Regulation by the H-2 gene complex of delayed type hypersensitivity

Identity at the major histocompatibility complex (MHC) was essential for successful transfer of delayed type hypersensitivity (DTH) in mice. The regions of the MHC involved differed according to the antigen used for sensitization. In the case of fowl gamma globulin (FGG), identity atI-A was necessary, whereas with dinitrofluorobenzene (DNFB), identity at theK, I, orD region was sufficient. These different genetic constraints probably reflect differences in the mechanisms by which antigens are presented to T lymphocytes. Cells from sensitized (CBA×C57BL)F₁ mice transferred DTH to FGG into parental-strain mice, but transfer was more effective in C57BL than in CBA with the same cell dose. This phenomenon is governed by the MHC, since there was better transfer intoH-2 ^b than intoH-2 ^k mice, regardless of their backgrounds. It may reflect the activity of an Ir gene-dependent process. Cells of one genotype (e.g., CBA), sensitized in chimeric mice derived from two MHC-incompatible strains (CBAC57BL), transferred DTH to both strains. These results do not support the notion that the genetic constraint observed in DTH transfer may be a result of the necessity for sensitized T and stimulator cells to match an identical MHC-coded cell interaction molecule. Rather, they favor the hypothesis that T cells recognize antigen, not as a naked determinant, but in close association with products of genes of the MHC.     

Ion transport across the isolated intestinal mucosa of the winter flounder,Pseudopleuronectes americanus

The isolated intestinal mucosa of the flounder, Pseudopleuronectes americanus, when bathed in a 20 mM HCO3-Ringer's solution bubbled with 1% CO2 in O2, generated a serosa-negative PD and, when short-circuited, absorbed Cl at almost 3 times the rate of Na. Reducing HCO3 to 5 mM decreased the net Cl flux by more than 60%. The following results suggest that, despite the PD, Na and Cl transport processes are nonelectrically coupled: replacing all Na with choline abolished both the PD and net Cl flux; replacing all Cl with SO4 and mannitol abolished the PD and the net Na flux; and adding ouabain (to 0.5 mM) abolished the PD and the net Cl flux. Nearly all of the unidirectional serosa-to-mucosa Cl flux (JClsm) seemed to be paracellular since it varied with PD and Cl concentration in a manner consistent with simple diffusion. JClsm was only about one-fourth of JNasm, suggesting that the paracellular pathway is highly cation-selective. The data can be explained by the following model: (i) Na and Cl uptake across the brush border are coupled 1 : 1; Na is pumped into the lateral space and Cl follows passively, elevating the salt concentration there; (ii) the tight junction is permeable to Na but relatively impermeable to Cl; and (iii) resistance to Na diffusion is greater in the lateral space (considered in its entirety) than in the tight junction. If these assumptions are correct, the serosa-negative transmural PD is due mainly to a salt diffusion potential across the tight junction and, under short-circuit condition, most of the Na pumped into the lateral space diffuses back into the luminal solution, whereas most of the Cl enters the serosal solution. Morphological features of the epithelium support this interpretation: the cells are unusually long (60 micrometer); there is little distension of the apical 12 micrometer of the lateral space during active fluid absorption; and distension distal to this region is intermittently constricted by desmosomes.     

Abscisic acid binding to subcellular fractions from leaves of Vicia faba

A centrifugation binding assay has been used to demonstrate the binding of [³H] (±) abscisic acid to membrane-rich fractions prepared from leaves of Vicia faba L. Kinetic analysis of this binding shows evidence of saturation of binding sites with increasing concentration of ligand. Scatchard analysis of these data yields a biphasic plot possibly indicating the presence of two types of binding sites. The dissocation constant for the high affinity site has been calculated to be 3.5×10^-8 mol 1^-1.     

DEVELOPMENTAL POTENTIAL OF AXILLARY BUDS OF WATER HYACINTH,EICHHORNIA CRASSIPES SOLMS. (PONTEDERIACEAE)

Buds axillary to foliage leaves of water hyacinth can elongate either as vegetative stolons or as renewal shoots produced in association with the terminal inflorescence. Stolons differ from renewal shoots in position within the shoot system, morphology, and function. Renewal shoot buds always expand, whereas stolon buds may or may not. A stolon bud develops in conjunction with the subtending leaf; as that leaf matures, the stolon bud reaches a critical period in development. At this point, the bud either continues to expand, producing a stolon, or it stops growth and matures. Maturation is not irreversible, but the probability of a bud expanding decreases as bud age increases. In the field, buds on plants at the water hyacinth mat edge frequently produce stolons, whereas buds on plants inside the mat rarely do so. Leaf morphology also varies between plants in these two regions of the mat. The particular association of leaf and branch type found in the field, however, can be reversed experimentally, indicating that although leaf and bud development are coordinated, the particular course of each is independent.     

Kinetic analysis of F-actin depolymerization in polymorphonuclear leukocyte lysates indicates that chemoattractant stimulation increases actin filament number without altering the filament length distribution

The rate of filamentous actin (F-actin) depolymerization is proportional to the number of filaments depolarizing and changes in the rate are proportional to changes in filament number. To determine the number and length of actin filaments in polymorphonuclear leukocytes and the change in filament number and length that occurs during the increase in F-actin upon chemoattractant stimulation, the time course of cellular F-actin depolymerization in lysates of control and peptide-stimulated cells was examined. F-actin was quantified by the TRITC-labeled phalloidin staining of pelletable actin. Lysis in 1.2 M KCl and 10 microM DNase I minimized the effects of F-actin binding proteins and G-actin, respectively, on the kinetics of depolymerization. To determine filament number and length from a depolymerization time course, depolymerization kinetics must be limited by the actin monomer dissociation rate. Comparison of time courses of depolymerization in the presence (pointed ends free) or absence (barbed and pointed ends free) of cytochalasin suggested depolymerization occurred from both ends of the filament and that monomer dissociation was rate limiting. Control cells had 1.7 +/- 0.4 x 10(5) filaments with an average length of 0.29 +/- 0.09 microns. Chemo-attractant stimulation for 90 s at room temperature with 0.02 microM N-formylnorleucylleucylphenylalanine caused a twofold increase in F-actin and about a two-fold increase in the total number of actin filaments to 4.0 +/- 0.5 x 10(5) filaments with an average length of 0.27 +/- 0.07 microns. In both cases, most (approximately 80%) of the filaments were quite short (less than or equal to 0.18 micron). The length distributions of actin filaments in stimulated and control cells were similar.     

Turnover of Dopamine and Serotonin and Their Metabolites in the Striatum of Aged Rats

Turnover of dopamine (DA), serotonin [5-hydroxytryptamine (5-HT)], and their metabolites has been measured in adult and aged rats. Turnover rates of 3,4-dihydroxyphenylacetic acid (DOPAC), homovanillic acid (HVA), and 5-hydroxy-3-indoleacetic acid (5-HIAA) have been assayed from the disappearance rates after blocking by pargyline inhibition of monoamine oxidase (MAO) and from the accumulation rates by probenecid inhibition of the probenecid-sensitive transport system. DA and 5-HT turnover rates have been measured as accumulation rates of 3,4-dihydroxyphenylalanine and 5-hydroxytryptophan, respectively, after central decarboxylase inhibition by 3-hydroxybenzylhydrazine (NSD-1015) and as accumulation rates of DA and 5-HT after pargyline inhibition of MAO. The DA turnover rate after NSD-1015 was 23.9% lower in aged rats than in adults, whereas after pargyline there was no significant difference between the two age groups. The HVA fractional rate constant and turnover after pargyline were lower in aged rats than in adults, and HVA turnover after probenecid was higher in aged rats than in adults. The DOPAC-HVA pathway seems to be reinforced at the expense of DOPAC conjugation. In aged and adult rats whose 5-HT steady-state levels were not statistically different, the 5-HT turnover rate after pargyline and NSD-1015 treatment was lower in aged rats than in adults. An increase of 5-HIAA levels after pargyline and probenecid treatment in aged rats could be due to the handling stress.     

Detection of salmonellas by DNA hybridization with a fluorescent alkaline phosphatase substrate.

This study evaluates a DNA hybridization assay for salmonella with AttoPhos (JBL Scientific, San Luis Obispo, CA), a fluorescent substrate for alkaline phosphatase. The probe used (50 ng/ml) was a biotinylated 600 bp fragment consisting of a tandem repeat of an insertion sequence (IS200) found in most Salmonella spp. evaluated. The hybridization was carried out at 65 degrees C for 2 h without prior prehybridization and hybrids were detected by the addition of a streptavidin-alkaline phosphatase conjugate. Circles (5 mm) were cut from the membrane and placed in a cuvette containing 1 ml of 1 mmol/l AttoPhos. The reaction was evaluated after 30 min at 37 degrees C with a fluorometer with an excitation wavelength of 440 nm and an emission wavelength of 550 nm. The sensitivity of the probe was estimated to be 10,000 copies of target DNA or 5 x 10(-20) mol of DNA. All 74 salmonella strains tested reacted with the probe but none of the 98 heterologous species tested gave positive results. The results of this study indicate that our assay method, which employs a biotinylated tandem repeat of IS200 and AttoPhos, is a specific and highly sensitive quantitative method for the detection of salmonellas.