in any physician office, or even by the patient himself. With respect to the putative association between periodontal disease and various systemic illnesses, this method could be integrated with various screening programs of oral pathogens.     

Effect of Dicyclohexylcarbodiimide (DCCD) on Transport Parameters in the Frog Cornea Epithelium

Dicyclohexylcarbodiimide (DCCD) is a carboxyl group modifier and it is an inhibitor of various ATPases. Present experiments, using an in vitro preparation, were designed to study whether DCCD affected the transporters of the bullfrog cornea epithelium, specifically, the Na⁺/K⁺ ATPase pump located in the basolateral membrane. For this purpose, corneas were impaled with microelectrodes and experiments were done under short-circuit current (I _sc ) conditions. Addition of DCCD to a concentration of 10⁻⁴ m to the tear solution gave a marked decrease in I _sc ; a marked depolarization of the intracellular potential, V _o ; and a significant decrease in the apical membrane fractional resistance, fR _o . There were small and variable although significant changes in the transepithelial conductance, g _t . The effects may be explained by a decrease in the basolateral membrane K⁺ conductance, in combination with a partial inhibition of the Na⁺/K⁺-ATPase pump located in the basolateral membrane. There is also evidence for an increase in the apical membrane Cl⁻ conductance. Received: 12 August 1999/Revised: 16 November 1999     

A small-scale survey on the status of intestinal parasite infections in rural villages in Nepal

The status of intestinal parasite infections was investigated in two rural villages (Chitrasar, Jerona) in Chitwan District, Nepal in 1999. Stool examination was performed with a total of 300 specimens from schoolchildren by formalin-ether sedimentation technique. The prevalence rate of intestinal parasite infections in the surveyed areas was 44.0%. The prevalence rate in Jerona was slightly higher than that in Chitrasar. The prevalence rate of intestinal parasite infections in female was slightly higher than that in male without statistically significant difference. Entamoeba coli was the most commonly found protozoan parasite (21.0%) followed by Giardia lamblia (13.7%) and others (5.3%). Hookworm was the most prevalent intestinal helminth (13.0%) followed by Trichuris trichiura (3.0%) and others (5.0%). Forty-three specimens (14.3%) showed mixed infections. It is necessary to implement large-scale treatment with anthelminthics, health education and sanitary improvement for intestinal parasite control in the surveyed areas.     

Stimulation of Neuropeptide Y Overflow in the Rat Paraventricular Nucleus by Corticotropin-Releasing Factor

Abstract: Neuropeptide Y (NPY) and corticotropin-releasing factor (CRF) are present at high concentrations in the hypothalamus where they mediate important endocrine and autonomic functions. Morphological and physiological studies have suggested an interaction between these peptides, and opposing actions of CRF and NPY have been reported on feeding and other behaviors. This study investigated the effect of CRF on NPY release in vivo, measured by push-pull techniques, in the anesthetized rat. Push-pull probes implanted into the paraventricular nucleus of the hypothalamus (PVN) were perfused with modified Ringer solution containing bovine serum albumin at 15 µl/min, and the perfusate was lyophilized prior to NPY radioimmunoassay. NPY overflow from the rat PVN was increased threefold by perfusion of a depolarizing concentration of potassium (50 mmol/L KCI). When CRF was administered into the PVN via the push-pull cannula at 1 or 5 µg/ml, dose-dependent increases in NPY overflow of two- and fivefold were observed ( p < 0.05). These increases were abolished by prior intracerebroventricular (i.c.v.) administration of the CRF antagonist [ d -Phe¹²,Nle^21,38,C^αMeLeu³²]CRF (12–41) at 1 or 5 µg/µl, respectively. NPY overflow returned promptly to resting levels following CRF administration. In contrast, when CRF was administered by i.c.v. bolus at a similar total dose (2 µg), no significant effect on NPY overflow was observed. These data provide in vivo evidence for an interaction between CRF and NPY at the level of the PVN.