Regulation of the Amount of Starch in Plant Tissues by ADP Glucose Pyrophosphorylase.

Starch, a major storage metabolite in plants, positively affects the agricultural yield of a number of crops. Its biosynthetic reactions use adenosine diphosphate glucose (ADPGlc) as a substrate; ADPGlc pyrophosphorylase, the enzyme involved in ADPGlc formation, is regulated by allosteric effectors. Evidence that this plastidial enzyme catalyzes a rate-limiting reaction in starch biosynthesis was derived by expression in plants of a gene that encodes a regulatory variant of this enzyme. Allosteric regulation was demonstrated to be the major physiological mechanism that controls starch biosynthesis. Thus, plant and bacterial systems for starch and glycogen biosynthesis are similar and distinct from yeast and mammalian systems, wherein glycogen synthase has been demonstrated to be the rate-limiting regulatory step.

Fungal pathogen protection in potato by expression of a plant defensin peptide.

Defensins are small cysteine-rich peptides with antimicrobial activity. We demonstrate that the alfalfa antifungal peptide (alfAFP) defensin isolated from seeds of Medicago sativa displays strong activity against the agronomically important fungal pathogen Verticillium dahliae. Expression of the alfAFP peptide in transgenic potato plants provides robust resistance in the greenhouse. Importantly, this resistance is maintained under field conditions. There have been no previous demonstrations of a single transgene imparting a disease resistance phenotype that is at least equivalent to those achieved through current practices using fumigants.

Hyperoxic training increases work capacity after maximal training at moderate altitude.

High-intensity training may be difficult to sustain due to limitations in systemic oxygen transport, particularly at high altitudes. The purpose of this study was to examine the effects of a high-intensity training protocol using hyperoxic gas breathing in athletes "maximally trained" at an altitude of 1,600 m. Five subjects underwent progressive cycle training until they reached a plateau of aerobic capacity, maximal workload, and endurance time at 85 percent maximal workload. Significant decreases (2 to 6 percent) in arterial oxygen saturation were found after the 85 percent maximal workload tests. Training intensity was then increased to 95 percent maximal workload while the subjects breathed a gas mixture containing at least 70 percent oxygen. After 6 weeks of hyperoxic training, exercise parameters were compared with the plateau values obtained during the baseline training period. Total time during maximal cycle testing increased from 19.1 to 19.6 min (p = 0.015), heart rate at 85 percent maximal workload decreased from 168 to 163 bpm (p = 0.047), and endurance time at 85 percent maximal workload increased from 6.2 to 8.2 min (p = 0.012). There was a trend toward improvement of maximal workload. We conclude that hyperoxic training increases work capacity after attainment of "maximal training" at moderate altitude.

Short-Term and medium-term effects of spinal cord tract transections on soleus H-reflex in freely moving rats.

Spinal cord function is normally influenced by descending activity from supraspinal structures. When injury removes or distorts this influence, function changes and spasticity and other disabling problems eventually appear. Understanding how descending activity affects spinal cord function could lead to new means for inducing, guiding, and assessing recovery after injury. In this study, we investigated the short-term and medium-term effects of spinal cord bilateral dorsal column (DC), unilateral (ipsilateral) lateral column (LC), bilateral dorsal column ascending tract (DA), or bilateral dorsal column corticospinal tract (CST) transection at vertebral level T8-T9 on the soleus H-reflex in freely moving rats. Data were collected continuously for 10-20 days before and for 20-155 days after bilateral DC (13 rats), DA (10 rats), CST (eight rats), or ipsilateral LC (seven rats) transection. Histological examination showed that transections were 98(+/- 3 SD)% complete for DC rats, 80(+/- 20)% complete for LC rats, 91(+/- 13 SD)% complete for DA rats, and 95(+/-13)% complete for CST rats. LC, CST, and DA transections produced an immediate (i.e., first-day) increase in H-reflex amplitude. LC transection also produced a small decrease in background activity in the first few posttransection days. Other than this small decrease, none of the transections produced evidence for the phenomenon of spinal shock. For all transections, all measures returned to or neared pretransection values within 2 weeks. DA and LC transections were associated with modest increase in H-reflex amplitude 1-3 months after transection. These medium-term effects must be taken into account when assessing transection effects on operant conditioning of the H-reflex. At the same time, the results are consistent with other evidence that, while H-reflex rate dependence and H-reflex operant conditioning are sensitive measures of spinal cord injury, the H-reflex itself is not.

A battery of potential alternatives to the Draize test: uridine uptake inhibition, morphological cytotoxicity, macrophage chemotaxis and exfoliative cytology.

Four assays that may serve as components of a battery of alternatives to the conventional Draize test were described. The exfoliative cytology assay is a refinement of the Draize test that may provide a more sensitive and more objective end-point. The macrophage migration assay addresses the inflammatory aspects of the physiological response to irritation. The uridine uptake inhibition assay uses a quantitative, reversible end-point to detect the short-term action of agents on cell membranes and cell phosphorylative potential. Finally, the cytological assay serves as a rapid, easily performed general indicator of cytotoxic action. The two latter assays have been demonstrated to correlate very well with Draize test results and with each other for a wide range of test agents.

Progress and problems in evaluating and validating alternative assays in toxicology.

Various techniques have been proposed and subsequently developed as potential replacements for whole-animal topical toxicology assays. Comparisons of in vitro and in vivo endpoints, the solubility of test agents, solvent effects, interactions between components of complex test samples and the standardization of data presentation are factors that influence not only individual laboratory results but interlaboratory reproducibility and the acceptance of newly developed alternative methods. Representative alternative test systems for evaluating topical toxicity are discussed. Examples of problems, data and solutions based on the authors' experience and on experience in other laboratories are reviewed. Specific problems in choosing endpoints, comparing results, recognizing responses of different types of target cell, evaluating the influences of support media in culture and interpreting published animal-based data are included.

PHA production, from bacteria to plants.

The genes encoding the polyhydroxyalkanoate (PHA) biosynthetic pathway in Ralstonia eutropha (3-ketothiolase, phaA or bktB; acetoacetyl-CoA reductase, phaB; and PHA synthase, phaC) were engineered for plant plastid targeting and expressed using leaf (e35S) or seed-specific (7s or lesquerella hydroxylase) promoters in Arabidopsis and Brassica. PHA yields in homozygous transformants were 12-13% of the dry mass in homozygous Arabidopsis plants and approximately 7% of the seed weight in seeds from heterozygous canola plants. When a threonine deaminase was expressed in addition to bktB, phaB and phaC, a copolyester of 3-hydroxybutyrate and 3-hydroxyvalerate was produced in both Arabidopsis and Brassica.

Susceptibility of laboratory rats, hamsters, and mice to wound infection with Staphylococcus aureus.

Response of rodents to experimentally induced subcutaneous infection was examined to determine whether laboratory rats used in invasive procedures have a superior ability to withstand wound infection than do hamsters and mice. Rats, hamsters, and mice were injected subcutaneously with 10(9), 10(7), and 10(5) colony-forming units of Staphylococcus aureus. Quantitative counts of viable S aureus from the injection site, bacteriologic cultures of heart blood, and histologic examinations of the subcutaneous tissues were performed. Multiple linear regression of the quantitative data and equality of regression lines among groups were determined. Results indicated that the ability to eliminate bacteria varied between species and depended on the dose injected within each species. Compared with hamsters and mice at all doses, rats eliminated bacteria faster and had the mildest and most rapidly organized inflammatory response after inoculation. Experimental bacteremia developed in 3.7% of all animals evaluated, with no species-specific pattern. The rat was more resistant to localized wound infection with S aureus than were hamsters and mice.

Rigid internal fixation for shoulder arthrodesis.

Fifteen patients underwent a shoulder arthrodesis utilizing standard dynamic compression plate fixation, but with limited postoperative immobilization with only an abduction pillow. In each case, the position of the extremity relative to the scapula and trunk was recorded immediately postoperatively, at regular intervals until fusion, and at follow-up evaluations. Thirteen of 15 shoulders fused without change of intraoperative position after an average postoperative period of 4 months. One patient lost position in the early postoperative period secondary to inadequate fixation, but subsequently fused. Another who demonstrated a persistent non-union at 2 1/2 years was subsequently explored and underwent a bone graft. Four patients complained of residual symptomatic hardware, with two requiring surgical removal of the plate and screws. All but one patient were satisfied with the clinical result at follow up. Only two patients were within 5 degrees of the preoperatively determined position of 30 degrees abduction, 30 degrees forward flexion, and 30 degrees internal rotation. However, almost all were able to function satisfactorily. The authors concluded that shoulder arthrodesis utilizing rigid internal fixation without postoperative cast or brace immobilization maximizes patient comfort without compromising the success of arthrodesis. However, control of arm position remains inexact and additional modifications are needed to ensure fusion position and to minimize disability.

Wire-bottom versus solid-bottom rodent caging issues important to scientists and laboratory animal science specialists.

Recent emphasis in the National Research Council's Guide for the Care and Use of Laboratory Animals and by the Association for Assessment and Accreditation of Laboratory Animal Care, International, related to the availability of bedding in rodent cages raises regulatory and accreditation issues in the toxicology-laboratory setting. This article reviews the results of a recent survey of 12 United States-based pharmaceutical and contract toxicology laboratories. The perceived benefits and issues related to the use of wire-bottom and bedded caging for rodent studies are presented. The 1999 survey showed that more than 80% of the rodents in surveyed toxicology facilities were housed in wire-bottom cages. Long-term budget expenses related to supplies and waste disposal are assessed. Considerable short-term and long-term costs to programs would be associated with a change from wire-bottom to solid-bottom caging. A review of the past and recent literature related to animal preferences and cage-associated animal lesions is included. The importance of IACUC review of caging chosen by the investigative staff is emphasized.

Discovering the invisible Puerto Rican slave family: demographic evidence from the eighteenth century.

Traditional notions that family life among slaves during the pre-plantation period in the non-Hispanic Caribbean was necessarily unstable are fading in light of new research. Although marriage among this segment of the population in Caguas, Cayey, San Germán, and Yauco--rural parishes in Puerto Rico--involved only a fraction of the overall number of marriages in these communities, the marriage of slaves was much more frequent than previously assumed. Family life among the eighteenth-century Puerto Rican slave population appears to have been quite stable, as shown by the reconstruction of birth intervals for both married and unmarried mothers. Married and unmarried mothers exhibited similar reproductive behavior. These results strongly suggest that a majority of the unmarried slave mothers lived in unions that were not institutionally recognized, but that were nevertheless stable, as indicated by the high percentage of their children born at intervals comparable to those of married mothers. If unmarried mothers were living in stable consensual unions, then our understanding of these slave family units during the colonial period must be reassessed not only for Puerto Rico but possibly for the rest of the Caribbean and Latin America.