Effect of amiloride on electrolyte concentrations and rubidium uptake in principal and mitochondria-rich cells of frog skin

The role of mitochondria-rich cells (MR cells) in transepithelial Na transport was investigated by determining electrolyte concentrations and Rb uptake in individual cells of frog skin epithelium using electron microprobe analysis. Measurements were performed under control conditions and after blocking the transepithelial Na transport with amiloride. Under control conditions, Na and Cl concentrations of MR cells scattered much more than those of principal cells and ranged from a few up to more than 30 mmol/kg wet weight. Rb uptake from the basal side into individual MR cells also showed a large variation and was, on the average, much less pronounced than into the principal cells. In principal cells, amiloride reduced the Na concentration and Rb accumulation. In contrast, no effect was observed upon electrolyte concentration and Rb uptake of MR cells. Rb uptake was correlated to the Na concentration of MR cells both under control conditions and after amiloride. It is concluded that, in contrast to the principal cells, MR cells are not involved in amiloride-sensitive transepithelial Na transport and that their Na/K-pump activity is very low.     

The osmotic behaviour of toad skin epithelium (Bufo viridis)

The effect of saline adaptation on the intracellular Na, K, Cl, P concentrations and dry weight content of the toad skin epithelium (Bufo viridis) was studied using the technique of electron microprobe analysis. The measurements were performed on isolated abdominal skins either directly after dissection or after additional incubation in Ussing-type chambers.Adaptations of the toads to increasing NaCl concentrations for 7 days resulted in increased blood plasma osmolarity and a parallel increase in the cellular electrolyte, P and dry weight concentrations of the epithelium, the K increase representing the most significant fraction of the intracellular osmolarity increase. No evidence was obtained to show that the nucleus and cytoplasm reacted differently from each other and all living epithelial cell types basically showed the same response.Incubation of the isolated skins under control conditions showed a drastic inhibition of the transepithelial Na transport after adaptation to high salinities. In spite of the large variations in the transport rate almost identical intracellular electrolyte concentrations were observed. In tap water adapted toads the average cellular concentrations were 8.8 mmole/kg wet weight for Na, 109.6 for K, 41.5 for Cl, and 135.3 for P, respectively. Incubation of the skin with Ringer's solution of different osmolarities demonstrated that the epithelial cells are in osmotic equilibrium with the inner bathing solution. The results are consistent with the view that the osmotic adaptation is mainly accomplished by the movement of water.This work was supported by grants from the Deutsche Forschungsgemeinschaft and the Stiftung Volkswagenwerk     

Chromosome mapping ofSoa,a gene influencing gustatory sensitivity to sucrose octaacetate in mice

Strain distribution patterns among recombinant inbred strains suggested that a locus influencing taste sensitivity to sucrose octaacetate was on chromosome 6. A location forSoa was established by linkage analysis of behavioral and electrophoretic data from outbred and congenic strains and from test-cross progeny. Haplotyping of 41 outbred CFW-Cr animals with a cDNA probe showed perfect cosegregation ofSoa andPrp, a gene for salivary proline-rich proteins. Five of twelve B6. SW-Soa ^a strains were found to retainLdr-1, lactate dehydrogenase regulator-1, on chromosome 6 as an allelic passenger from the SWR/J donor strain (source of theSoa ^a Taster allele). Centimorgan distance was estimated using the ABP/Le linkage-testing strain (non-Taster,Soa ^b) and the SWR/J strain (Taster,Soa ^a) in a testcross breeding system. The data are consistent with a position for theSoa locus on mouse chromosome 6, 62 cM from the centromere.This research was supported in part by Grants DC00150 (G. W.) and DE003658 (E.A.A.). This paper is based on a thesis submitted to the Florida State University by the first author in partial fulfillment of the requirements for the Master of Sciences degree.     

2000年～2004年淋球菌深圳分离株的耐药性变迁

目的了解深圳地区淋球菌流行状况和耐药趋势,为淋病的预防和治疗提供依据.方法琼脂稀释法测定青霉素、四环素、壮观霉素、头孢三嗪和环丙沙星对246株淋球菌的最低抑茵浓度,头孢硝噻吩纸片法检测β-内酰胺酶.结果淋球茵PPNG和TRNG阳性率呈逐年上升趋势;淋球菌对5种常规抗生素的耐药性均有不同程度增强,尤以环丙沙星耐药性增强突出.结论深圳地区淋球菌耐药状况较为严重,大观霉素可作为淋病治疗的首选药物.临床应加强淋球茵耐药性监测并根据药物敏感试验合理使用抗生素.     

Enhanced zinc consumption causes memory deficits and increased brain levels of zinc

Zinc deficiency has been shown to impair cognitive functioning, but little work has been done on the effects of elevated zinc. This research examined the effect on memory of raising Sprague-Dawley rats on enhanced levels of zinc (10 ppm ZnCO3; 0.153 mM) in the drinking water for periods of 3 or 9 months, both pre- and postnatally. Controls were raised on lab water. Memory was tested in a series of Morris Water Maze (MWM) experiments, and zinc-treated rats were found to have impairments in both reference and working memory. They were significantly slower to find a stationary platform and showed greater thigmotaxicity, a measure of anxiety. On a working memory task, where the platform was moved each day, zinc-treated animals had longer latencies over both trials and days, swam further from the platform, and showed greater thigmotaxicity. On trials using an Atlantis platform, which remained in one place but was lowered on probe trials, the zinc-treated animals had significantly fewer platform crossings, spent less time in the target quadrant, and did not swim as close to the platform position. They had significantly greater latency on nonprobe trials. Microprobe synchrotron X-ray fluorescence (microSXRF) confirmed that brain zinc levels were increased by adding ZnCO3 to the drinking water. These data show that long-term dietary administration of zinc can lead to impairments in cognitive function.     

Cellular site of active K absorption in the guinea-pig distal colonic epithelium

 The mammalian distal colon, which is composed of different cell types, actively transports Na, K and Cl in absorptive and K and Cl in secretory directions. To further characterize the K absorption process and to identify the cells involved in K absorption, unidirectional Rb fluxes and luminal Rb uptake into different epithelial cell types were determined in isolated guinea-pig distal colon. Net Rb absorption (1.5–2.5 μmol·h^–1·cm^–2) was not influenced by inhibition of Na transport with amiloride or by incubating both sides of the epithelium with Na-free solutions, but was almost completely abolished by luminal ouabain, ethoxzolamide or by incubating both sides of the epithelium with Cl-free solutions. Luminal Rb uptake, blockable by luminal ouabain, preferentially occurred in columnar surface and neck cells, to a lesser extent in surface goblet cells and to an insignificant degree in lower crypt cells. Employing a luminal Rb-Ringer (5.4 mM Rb) the Rb concentration increased within 10 min in columnar surface and neck, surface goblet and lower crypt cells to 70, 32 and about 10 mmol·kg^–1 wet weight, respectively. The presence of 5.4 mM K in the luminal incubation solution reduced Rb uptake almost completely indicating a much higher acceptance of the luminal H-K-ATPase for K than for Rb. The increase in Na and decrease in K concentrations in surface and neck cells induced by luminal ouabain might indicate inhibition of the basolateral Na-K-ATPase or drastic enhancement of cellular Na uptake by the Na-H exchanger. Bilateral Na-free incubation did not alter Rb uptake, but bilateral Cl-free incubation drastically reduced it. Inhibition of net Rb absorption by ethoxzolamide and inhibition of both Rb absorption and Rb uptake by bilateral Cl-free incubation support the notion that cellular CO₂ hydration is a necessary prerequisite for K absorption and that HCO₃ leaves the cell via a Cl-HCO₃ exchanger. Since ouabain-inhibitable transepithelial Rb flux and luminal Rb uptake rate by surface and neck cells were about the same, Rb(K) absorption seems to be accomplished mainly by columnar surface cells. Received: 4 August 1997 / Received after revision: 12 November 1997 / Accepted: 4 December 1997     

Effect of epitalon on the lifespan increase in Drosophila melanogaster

The geroprotector activity of epitalon, a synthetic tetrapeptide Ala–Glu–Asp–Gly, was studied on the Drosophila melanogaster wild strain Canton-S. The substance was added to the culture medium only at the developmental stage (from egg to larva). Epitalon significantly increased the lifespan (LS) of imagoes by 11–16% when applied at unprecedented low concentrations — from 0.001×10^–6 to 5×10^–6 wt.% of culture medium for males and from 0.01×10^–6 to 0.1×10^–6 wt.% of culture medium for females. The increase in LS did not depend on the substance dose. Effective concentrations of epitalon were 16 000–80 000 000 times lower than those of melatonin. The possible mechanisms of the antioxidant and regulatory effects of epitalon are discussed.     

Isolation and partial characterisation of neuronal growth cones from neonatal rat forebrain

We have devised a method for the isolation of viable neuronal growth cones from neonatal rat forebrain. The method involves differential and density gradient centrifugation and exploits the relatively low buoyant density (approximately 1.018 g/cm3) of growth cones. There are no known biochemical markers for growth cones and it was necessary therefore to monitor for their presence during the isolation using transmission electron microscopy. Several criteria were used to identify isolated growth cones including the presence of filopodia, an extensive system of branching, tubular smooth endoplasmic reticulum and a region rich in microfilaments subjacent to the plasma membrane. These morphological features are similar to those of growth cones identified unequivocally in intact developing brain and in tissue culture. Electron microscopical analysis showed that greater than 90% of membrane-bound, identifiable objects in one fraction were growth cones by these criteria. The major contaminant consisted of membrane sacs and vesicles of unidentified origin. There were only small amounts of isolated rough endoplasmic reticulum and mitochondria. Isolated growth cones were roughly spherical in shape with a diameter of 1.9 +/- 0.5 micron (mean +/- 1 SD). They usually contained mitochondria, large granular vesicles and small vesicles, and occasionally contained coated vesicles, lysosomes, lamellar bodies and multivesicular bodies, and only very rarely, intermediate filaments. Occasionally, growth cones had rudimentary synapses on them. The viability of isolated growth cones was investigated by observing their behaviour in short-term culture. After a few hours in culture on poly-D-lysine-coated coverslips, growth cones flattened down and extended filopodia-like processes. This behaviour was inhibited by cytochalasin B and reversibly by cold (4 degrees C). We conclude that physiologically active growth cones can be isolated rapidly and in large numbers by the method described here.     

中药配方颗粒辅料麦芽糊精的HPLC定量检测与中红外光谱半定量检测

目的 建立中药配方颗粒辅料麦芽糊精的检测方法,为中药配方颗粒质量评价提供分析技术支持。方法 建立配方颗粒中果糖、葡萄糖、蔗糖、麦芽糖、乳糖的HPLC定量检测方法,根据样品经糖化酶水解后的葡萄糖增加量换算出麦芽糊精含量。基于中药浸膏与麦芽糊精不同比例混合物的中红外光谱（mid-infrared spectroscopy,MIRS）特征峰差异,建立配方颗粒中麦芽糊精的半定量检测方法。结果 所建立的HPLC方法可以准确定量检测配方颗粒中果糖、葡萄糖、蔗糖、麦芽糖、乳糖。如果中药浸膏自身含有较多可经糖化酶水解产生葡萄糖的成分（淀粉、蔗糖等）,HPLC检测配方颗粒中麦芽糊精时存在系统性正误差,可通过中药浸膏酶解后葡萄糖增加量、待测样品酶解后蔗糖减小量等部分修正正误差。如果中药浸膏自身少含或不含淀粉、蔗糖等可经糖化酶水解产生葡萄糖的成分,HPLC检测配方颗粒中麦芽糊精时存在系统性负误差,换算公式偏差、麦芽糊精纯度、样品处理损失等导致根据样品酶解后葡萄糖增加量计算的麦芽糊精“检测含量”预期低于根据生产投料计算的麦芽糊精“名义含量”。麦芽糊精含量越高,配方颗粒MIRS中1200～900 cm-1区域的...     

基于指纹图谱和多指标含量测定的蜜远志质量评价

目的 建立指纹图谱和多指标定量的蜜远志质量评价方法。方法 Kromasil 100-5-C₁₈色谱柱（250 mm×4.6 mm,5 μm）为固定相,以乙腈（A）-0.2%磷酸水溶液（B）为流动相进行梯度洗脱,体积流量为1.0 mL/min,检测波长318 nm。对16批蜜远志建立指纹图谱,确定共有峰并进行相似度分析结合化学计量学分析。采用Waters Xbridge Shiled RP C₁₈色谱柱（250 mm×4.6 mm,5 μm）进行同时测定远志（口山）酮III、3,6''-二芥子酰基蔗糖、细叶远志皂苷、远志酸、远志皂苷元的含量。结果 在指纹图谱研究中,标定了27个共有峰,结合对照品和HPLC-Q-TOF-MS共指认8个成分,分别为西伯利亚远志糖A5、西伯利亚远志糖A6、远志（口山）酮IX、远志（口山）酮III、3,6''-二芥子酰基蔗糖、细叶远志皂苷、远志酸、远志皂苷元。聚类分析将16批样品分为3类;经主成分分析,主成分1～3是影响蜜远志质量评价的主要因子。多指标含量测定中远志（口山）酮III、3,6''-二芥子酰基蔗糖、细叶远志皂苷、远志酸、远志皂苷元的质量分数分别为0.027%～0.068%、0.074%～0.798%、1.1%～1.4%、0.15%～0.36%、0.15%～0.37%。经方法学验证,线性关系良好（r≥0.999 6）,平均加样回收率为97.15%～100.9%。结论 所建立的指纹图谱结合多指标含量测定方法准确、高效,特征性强,可为蜜远志饮片的质量评价和临床应用提供有效参考。