Basolateral K channels in an insect epithelium. Channel density, conductance, and block by barium

K channels in the basolateral membrane of insect hindgut were studied using current fluctuation analysis and microelectrodes. Locust recta were mounted in Ussing-type chambers containing Cl-free saline and cyclic AMP (cAMP). A transepithelial K current was induced by raising serosal [K] under short-circuit conditions. Adding Ba to the mucosal (luminal) side under these conditions had no effect; however, serosal Ba reversibly inhibited the short-circuit current (Isc), increased transepithelial resistance (Rt), and added a Lorentzian component to power density spectra of the Isc. A nonlinear relationship between corner frequency and serosal [Ba] was observed, which suggests that the rate constant for Ba association with basolateral channels increased as [Ba] was elevated. Microelectrode experiments revealed that the basolateral membrane hyperpolarized when Ba was added: this change in membrane potential could explain the nonlinearity of the 2 pi fc vs. [Ba] relationship if external Ba sensed about three-quarters of the basolateral membrane field. Conventional microelectrodes were used to determine the correspondence between transepithelially measured current noise and basolateral membrane conductance fluctuations, and ion-sensitive microelectrodes were used to measure intracellular K activity (acK). From the relationship between the net electrochemical potential for K across the basolateral membrane and the single channel current calculated from noise analysis, we estimate that the conductance of basolateral K channels is approximately 60 pS, and that there are approximately 180 million channels per square centimeter of tissue area.     

Pathways involved in fluid phase and adsorptive endocytosis in neuroblastoma

The endocytosis of ricin, horseradish peroxidase (HRP), and a conjugate of ricin-HRP by monolayer cultures of murine neuroblastoma was studied using morphological and biochemical techniques. The binding of (125)I-ricin and (125)I-ricin-HRP to cells at 4 degrees C, as a function of ligand concentration, was a saturable process. The apparent affinity constants, determined at equilibrium, were 2.8 X 10(6) M(-1) for ricin and 1 x 10(6) M(-1) for ricin-HRP. The number of binding sites per cell was 8 x 10(7) and 3 x 10(7) for the lectin and the conjugate, respectively. The binding of (125)I-ricin to monolayers as not proportional to cell density. We found reduced binding at higher cell concentrations, suggesting a decrease in the accessibility of the ligand for the receptor site or fewer sites with increasing cell population. Neuroblastoma cells have an acid-phosphatase-positive network of cisternae and vesicles near the Golgi apparatus (GERL). Ricin-HRP undergoes endocytosis in vesicles and cisternae corresponding to GERL, and in residual bodies (dense bodies). The cellular uptake of ricin-HRP was 100-200 times greater than free HRP and there was no stimulation of fluid phase endocytosis by ricin. When monolayers were exposed to concentrations of native HRP 100-fold that of the conjugate, cellular uptake of peroxidase was comparable, but HRP was localized only in residual bodies and never in elements of GERL. These results support the conclusion that GERL is involved in the adsorptive endocytosis of ricin-HRP, while residual bodies are involved in the bulk uptake of HRP. In addition, the binding, uptake, and possible recycling of (125)I- subunit B (the binding subunit) of ricin and of (125)I-ricin was examined by quantitative electron microscope autoradiography. Both ricin and its binding subunit displayed similar autoradiographic grain distributions at 4 degrees C, and there was no evidence of their breakdown or recycling to the plasma membrane during endocytosis for 2 h.     

Mapping quantitative trait loci associated with aluminum toxin tolerance in NJRIKY recombinant inbred line population of soybean (Glycine max)

To investigate the genetic mechanism of AI-tolerance in soybean,a recombinant inbred line population (RIL) with 184 F2:7:11 lines derived from the cross of Kefeng No.1 x Nannong 1138-2 (AI-tolerant x AI-sensitive) were tested in pot experimentwith sand culture medium in net room in Nanjing.Four traits,i.e.plant height,number of leaves,shoot dry weight and root dry weight at seedling stage,were evaluated and used to calculate the average membership index (FAi) as the indicator of AI-tolerance.The composite interval mapping (ClM) under WinQTL Cartographer v.2.5 detected five QTLs (i.e.qFAiol,qFAi-2,qFAi-3,qFAi-4 and qFAi-5),explaining 5.20%-9.07% of the total phenotypic variation individually.While with the multiple interval mapping (MIM) of the same software,five QTLs (qFAi-1,qFAi-5,qFAi-6,qFAi-7,and qFAi-8) explaining 5.7%-24.60% of the total phenotypic variation individually were mapped.Here qFAi-1 and qFAi-5 were detected by both CIM and MIM with the locations in a same flanking marker region,GMKF046-GMKF080 on B1 and satt278-sat_95 on L,respectively.While qFAi-2 under CIM and qFAi-6 under MIM both on D1b2 were located in neighboring regions with their confidence intervals overlapped and might be the same locus.Segregation analysis under major gene plus polygene inheritance model showed that Al-tolerance was controlled by two major genes (h2mg =33.05%) plus polygenes (h2pg=52.73%).Both QTL mapping and segregation analysis confirmed two QTLs responsible for Al-tolerance with relatively low heritability,and there might be a third QTL,confounded with the polygenes in segregation analysis.     

Effects of feed intake level on efficiency of microbial protein synthesis and nitrogen balance in Boran steers consuming tropical poor-quality forage

This study aimed at evaluating the effects of feed intake level on the efficiency of rumen microbial protein synthesis (EMPS), nitrogen (N) excretion, and N balance in twelve 18-months old Boran (Bos indicus) steers with initial average liveweight of 183 kg (standard deviation (SD) 15.2). The experiment followed a 4 × 4 complete Latin Square design with four dietary treatments tested in four periods. Each period ran for 5 weeks with 3 weeks of adaptation and 2 weeks of sample collection; separated by 2 weeks of re-feeding. Steers were fed at 100%, 80%, 60%, and 40% of their metabolisable energy requirement for maintenance (MER, referred to as MER100, MER80, MER60, and MER40, respectively). Steers receiving MER80, MER60, and MER40 were only fed Rhodes grass hay. MER100 steers were offered Rhodes grass hay at 80% of their MER and cottonseed meal and sugarcane molasses at each 10% of MER. Mean daily dry matter intake differed between treatments (p < 0.001) and ranged between 2.1 kg/animal (SD 0.13) in MER40 and 4.5 kg/animal (SD 0.31) in MER100. Urinary N excretion and N balance did not differ between MER80, MER60, and MER40. According to contrast test, declining feed intake level from MER80 to MER40 reduced duodenal microbial crude protein flow (p < 0.001), but did not alter the EMPS (g microbial N/kg digestible organic matter intake). Yet, if scaled to N intake, EMPS increased (p < 0.049), whereas total N and faecal N excretions decreased linearly with declining intake level (p < 0.001 for both variables). At similar grass hay intake, duodenal microbial crude protein flow was 41% higher in MER100 than in MER80 steers (p < 0.001). In cattle offered poor-quality tropical forage below their MER, the very low EMPS and thus microbial protein supply aggravate the negative effects of low dietary nutrient and energy intakes in periods of feed shortage.