Cation Transport in Escherichia coli : VII. Potassium requirement for phosphate uptake

When Escherichia coli K-12 is grown in media containing limiting amounts of K, growth continues normally until all the extracellular K has been consumed. Thereafter the rates of growth, glucose consumption, and oxygen consumption decrease progressively, and the cell contents of K and P fall. These changes, referred to as K limitation, are all reversed by the addition of K. By specifically altering the ionic composition of the cells it was shown that these metabolic disturbances are not due to changes in the cell content of K or Na, but are directly related to the absence of K from the extracellular medium. The cell pool of inorganic P and the uptake of PO₄ from the medium are low in K-limited cells and are immediately stimulated by the addition of K, suggesting that the primary effect of K limitation is to inhibit PO₄ uptake. All the metabolic effects of K limitation can be attributed to inhibition of PO₄ uptake. The requirement of extracellular K for PO₄ uptake may be due to a coupling between the uptake of K and PO₄.     

Ion Transport in Isolated Rabbit Ileum : III. Chloride fluxes

Unidirectional Cl fluxes across in vitro segments of rabbit ileum have been determined both in the absence and in the presence of an electrochemical potential gradient. The results indicate that Cl transport in this preparation can be attributed to purely passive forces uninfluenced by solvent drag or exchange diffusion. Furthermore, on the basis of this and previous studies, it has been demonstrated that the sum of the partial ionic conductances of Na and Cl accounts for at least 90 per cent of the total tissue conductance.     

Kinetic relations of the Na-amino acid interaction at the mucosal border of intestine

The relation between unidirectional influxes of Na and amino acids across the mucosal border of rabbit ileum was studied under a variety of conditions. At constant Na concentration in the mucosal bathing solution, amino acid influx followed Michaelis-Menten kinetics permitting determination of maximal influx and the apparent Michaelis constant, K_t. Reduction in Na concentration, using choline as substitute cation, caused an increase in K_t for alanine but had no effect on maximal alanine influx. The reciprocal of K_t was a linear function of Na concentration. Similar results were obtained for valine and leucine and these amino acids competitively inhibited alanine influx both in the presence and in the absence of Na. These results lead to a model for the transport system which involves combination of Na and amino acid with a single carrier or site leading to penetration of both solutes. The model predicts that alanine should cause an increase in Na influx and the ratio of this extra Na flux to alanine flux should vary with Na concentration. The observed relation agreed closely with predicted values for Na concentrations from 5 to 140 mM. These results support the hypothesis that interactions between Na and amino acid transport depend in part on a common entry mechanism at the mucosal border of the intestine.     

Apparatus for monitoring the oxygen uptake and carbon dioxide production of fermentations

The requirements of the continuous analysis of effluent gas streams from aerated flash and tank fermentors are described, as are instrumental devices for measuring the oxygen and carbon dioxide content of fermentor gases. The use of a specially designed sequential gas sample for monitoring four fermentations simultaneously and a system for precise control of low air flow and pressure is explained. Equations for calculating carbon dioxide production or oxygen consumption rates and respiratory quotients are given. A discussion of the operating characteristics of a device for automatic translation of aeration data between fermentors is presented.     

Identification of Fluorescent-Antibody Labeled Group A Streptococci by Fluorometry

A quantitative system, amenable to automation, for determining the presence of group A streptococci in broth culture is described. After separation from the broth, the cells' protein coats are removed by digestion with 0.1% trypsin, and they are then stained with anti-A fluorescent antibody (FA). Excess FA is removed, and bound FA is put into solution by dissociation with demineralized distilled water. The amount of FA bound to the cells is quantitated by fluorometry of the solution. The level of nonspecific staining is measured by staining the cells with fluorescein-conjugated normal rabbit globulin absorbed with group A cells, dissociating, and quantifying, as above. The two quantities are subtracted to measure specific binding of FA to group A cells. A clinical trial showed 92% agreement with microscopists.     

Ultraviolet Radiation Effects on Ehrlich Ascites Tumor Cells : Observations Using a Flying Spot Ultraviolet Microscope

A flying spot ultraviolet microscope, employing a fast scan and pulsed operation of the raster, has been used to induce radiation damage in ascites tumor slide cultures, and to study by time-lapse cinematography the progressive stages of cell damage. The cells observed came from a strain (EF₇) of the Ehrlich ascites carcinoma. Irradiated cells were found to show a characteristic syndrome of damage, involving blebbing at the cell surface, while control cells in the adjacent areas of the preparation remained unchanged. The end of the blebbing period is marked by swelling of the cells, and the time taken for this phenomenon to occur was used as a measure of the severity of the damage. It was found that the time required for swelling is dependent on the size of the dose employed, as well as on the sensitivity of the cells. This latter sensitivity was found to decline as the physiological age of the tumor increased. If ultraviolet illumination below 255 mµ is excluded, no symptoms of damage occur, even when very large doses are used. These observations are discussed in relation to the nature of the system in the cell which is affected.     

Hindlimb suspension suppresses muscle growth and satellite cell proliferation

The effects of long-term hindlimb unweighting by tail suspension on postnatal growth of 20-day rat extensor digitorum longus (EDL) and soleus muscles were studied. Morphological assay indicated that radial growth of soleus myofibers was completely inhibited between 3 and 10 days of suspension and reduced thereafter, leading to a severe attenuation (-76% from control) over the total experimental period. Longitudinal growth rate, however, was accelerated 40% over weight-bearing controls. In addition, myofibers were arranged parallel to the long axis of the muscle, an orientation associated with chronologically younger muscles, suggesting morphological maturation of the soleus muscle had been delayed by suspension. In contrast, radial and longitudinal growth of EDL myofibers were minimally affected under similar conditions and remained within approximately 5% of control at all times. Suspension also influenced the normal changes that occur in satellite cell and myonuclear populations during postnatal growth. Both the number and proliferative activity of satellite cells were severely reduced in individual myofibers after only 3 days in both soleus and EDL muscles. The reduced number of satellite cells within 3 days of initiating hindlimb suspension appeared to be the result of their incorporation into myofibers while the long-lasting reduction appeared to be the added effects of decreased proliferative activity. In the soleus, this reduction in number and proliferation of satellite cells persisted throughout the experimental period and resulted in an overall 43% fewer myonuclei and 45% fewer satellite cells than control at 50 days of age. In contrast, both the total number and mitotic activity of satellite cells in the EDL rapidly returned to weight-bearing control levels by day 10 of suspension, resulting in no overall reduction in myonuclear accretion.     

A simple Hill element-nonlinear spring model of muscle contraction biomechanics

The purpose of this study was to develop a model to predict the mechanical response of muscles during isometric tetanic, afterloaded isotonic and isovelocity shortening contractions. Two versions of the model were developed. Both incorporated a contractile element that obeyed a Hill force-velocity relationship and a series elastic element. In a quadratic spring version, the series elastic element force was represented as proportional to the square of the stretch; in a cubic spring version, it was represented as proportional to the cube of the stretch. Both versions provided closed-form equations for response predictions that involved four independent parameters. Once the four parameters were chosen, each of these responses could be predicted. Model validity was established by comparing predicted and observed responses in slow and fast hindlimb muscles of rodents. Significant model-predicted responses seldom differed by more than 15% from experimental values. The model can provide insights into how changes in individual properties affect the overall mechanical behavior of muscles in a variety of circumstances and reduce the need for collection of experimental data.     

Pulmonary hemodynamic reaction to foreign blood in goats and rabbits.

We have found that the goat is extraordinarily sensitive to very small quantities of rabbit or rat blood. As little as 0.004 ml/kg induces transient pulmonary hypertension [maximal rise in pulmonary arterial pressure 32 +/- 10 (SD) cmH2O] in goats. We hypothesized that this reaction may be related to the presence of the resident population of intravascular macrophages that reside in the pulmonary capillaries of goats. If that is so, then rabbits or rats, which have few or no intravascular macrophages, should not be reactive to foreign blood. We compared pulmonary hemodynamics and changes in blood thromboxane B2 concentrations among goats, rabbits, and rats in response to graded doses of foreign blood. The pulmonary reaction to foreign blood was much greater in goats than in rabbits or rats, even though we injected up to 10- or 60-fold larger amounts into the latter species. In goats the pulmonary vascular pressure response to rabbit blood was dose dependent in goats and correlated well with changes in systemic arterial thromboxane B2 concentrations [change in pulmonary arterial pressure = 0.07 (thromboxane B2) + 8.3, r = 0.79]. We also tested the prostaglandin H2 endoperoxide analogue (U-46619) and found that the goats are somewhat more reactive than rabbits. We conclude that the pulmonary hemodynamic reaction to foreign blood is consistent with the concept that the foreign erythrocytes are reacting with the pulmonary intravascular macrophages in goats. The lower reactivity of the rabbit pulmonary circulation to thromboxane may also have a role.     

Crystallization of Escherichia coli catabolite gene activator protein with its DNA binding site. The use of modular DNA

To obtain crystals of the Escherichia coli catabolite gene activator protein (CAP) complexed with its DNA-binding site, we have searched for crystallization conditions with 26 different DNA segments greater than or equal to 28 base-pairs in length that explore a variety of nucleotide sequences, lengths, and extended 5' or 3' termini. In addition to utilizing uninterrupted asymmetric lac site sequences, we devised a novel approach of synthesizing half-sites that allowed us to efficiently generate symmetric DNA segments with a wide variety of extended termini and lengths in the large size range (greater than or equal to 28 bp) required by this protein. We report three crystal forms that are suitable for X-ray analysis, one of which (crystal form III) gives measurable diffraction amplitudes to 3 A resolution. Additives such as calcium, n-octyl-beta-D-glucopyranoside and spermine produce modest improvements in the quality of diffraction from crystal form III. Adequate stabilization of crystal form III is unexpectedly complex, requiring a greater than tenfold reduction in the salt concentration followed by addition of 2-methyl-2,4-pentanediol and then an increase in the concentration of polyethylene glycol.