Response properties of single units in an association area of the kitten neocortex

Two percept-genetic methods, the Meta-Contrast Technique (MCT) and the Serial Afterimage Test (AI), were used to study adaptive aspects of perception in a group of 37 subjects suffering from manifest anxiety. The perceptual processes laid bare by these methods reflected not only anxiety but also defensive formations. MCT and AI correlated with each other in these respects and, when combined, correlated with rated symptoms. In accordance with the theory behind percept-genetic methods, anxiety signs appearing late in the test serials were more likely to correspond to manifest symptoms than signs which appeared early. The AI test scores seemed to reflect a difference between anxiety experienced as an inherent quality of life and anxiety as a foreign force struggling for dominance.     

Two functional effects of decreased conductance EPSP's: synaptic augmentation and increased electrotonic coupling

Three electronically coupled motor neurons, which mediate inking behavior in Aplysia californica, receive both increased and decreased conductance excitatory postsynaptic potentials (EPSP's). The increased conductance EPSP's reduce electrical coupling among the cells, whereas the decreased conductance EPSP's increase electrical coupling. The decreased conductance EPSP's also augment the action of a previously ineffective sensory input and this augmentation is enhanced by the increase in electrical coupling. Both effects combine to trigger a stereotypic behavioral response.     

Time to clinical stability in patients hospitalized with community-acquired pneumonia: implications for practice guidelines

CONTEXT: Many groups have developed guidelines to shorten hospital length of stay in pneumonia in order to decrease costs, but the length of time until a patient hospitalized with pneumonia becomes clinically stable has not been established. OBJECTIVE: To describe the time to resolution of abnormalities in vital signs, ability to eat, and mental status in patients with community-acquired pneumonia and assess clinical outcomes after achieving stability. DESIGN: Prospective, multicenter, observational cohort study. SETTING: Three university and 1 community teaching hospital in Boston, Mass, Pittsburgh, Pa, and Halifax, Nova Scotia. PATIENTS: Six hundred eighty-six adults hospitalized with community-acquired pneumonia. MAIN OUTCOME MEASURES: Time to resolution of vital signs, ability to eat, mental status, hospital length of stay, and admission to an intensive care, coronary care, or telemetry unit. RESULTS: The median time to stability was 2 days for heart rate (< or =100 beats/min) and systolic blood pressure (> or =90 mm Hg), and 3 days for respiratory rate (< or =24 breaths/min), oxygen saturation (> or =90%), and temperature (< or =37.2 degrees C [99 degrees F]). The median time to overall clinical stability was 3 days for the most lenient definition of stability and 7 days for the most conservative definition. Patients with more severe cases of pneumonia at presentation took longer to reach stability. Once stability was achieved, clinical deterioration requiring intensive care, coronary care, or telemetry monitoring occurred in 1% of cases or fewer. Between 65% to 86% of patients stayed in the hospital more than 1 day after reaching stability, and fewer than 29% to 46% were converted to oral antibiotics within 1 day of stability, depending on the definition of stability. CONCLUSIONS: Our estimates of time to stability in pneumonia and explicit criteria for defining stability can provide an evidence-based estimate of optimal length of stay, and outline a clinically sensible approach to improving the efficiency of inpatient management.     

Apoptotic induction in transformed follicular lymphoma cells by Bcl-2 downregulation

The roles of Bcl-2 protein and the protein ratio of Bcl-2/Bax in regulating cell growth in various lymphoma cell lines were examined. A dose-dependent decrease in Bcl-2 protein expression was observed in the different lymphomas incubated with lipid-incorporated bcl-2 antisense oligonucleotides (L-bcl-2). Growth inhibition was observed in a transformed follicular lymphoma (FL) cell line, which has the t(14;18) translocation and Bcl-2 protein overexpression. One of the mechanisms by which L-bcl-2 growth inhibition is mediated in these transformed FL cells might be through apoptotic induction, because the treated cells had an increased apoptotic index and showed the typical DNA fragmentation. These studies indicate that Bcl-2 protein is critical in the growth regulation of transformed FL cells. L-bcl-2 did not induce growth inhibition in lymphoma cells not expressing Bcl-2 or Bax protein. Thus, the protein ratio of Bcl-2/Bax may also be important in regulating the growth of these lymphomas.     

Protein thermostability above 100 degreesC: a key role for ionic interactions

The discovery of hyperthermophilic microorganisms and the analysis of hyperthermostable enzymes has established the fact that multisubunit enzymes can survive for prolonged periods at temperatures above 100 degreesC. We have carried out homology-based modeling and direct structure comparison on the hexameric glutamate dehydrogenases from the hyperthermophiles Pyrococcus furiosus and Thermococcus litoralis whose optimal growth temperatures are 100 degreesC and 88 degreesC, respectively, to determine key stabilizing features. These enzymes, which are 87% homologous, differ 16-fold in thermal stability at 104 degreesC. We observed that an intersubunit ion-pair network was substantially reduced in the less stable enzyme from T. litoralis, and two residues were then altered to restore these interactions. The single mutations both had adverse effects on the thermostability of the protein. However, with both mutations in place, we observed a fourfold improvement of stability at 104 degreesC over the wild-type enzyme. The catalytic properties of the enzymes were unaffected by the mutations. These results suggest that extensive ion-pair networks may provide a general strategy for manipulating enzyme thermostability of multisubunit enzymes. However, this study emphasizes the importance of the exact local environment of a residue in determining its effects on stability.     

The Homothorax homeoprotein activates the nuclear localization of another homeoprotein, extradenticle, and suppresses eye development in Drosophila

The Extradenticle (Exd) protein in Drosophila acts as a cofactor to homeotic proteins. Its nuclear localization is regulated. We report the cloning of the Drosophila homothorax (hth) gene, a homolog of the mouse Meis1 proto-oncogene that has a homeobox related to that of exd. Comparison with Meis1 finds two regions of high homology: a novel MH domain and the homeodomain. In imaginal discs, hth expression coincides with nuclear Exd. hth and exd also have virtually identical, mutant clonal phenotypes in adults. These results suggest that hth and exd function in the same pathway. We show that hth acts upstream of exd and is required and sufficient for Exd protein nuclear localization. We also show that hth and exd are both negative regulators of eye development; their mutant clones caused ectopic eye formation. Targeted expression of hth, but not of exd, in the eye disc abolished eye development completely. We suggest that hth acts with exd to delimit the eye field and prevent inappropriate eye development.     

Structural and functional analogy between pneumolysin and proaerolysin

Pneumolysin and proaerolysin are bacterial toxins that form pores in host cells by oligomerization. We propose that they may have similar structures despite a poor sequence identity. The crystal structure of proaerolysin reveals a protein composed of four domains, arranged in the shape of an elongated comma. Electron microscopy of the pneumolysin monomer shows a similar arrangement of domains. The sequence of pneumolysin recognizes the template of proaerolysin from a library of protein folds. A three-dimensional model of pneumolysin has been constructed by the comparative approach using the structure of proaerolysin. This model, together with results on the activity of site- specific mutants and the positions of antigenic sites, has been used to propose functional roles of individual domains.      

Production of human normal adult and fetal hemoglobins in Escherichia coli

A hemoglobin expression system in Escherichia coli is described. In order to produce authentic human hemoglobin, we need to co-express both methionine aminopeptidase and globin genes under the control of a strong promoter. We have constructed three plasmids, pHE2, pHE4 and pHE7, for the expression of human normal adult hemoglobin and a plasmid, pHE9, for the expression of human fetal hemoglobin, in high yields. The globin genes can be derived from either synthetic genes or human globin cDNAs. The extra amino-terminal methionine residues of the expressed globins can be removed by the co-expressed methionine aminopeptidase. The heme is inserted correctly into the expressed alpha- globin from our expression plasmids. A fraction (approximately 25%) of the heme is not inserted correctly into the expressed beta- or gamma- globin. However, the incorrectly inserted hemes can be converted into the correct conformation by carrying out a simple oxidation-reduction process on the purified hemoglobin molecule. We have investigated the functional properties of the expressed hemoglobins by measuring their oxygen-binding properties and their structural features by obtaining their 1H-NMR spectra. Our results show that authentic human normal adult and fetal hemoglobins can be produced from our expression plasmids in E. coli and in high yields. Our expression system allows us to design and to produce any recombinant hemoglobins needed for our research on the structure-function relationship in hemoglobin.      

Foreign serum-induced pancreatitis in mice. II. Secretory disturbances of acinar cells

As previously reported, pancreatic acinar cell necrosis and inflammation develop in mice a few hours after one intraperitoneal injection of foreign serum. However, sublethally injured acinar cells exhibited notable increases in both zymogen granule numbers and amylase activity, observed within 3 hours and increasing with time. These two changes were coupled with a progressive decrease in the secretory response to pilocarpine and were preceded by significant disturbances in pancreatic tissue concentrations of sodium and potassium. We conclude that (1) the granule increase results from an induced disturbance of the granule exocytosis mechanism while granule formation continues and, therefore, (2) the granule secretory process is more sensitive to the serum injury mechanism than is the zymogen synthesis process. Although the granule increases developed in acinar cells throughout most of the nonnecrotic gland and persisted for at least 24 hours, acinar cell necrosis was maximal in extent--approximately 25% of the gland in severest form--by 12 to 15 hours. We conclude, therefore, that the increase in granules is neither the primary determinant nor initiator of acinar cell death. The latter is likely caused by disturbed plasma membrane functions, sufficient in some cells to result in lethal changes in ion and fluid composition. The injury mechanism, which permits granule formation to go on in the face of impaired granule exocytosis, is yet to be worked out. The possibilities are discussed in relationship to the reactivity of foreign sera for target cell plasma membranes.     

The effects of tolbutamide on the myocardium in experimental diabetes

The effects of chronic tolbutamide treatment were examined in a diabetic animal model in which abnormal myocardial function and composition have previously been demonstrated. Eight diabetic dogs were given tolbutamide 250 mg/day orally and compared with seven untreated diabetics, five healthy dogs receiving tolbutamide, and eight normal controls. After one year, resting hemodynamic studies in the intact anesthetized state showed that treated diabetic dogs had a significantly higher left ventricular end-diastolic pressure of 12.1+/-1.3 mm Hg associated with normal end-diastolic volume, compared to 6.1+/-0.8 mm Hg in untreated diabetics (P less than 0.01) and 6.3+/-0.5 in normals. Stroke work and ejection fraction were similar to normals. Acute volume expansion revealed a larger rise of left ventricular end-diastolic pressure in treated and untreated diabetics than normals, without a significant stroke volume response in treated diabetics. Enhanced stiffness of myocardium appeared to be related to interstitial accumulation of periodic acid-Schiff staining material, further intensified in treated diabetics by triglyceride accumulation observed on electron microscopy and by chemical analysis. Thus treatment of diabetes with tolbutamide, despite improved glucose tolerance, effected further reduction of left ventricular function and altered morphology of myocardium.