Interaction of Mg-2+ with peptidoglycan and its relation to the prevention of lysis of a marine pseudomonad

Intact cells of marine pseudomonad B-16 (ATCC 19855) which have been washed with a solution of NaCl require only 0.001 M MgSO4 and 100 to 300 times this concentration of NaCl or KCl to prevent lysis. Conversion of intact cells to mureinoplasts, a process involving removal of the outer double-track layer (outer membrane) and the periplasmic space layer of the cell wall, approximately doubled the requirement for the three salts to prevent lysis. The formation of protoplasts from mureinoplasts by removing the peptidoglycan layer again doubled the requirement for Na+ and K+ salts but increased the requirement for the Mg-2+ salt 200- to 300-fold. Cells of the marine pseudomonad suspended in solutions containing Mg-2+ salts failed to lyse on subsequent repeated suspension in distilled water, whereas cells presuspended in NaCl lysed immediately. Isolated envelope layers including the peptidoglycan layer, when dialyzed against solutiions containing Mg-2+ salts, retained Mg-2+ after subsequent suspension in distilled water. Envelope layers exposed to solutions of Na+ or K+ salts failed to retain these ions after exposure to distilled water. Na+ displaced Mg-2+ from the cell envelope layers. The results obtained indicate that the capacity of Mg-2+ salts at very low concentration to prevent lysis of intact cells and mureinoplasts of this organism is due primarily to the interaction of Mg-2+ with the peptidoglycan layer of the cell wall. Ion interaction with the layers lying outside of the peptidoglycan layer contributes only a small amount to the mechanical strength of the wall.     

Pharmacological inhibition of protein kinases in intact cells: antagonism of beta adrenergic receptor ligand binding by H-89 reveals limitations of usefulness

The use of pharmacological inhibitors of protein kinases represents a potentially powerful tool in dissecting the regulatory features of intracellular signaling pathways. However, although the in vitro potency, selectivity, and efficacy of numerous kinase inhibitors have been characterized, little is known regarding the usefulness of these compounds as inhibitors in intact cells. In attempting to characterize the role of protein kinase A (PKA) in regulating the beta-2 adrenergic receptor (AR) in human airway cells, we observed a seemingly profound capacity of the isoquinoline H-89, a potent and widely used PKA inhibitor, to attenuate agonist-mediated desensitization of the beta-2 AR. Although additional experiments identified H-89 as an effective inhibitor of intracellular PKA, extended analysis of the compound determined the principal effect of H-89 was via its action as a beta-2 AR antagonist. Pretreatment with or the acute addition of H-89 significantly attenuated isoproterenol-stimulated cAMP accumulation. In cells pretreated with H-89 and then washed extensively, the subsequent dose-dependent response to isoproterenol suggested beta-2 AR antagonism by retained H-89. Competition binding of [125I]iodopindolol established Ki values of approximately 180 nM and 350 nM for H-89 antagonism of beta-2 AR and beta-1 AR, respectively. Additional receptor binding studies suggest selectivity of H-89 for the beta-2 AR and beta-1 AR, although a weak antagonism (Ki values of approximately 10 microM or greater) of other G protein-coupled receptors was observed. Results from additional pharmacological and biochemical analyses of various protein kinase inhibitors further established the need for careful characterization of pharmacological inhibitors when used in intact cell models.     

Television images and adolescent girls' body image disturbance

The impact of media images on adolescents' body image and thin ideal endorsement has been consistently asserted in the body image literature, yet has remained inadequately tested. I used social comparison theory and critical viewing with a sample of 214 high school girls to predict adolescents' body-image disturbance and thin ideal endorsement. Media variables accounted fr 15% of the variance for drive for thinness, 17% for body dissatisfaction, 16% for bulimic behaviors, and 33% for thin ideal endorsement. Results suggest that body image processing is the boy to understanding how television images affect adolescent girls' body-image attitudes and behaviors.     

A study of injected dose for brain mapping on the ECAT HR+: activation maps for a parametric verbal working memory task

The success of the 15O-water PET technique to localize statistically significant changes in regional cerebral blood flow is dependent on factors such as the activity level injected and the magnitude of the flow change. Undetectable changes may occur if insufficient activity is injected leading to high levels of statistical noise or the task performed results in only small changes in blood flow. To explore the relationship between injected activity and statistical significance, we performed a series of studies with the ECAT EXACT HR+, a high resolution PET tomograph. A parametric verbal working memory task (the N-back task) was selected to examine the relationship between regional cerebral blood flow and working memory load across a range of injected doses of 15O-water. At each activity level the volunteers were required to perform four different levels of the N-back task, a task in which a letter displayed on a monitor is matched with the letter displayed N letters previously. With increasing N, this task places increased load on working memory. For this study, 5, 10, and 15 mCi of 15O-water were injected into nine normal volunteers. The complete sequence of four tasks (N = 0, 1, 2, and 3) at three activity levels was repeated twice, for a total of 24 injections of 15O-water. We show that the peak count rate performance for the HR+ is approached at injected activity levels of 15O-water around 15 mCi. For this particular choice of N-back task, robust activation maps can nevertheless be obtained with as little as 5 mCi injected dose.     

Can duplex scan arterial mapping replace contrast arteriography as the test of choice before infrainguinal revascularization?

PURPOSE: Arteriography is the diagnostic test of choice before lower extremity revascularization, because it is a means of pinpointing stenotic or occluded arteries and defining optimal sites for the origin and termination of bypass grafts. We evaluated whether a duplex ultrasound scan, used as an alternative to arteriography, could be used as a means of accurately predicting the proximal and distal anastomotic sites in patients requiring peripheral bypass grafts and, therefore, replace standard preoperative arteriography. METHODS: Forty-one patients who required infrainguinal bypass grafts underwent preoperative duplex arterial mapping (DAM). Based on these studies, an observer blinded to the operation performed predicted what operation the patient required and the best site for the proximal and distal anastomoses. These predictions were compared with the actual anastomotic sites chosen by the surgeon. RESULTS: Whether a femoropopliteal or an infrapopliteal bypass graft was required was predicted correctly by means of DAM in 37 patients (90%). In addition, both anastomotic sites in 18 of 20 patients (90%) who had femoropopliteal bypass grafts and 5 of 21 patients (24%) who had infrapopliteal procedures were correctly predicted by means of DAM. CONCLUSION: DAM is a reliable means of predicting whether patients will require femoropopliteal or infrapopliteal bypass grafts, and, when a patient requires a femoropopliteal bypass graft, the actual location of both anastomoses can also be accurately predicted. Therefore, DAM appears able to replace conventional preoperative arteriography in most patients found to require femoropopliteal reconstruction. Patients who are predicted by means of DAM to require crural or pedal bypass grafts should still undergo preoperative contrast studies to confirm these results and to more precisely locate the anastomotic sites.