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Biomedical subjects

A A Brayman

Publications and source records attributed to A A Brayman.

16 recordsLinked to original sources

Bubble cycling and standing waves in ultrasonic cell lysis.

Two quite different but potentially complementary hypothetical mechanisms have been proposed to explain the dependence of ultrasonic cell lysis in vitro on exposure vessel rotation. One mechanism proposes that resonant-size bubbles are trapped in planar arrays by standing waves in the sample and that exposure vessel rotation sweeps cells through the bubble arrays. The other mechanism, for which there is now considerable support, proposes that exposure vessel rotation simply allows bubbles to recycle after having been driven through the medium by the ultrasound. We report here the results of efforts to further discriminate between, and quantify, the relative contributions of these two mechanisms. Murine P388 cells were exposed to 1.07 MHz continuous-wave (CW) ultrasound for 5 min at 5 W/cm2 (ISP) using various exposure vessel compositions and exposure configurations. Experimental treatments were designed either to minimize or maximize standing waves in the sample tube, or to minimize the potential for bubble recycling while maximizing standing wave formation. The results of these experiments indicate that bubble cycling is responsible for the majority of cell lysis occurring in the rotating exposure vessels, but that a significant contribution to total lysis is provided by trapped bubbles under some conditions.

Animals

Modest enhancement of ultrasound-induced mutations in V79 cells in vitro.

V79 cells were insonated (1 MHz, 35 W/cm2, 2 min continuous wave) under temperature conditions previously reported to cause a large increase in acoustic cavitation. The goal was to increase the ease of detecting ultrasound-induced mutations. The data supported the hypothesis that a 3 degrees C gas-equilibration period followed by insonation at 37 degrees C resulted in a detectable increase in resistance to 6-thioguanine with 6 trials per regimen.

Animals

Apparent contribution of respiratory gas exchange to the in vitro "cell density effect" in ultrasonic cell lysis.

Ultrasonic damage to cells in vitro has been reported to vary dramatically with cell density; the particle density per se has been proposed to be responsible for this effect via interference with gas body activity. Reported here are the results of experiments designed to test the general postulate that the "cell density effect" can be explained in part by cellular modification of the suspension medium by respiratory gas exchange. Ultrasonic cell lysis is shown to occur at a much higher level in suspensions supplemented with the respiratory inhibitor NaCN than in control suspensions when suspension densities exceed 2 x 10(7) cells/mL. Moreover, at constant cell density, cell lysis diminishes with increasing pre-insonation incubation time at 37 degrees C. The rate of change in cell lysis with incubation time was diminished significantly by cyanide treatment. These observations are consistent with the postulate that respiratory O2:CO2 exchange enriches the medium in CO2 while depleting the medium of oxygen, which cavitates more readily than CO2, thereby diminishing the potential for cavitation-related cellular damage. However, this is only a partial explanation of the cell concentration dependence of cell lysis; cell density per se is an important factor in the "cell density effect."

Animals

Ultrasonically induced in vitro cell lysis: node-antinode interactions.

An attempt was made to discriminate between two hypotheses (standing wave, bubble recycling) of the mechanism of ultrasonically induced cell lysis in a rotating tube. A tube containing an aqueous suspension of P-388 cells was moved back and forth (+/- 3 or +/- 7 mm) during insonation (1 MHz, 5 W/cm2, continuous wave, 5 min). Cell lysis (approximately 20%) occurred. As a positive control, some tubes were also partially or completely rotated during insonation; considerable cell lysis (approximately 60%) occurred. The results are interpreted to suggest that both hypotheses are simultaneously useful in explaining the observed effect of cell lysis in a rotating tube.

Air

Failure to reproduce increased calcium uptake in human lymphocytes at purported cyclotron resonance exposure conditions.

An attempt was made to verify a report (Liboff et al. 1987) that a unique combination of DC and AC magnetic field exposures at room temperature results in a 3-fold increase in 45Ca2+ uptake by human lymphocytes in vitro. Exposures at "resonance condition", as well as at frequencies and amplitudes above and below the reported effective exposure conditions, were without effect, as were exposures at 37 degrees C. Treatment with ionomycin (0.25 microM), a positive control, resulted in a highly significant increase 45Ca2+ uptake. Some experiments were performed simultaneously by different investigators. Their results did not differ significantly. All experiments were conducted "double blindly".

Analysis of Variance

Proportionality of ELF electric field-induced growth inhibition to induced membrane potential in Zea mays and Vicia faba roots.

The postulate that 60-Hz electric field-induced bioeffect severity is proportional to induced transmembrane potential [Vmi] magnitude was tested and supported using a plant root model cell system. Statistically significant correlations were obtained upon regression of the relative rates of exposed Vicia faba and Zea mays root segment growth on the average Vmi (calculated) arising in those segments under specified 60 Hz field exposure conditions. The Vmi associated with the apparent threshold for growth inhibition was similar in Zea and Vicia roots (2.5 vs 2.4 mV, respectively). At Vmi greater than this threshold, Zea root growth declined by about 9% per mV, and Vicia root growth by about 19% per mV induced potential.

Electricity

60-Hz electric field exposure inhibits net apparent H(+)-ion excretion from excised roots of Zea mays L.

Plant root model cell systems have provided insight into the biophysical mechanism by which extremely low frequency electric fields (EF; f less than or equal to 100 Hz) affect nonexcitable eukaryotic cells. The evidence indicates that the plasma membrane is the site of interaction with applied extremely low frequency EF, and that cells respond to field exposure via a sensing mechanism involving the induction of extremely low frequency membrane potentials (Vim). We suggest a mechanism by which Vim may be transduced into EF-induced root growth inhibition. Suspensions of excised Zea root tips were used to test the hypothesis that growth-inhibiting extremely low frequency EF exposures inhibit net H+ excretion from protoplasts, a process mediated by a plasma membrane H(+)-ATPase which is intimately involved in cellular extension. Rates of acidification of root tip suspensions were measured as an analog for net H+ efflux. The experimental results support this hypothesis. At the apparent threshold for inhibition of H+ excretion, the associated 60-Hz EF strength was about 220 V.m-1 (root mean square). Estimates of Vim associated with inhibition of net H+ excretion are in agreement with those known to affect Na+/K+ transport in human erythrocytes.

Adenosine Triphosphatases

Repetitive pulsed-train "off" duration mitigates reductions in root growth rates of Pisum sativum L. induced by 60-Hz electric field.

An investigation was undertaken to define a 60-Hz electric field exposure system which would affect a eukaryotic cell system while mitigating a potential thermal rise. The biological effectiveness of pulsed 60-Hz electric fields on a cell system of defined sensitivity to continuous-wave 60-Hz electric fields was sought. Roots of garden pea (Pisum sativum L.) were exposed to pulsed trains of 60-Hz, 430 V/m electric fields. The "on" time was constant at 1 s and the "off" time varied. The repetitive on:off regimens used were 1:20, 1:50, 1:100, 1:130, 1:200, and 1:300. With continuous or 1:20 pulsed fields the growth response was equivalent (representing a 60% depression in root growth rate). The severity of the growth effect diminished as the off time increased; for the 1:100 regimen, the relative growth rate was depressed by about 30%; for the 1:300 regimen, the relative growth rate was equal to that of the controls.

Electromagnetic Fields

An explanation for the decrease in cell lysis in a rotating tube with increasing ultrasound intensity.

Previous observations indicate that for in vitro mammalian cells insonated in rotating test tube the amount of cell lysis initially increases to some maximum and then decreases with further increase in ultrasound exposure. The results of the present investigation support the postulate that the reduction in cell lysis with increase in ultrasound intensity is related to the development of an ultrasonically induced "cloud" of bubbles in the fluid between the transducer and test tube; these bubbles mitigate against acoustic transmission thus reducing cell lysis in the insonated test tube.

Animals

Proportionality of 60-Hz electric field bioeffect severity to average induced transmembrane potential magnitude in a root model system.

The postulate that electric field-induced bioeffects in the root model system are related to the induction of 60-Hz transmembrane potentials (Vim) was quantitatively tested. Root segment growth rate data and the calculated mean 60-Hz Vim which would arise in the cortical cells of a segment under specified exposure conditions were subjected to regression analysis. Statistically significant correlations between segmental growth rate and segmental-average Vim were obtained using data analyzed (1) within species at a constant applied field strength, (2) within species and pooled across field exposures, and (3) pooled across both species and exposures. In C. sativus roots, segmental growth is inhibited when segmental-average Vim attain a value of 3.4-3.6 mV. In C. maxima roots, growth inhibition occurs when Vim attain or exceed 2.3-2.7 mV. Segmental growth cessation is predicted to occur when segmental-average Vim exceed 7-9 mV.

Cell Physiological Phenomena

Lack of inductively-generated 60-Hz, 3 V.m-1 electric field effect on CHO cell plating efficiency.

Chinese hamster ovary (CHO) fibroblasts were exposed or sham-exposed to inductively-generated 60-Hz, 3.1 V.m-1 fields for 24 h. The absolute plating efficiency of the cells was determined immediately following the field exposure/sham-exposure, and after one week of post-exposure incubation. One-way analyses of variance performed on replicate experimental treatment means indicated no treatment effect on cell colony-forming ability.

Analysis of Variance

Weak low frequency electromagnetic fields and chick embryogenesis: failure to reproduce positive findings.

Fertilised chicken eggs were incubated for 48 hours while exposed to pulsed trains of magnetic fields having a duration of 0-5 ms, a rise time of 42 microsecond, and a pulse repetition rate of 100 at a magnetic field flux density of 1 microT. Some eggs were exposed to 1,552 rad X-rays as a positive control. After exposure the embryos were scored blind for a variety of morphological features. X-irradiated eggs displayed highly significant and repeatable anatomical alterations. There were no differences between magnetic field-exposed, sham-exposed and control eggs.

Animals

Effects of 60-Hz electric fields on cellular elongation and radial expansion growth in cucurbit roots.

Serial longitudinal and transverse sections were prepared from roots of Cucumis sativus and Cucurbita maxima that had been exposed/sham-exposed to 60-Hz electric fields for 0-2 days. Field exposures were selected to produce a 10-20% or a 70-80% growth inhibition in whole roots of both species. Cortical cell length and diameter were measured using a microscope and eyepiece micrometer; measurements were conducted "blind." In both species, inhibition of cellular elongation was associated with exposure to electric fields (EF). Cellular radial expansion was apparently unaffected by exposure to electric fields. The diameters of radially unexpanded or fully expanded C. sativus cortical cells were about 25-30% smaller than those of comparable cells in C. maxima roots. Previous studies of the relationship between rates of root growth and applied EF strength showed that the response thresholds of C. sativus and C. maxima differed by a similar relative amount. These results are consistent with the postulate that EF-induced effects in roots are elicited by induced transmembrane potentials.

Analysis of Variance

Induction of ELF transmembrane potentials in relation to power-frequency electric field bioeffects in a plant root model system. I. Relationship between applied field strength and cucurbitaceous root growth rates.

Seminal roots of Cucumis sativus and Cucurbita maxima were exposed to 60 Hz electric fields of 100-500 V X m-1 in a conducting aqueous inorganic growth medium. Root growth rates were measured to produce a dose-response relationship for each species. The species were selected for study because of their familial relationship, reported sensitivity to 60 Hz, 360 V X m-1 electric fields, and differing average root cell sizes. The latter characteristic influences the magnitude of ELF membrane potentials induced by constant-strength applied electric fields, but does not affect the magnitude of the electric field strength tangent to the cell surface. The difference in average root cell size between C. sativus (smaller cells) and C. maxima (larger cells) was used to evaluate two alternate hypotheses that the observed effect on root growth is stimulated by the electric field tangent to the cell surface, or a field-induced perturbation in the normal transmembrane potential of the cells. The results of the dose-response relationship studies are qualitatively consistent with the hypothesis that the effect is elicited by induced transmembrane potentials. The smaller-celled roots showed a substantially higher response threshold [C. sativus; E0TH approximately 330 V X m-1] than did the larger-celled species [C. maxima; E0TH approximately 200 V X m-1]. At field strengths above the response thresholds in both species, the growth rate of C. sativus roots was less affected than that of C. maxima roots exposed to the same field strength.

Cell Division

Induction of ELF transmembrane potentials in relation to power-frequency electric field bioeffects in a plant root model system. II. The effect of 60 Hz electric fields on the growth of different regions of the cucurbit root elongation zone.

The region of elongation in Cucumis sativus and Cucurbita maxima roots was marked at increasing distances from the apex to provide an analog of increasing cell size. These roots were exposed/sham-exposed to 60 Hz electric fields and the growth rates of the root segments measured. The growth rate effect magnitude varied with increasing distance from the root tip at constant field strength, and with increasing applied field strength. These results provide strong, qualitative support for the postulate that ELF transmembrane potential induction is involved in the stimulation of ELF electric field effects in the plant root model system.

Cell Division

The relationship between sensitivity to 60-Hz electric fields and induced transmembrane potentials in plants root cells.

Growth rates and cell diameters were determined from 12 species of plant roots exposed to a 60-Hertz (Hz) electric field of 360 Volts per meter (V/m) in an aqueous inorganic nutrient medium [conductivity: 0.07-0.09 Siemens per meter (S/m)]. The degree of growth depression ranged from zero to nearly 100 percent of control. Cell diameters ranged from 13.5 to 31.8 micron as an averaged value for procambial, cortical, and meristem cells. Sensitivity to the electric field as determined by root growth rate reduction increased with increasing cell size. Sensitivity also increased with increase in 60 Hz induced transmembrane potentials; the transmembrane potential threshold for growth reduction was about 6.0 mV and the potential for near-complete cessation of growth was about 10-11 mV. Two different hypothetical mechanisms of action by which applied electric fields induce biological effects at the cellular level were tested. The two mechanisms pertain to different possible modes of action of applied electric fields: one mechanism postulates the involvement of the transmembrane field, the other mechanism postulates the tangential electric field as the important factor for inducing biological effects. The data support the transmembrane and not the tangential field mechanism. It is concluded that the effects observed are consistent with a membrane related mechanism and that there is a narrow range (a few mV) between threshold and debilitating induced membrane potentials.

Electromagnetic Fields