Biochemical differentiation in the tobacco flower probed with monoclonal antibodies

We have isolated a series of monoclonal antibodies that react to antigens in flowers of Nicotiana tabacum L. (tobacco) displaying specificity or preferentiality in their cell and tissue distributions. We immunized mice with extracts from tobacco flowers and then screened the hybridomas by enzyme-linked immunosorbent assay (ELISA) against extracts from leaves, sepals, petals, stamens and pistils; twenty five were chosen from the total screened. The antigens detected by about half of the antibodies were periodate-sensitive, implying that the epitopes were carbohydrate. Competition ELISA assays were used to determine if any antibodies were reacting to the same epitopes. Western blot analysis showed that while some antibodies reacted to specific bands, the bulk either failed to react or reacted to multiple bands, consistent with a glyco-conjugate nature for many of the antigens. Analysis of the spatial pattern of antigen distribution within tobacco flowers by immunolocalization showed that some antibodies recognized epitopes that were limited to very specific cells and tissues. We used the immunolocalization technique to analyze a mutant with stigmoid anthers: an antibody recognizing a pistil transmitting-tract antigen also reacted to cells in stigmoid anthers. Our results with this antibody set imply that biochemical differentiation within the tobacco flower includes cell-and tissue-specific glyco-moeities, and also that similarities, at the biochemical level, exist between a normal floral organ and the abnormal organ in a phenotype with a developmental switch.Abbreviations ELISA enzyme-linked immunosorbent assay - Fg immunoglobulin - kDa kilodalton     

Treatment of canine aspiration pneumonitis: fluid volume reduction vs. fluid volume expansion

The aspiration of gastric acid causes pulmonary edema and hypoxemia. One approach to the management of this syndrome is to raise cardiac output (Qt) and O2 delivery (QO2) to ensure tissue oxygenation (VO2) at the risk of increasing the edema. Another approach reduces the edema by reducing pulmonary microvascular pressure (Pmv) at the risk of reducing QO2 and VO2. We compared these approaches in 24 anesthetized, ventilated dogs with pulmonary wedge pressure (Ppw), a clinical approximation of Pmv, of 12.5 mmHg. Before and again 1 h after endobronchial instillation of 0.1 N HCl, we measured Qt, QO2, VO2, venous admixture, and in vivo extravascular lung liquid. The dogs were then randomly divided into four equal groups: 1) 12.5 mmHg Ppw, high Qt; 2) 7.5 mmHg Ppw, intermediate Qt; 3) 4.5 mmHg Ppw, low Qt; and 4) 4.5 mmHg Ppw plus dopamine, intermediate Qt. Measured values were followed for 4 more h, after which the lungs were excised to compare wet weight-to-body weight ratios (W/B). When plasmapheresis reduced Ppw at 1 h, edema did not increase further and W/B of groups 2 (21 +/- 3), 3 (18 +/- 3), and 4 (22 +/- 3) were significantly less than in group 1 (27 +/- 3) (P less than 0.001). Although Qt decreased with Ppw, increased hematocrit and reduced venous admixture maintained QO2 in group 2 but not in group 3. In group 4 an intermediate Qt maintained QO2 even at 4.5 mmHg Ppw but edema increased to the group 2 level presumably because Pmv rose with Qt on dopamine. VO2 remained constant over time in each group. These data demonstrate that canine HCl-induced pulmonary edema, measured in vivo or gravimetrically, is very sensitive to reductions in Pmv. Moreover, the lowest Pmv (and QO2) was well tolerated because an O2 supply dependency of VO2 was not observed.     

Estimated feeding rates and energy requirements of gentoo penguins,Pygoscelis papua,at Macquarie Island

Summary Water and sodium turnovers of 6–7 week old gentoo penguin chicks and breeding adults were measured using isotopically labelled water and sodium. Influx rates for chicks averaged 188 ml·kg^-1·day^-1 and 13.9 mmol·kg^-1·day^-1 for water and sodium, respectively. Chicks consumed an estimated 228 g·kg^-1·day^-1 fresh food or 886 kJ kg^-1 day. These values correspond to 761 g·day^-1 or 2945 kJ·day^-1 for a gentoo chick mid-way through the growth period. Flux rates for adults attending chicks ranged from 199 to 428 ml·kg^-1·day^-1 for water and from 15 to 36 mmol·kg^-1·ay^-1 for sodium.     

Bias in leaf area — sapwood area ratios and its impact on growth analysis in Pinus contorta

Summary Two alternative estimators of individual tree leaf area (A₁) area are used to derive estimates of leaf-area index (L) for 40 plots in Pinus contorta Dougl. stands. One estimator of A₁ is based on the common assumption of a constant ratio between A₁ and sapwood cross-sectional area at breast height (A_s). The second estimator of A₁ accounts for tree-to-tree variation in the relation between A₁ and A_s. The apparent relationship between stand growth and leaf-area index is strongly dependent on the way leaf area is estimated. When L is derived from a constant A₁A_s ratio, stand growth appears to be strongly correlated with L. However, when L is based on estmates of A₁ that account for tree-to-tree variation in the A₁ — A_s relation, stand growth is seen to be only weakly related to L. Stand structure, quantified as percent live-crown, accounts for a great deal of the observed variation in leaf-area efficiency. These contrasting relationships illustrate the importance of unbiased estimates of L in interpreting the link between stand-level processes and leaf area.Utah Aggriculural Experiment Station Journal Paper No. 3333     

Nonenzymatic isolation and culture of adult islets from atrophic pancreata of copper-deficient rats: A morphologic analysis

Summary The purpose of this study was to develop a nonenzymatic method of isolating adult islets using atrophied pancreata from copper-deficient rats and to analyze their morphologic characteristics and behavior in culture. This unusual model of isolation was studied because islets remain intact in the course of dietary copper deficiency while the acinar glandular component of the pancreas undergoes selective atrophy and lipomatosis. Small fragments containing islets were readily microdissected from atrophied glands and placed in culture. Within 24 h the fragments congealed into small irregular- to spherical-shaped masses within which the darker profile of islets could be distinguished. Within a period of 3 to 5 d, islet tissue began to bud from the lipocytic mass until by Day 7 spherical aggregates of intact islet tissue separated from the residual fragments. Subsequent to further in vitro treatment, these islets could be maintained as free viable spherical masses if periodically agitated, as attached stationary islets which developed monolayer growth if left undisturbed and as aggregated masses of islet tissue forming megaislets if combined in small groups. Grouped islets treated with actinomycin D and cycloheximide did not exhibit aggregation when incubated with these inhibitors. This suggests that megaislet formation was an active process requiring protein-RNA synthesis rather than passive clumping or aggregation that can accompany metabolically altered or dying islets undergoing cellular shedding and adhesion. Immunohistochemical localization demonstrated that insulin, glucagon, somatostatin, and pancreatic polypeptide-immunoreactive cell types were present within the islets derived from this technique. The cellular topography of these islets was not unlike that described by others for islets cultured from enzymatic isolation. This culture model may serve as a resource for mature, viable islets isolated without mechanical or enzymatic disaggregation which can have attenuating effects on islet function. This work was supported by a research grant from the Diabetes Research and Education Foundation.     

Feedback delays and relaxation expectancies in EMG biofeedback

The use of noncontingent feedback controls in studies of the efficacy and process of electromyographic (EMG) biofeedback may yield results confounded by differential expectancies for relaxation. Furthermore, the role of expectancies in producing psychological and physical relaxation as well as reducing muscle activity is unclear. This study investigated the effects of feedback delays and induced relaxation expectancies on EMG activity and experienced relaxation. One hundred four non-clinical subjects participated in one auditory frontal EMG biofeedback training session. Subjects were assigned to one of four computerized feedback delay conditions (0.0037, 0.7493, 2.2481, 6.7444 s) and to one of two relaxation expectancy conditions (positive or negative). During 20 minutes of biofeedback training, all groups decreased frontal activity. Feedback delays interacted with training epochs in affecting EMG; the longest delay group reduced frontal activity more slowly than the shortest delay group during training. Positive relaxation expectancies produced greater experienced relaxation than did negative relaxation expectancies. Instrumental and expectancy factors in EMG biofeedback appear to operate independently of each other by reducing physiological activity and producing psychological relaxation respectively.This study was completed by the first author under the direction of the second author in partial fulfillment of the requirements for the Master of Arts degree. We gratefully acknowledge the computerization advice and assistance provided by Larry Wheeler, and the assistance in data collection provided by Dawn Dexter and Michael Winstanley.     

Interaction of opioid peptides and other drugs with multiple delta ncx binding sites in rat brain: further evidence for heterogeneity.

Recent pharmacological data strongly support the hypothesis of δ receptor subtypes as mediators of both supraspinal and spinal antinociception (δ₁ and δ₂ receptors). In vitro ligand binding data, which are fully supportive of the in vivo data, are still lacking. A previous study indicated that [³H][ -Ala², -Leu⁵]enkephalin labels two binding sites in membranes depleted of μ binding sites by pretreatment with the site-directed acylating agent, 2-(p-ethoxybenzyl)-1-diethylaminoethyl-5-isothiocyanatobenzimidazole-HCI (BIT). The main goal of the present study was to develop a ligand-selectivity profile of the two δ_ncx binding sites. The data indicated that naltrindole and oxymorphindole were relatively selective for site 1 (20-fold). [ -Ser²,Thr⁶]Enkephalin and deltorphin-II were only 2.7-fold and 2.2-fold selective for site 1. [ -Pen², -Pen⁵]Enkephalin and deltorphin-I were 80-fold and 38-fold selective for site 2.3-Iodo-Tyr- -Ala-Gly-Phe- -Leu was 52-fold selective for site 1. Morphine had moderate affinity for site 1 (K_i = 16 nM), and was about 11-fold selective for site 1. Thus, of the 10 drugs studied, only DPDPE and DELT-I were selective for site 2. Viewed collectively with other data, it is likely that the δ₁ receptor and the δ_ncx binding site are synonymous.     

Long term incubation of cardiac myocytes with oleic acid and very-low density lipoprotein reduces heparin-releasable lipoprotein lipase activity

An exogenous [³H]triolein emulsion was hydrolyzed by intact cardiac myocytes with functional LPL located on the cell surface. This surface-bound LPL could be released into the medium when cardiac myocytes were incubated with heparin. Incubation of cardiac myocytes with VLDL, or the products of TG breakdown, oleic acid or 2-monoolein, did not increase LPL activity in the medium. However, incubation of cardiac myocytes with either VLDL or oleic acid for > 60 min did reduce heparin-releasable LPL activity. In the heart, this inhibitory effect of FFA could regulate the translocation of LPL from its site of synthesis in the cardiac myocyte to its functional site at the capillary endothelium.Abbreviations LPL lipoprotein lipase - TG triacylglycerol - FFA free fatty acids - VLDL very-low density lipoprotein