Electrical properties of cell pellets and cell electrofusion in a centrifuge.

A new approach is proposed for studying cell deformability by centrifugal force, electrical properties of cell membranes in a high electric field, and for performing efficient cell electrofusion. Suspensions of cells (L929 and four other cell types examined) are centrifuged in special chambers, thus forming compact cell pellets in the gap between the electrodes. The setup allows measurement of the pellet resistance and also the high-voltage pulse application during centrifugation. The pellet resistance increases sharply with the centripetal acceleration, which correlates with reduction of the cell pellet porosity due to cell compression and deformation. Experiments with cells pretreated with cytochalasin B or colcemid showed that cell deformability depends significantly on the state of cytoskeleton. When the voltage applied to the cell pellet exceeds a 'critical' value, electrical breakdown (poration) of cell membranes occurs. This is seen as a deflection in the I(V) curve for the cell pellet. The electropores formed during the breakdown reseal in several stages: the fastest takes 0.5-1 ms while the whole process completes in minutes. A novel effect of colloid-osmotic compression of cell pellets after electric cell permeabilization is described. Supercritical pulse application to the cell pellet during intensive centrifugation leads to massive cell fusion. The fusion index grows with the increase of centripetal acceleration, and drops drastically when the pulse is applied after the centrifuge is stopped. The colloid-osmotic pellet compression enhances the fusion efficiency. No fusion occurs when cells are brought in contact after the pulse treatment. The data suggest that tight intermembrane contact formed prior to pulse application is a prerequisite condition for efficient cell electrofusion. The capacities of the technique proposed and the mechanism of membrane electrofusion are discussed.

Electrofusion of fragile mutants of Saccharomyces cerevisiae.

This communication presents experimental evidence that intact fragile (osmotic sensitive) yeasts can be electrofused and give viable hybrids. The yield increases with one order of magnitude for electrofusion of intact fragile yeasts with protoplasts of non-fragile ones. The yield of viable hybrids, obtained by electrofusion of protoplasts of fragile and non-fragile yeasts, is one order of magnitude higher than the yield from protoplasts of non-fragile yeasts. The destabilized cell wall and plasma membrane of the mutant yeasts could be a possible explanation for this phenomenon.

Superimposing spatially coherent electromagnetic noise inhibits field-induced abnormalities in developing chick embryos.

Living cells exist in an electrically noisy environment. This has led to the so-called "signal-to-noise" problem whereby cells are observed to respond to extremely-low-frequency (ELF) exogenous fields that are several orders of magnitude weaker than local endogenous fields associated with thermal fluctuations. To resolve this dilemma, we propose that living cells are affected only by electromagnetic fields that are spatially coherent over their surface. The basic idea is that a significant number of receptors must be simultaneously and coherently activated (biological cooperativity) to produce effects on the biochemical functioning of the cell. However, like all physical detection systems, cells are subject to the laws of conventional physics and can be confused by noise. This suggests that a spatially coherent but temporally random noise field superimposed on a coherent ELF signal will defeat the mechanism of discrimination against noise, and any observed field-induced bioeffects would be suppressed. An experimental test of this idea was conducted using morphological abnormalities in developing chick embryos caused by electromagnetic field exposure as the endpoint. At an impressed noise amplitude comparable to the ELF field strength (but roughly one-thousandth of the thermal noise field), the increased abnormality rate observed with only the ELF field present was reduced to a level essentially the same as for the control embryos.

The effect of pulsed and sinusoidal magnetic fields on the morphology of developing chick embryos.

Several investigators have reported robust, statistically significant results that indicate that weak (approximately 1 microT) magnetic fields (MFs) increase the rate of morphological abnormalities in chick embryos. However, other investigators have reported that weak MFs do not appear to affect embryo morphology at all. We present the results of experiments conducted over five years in five distinct campaigns spanning several months each. In four of the campaigns, exposure was to a pulsed magnetic field (PMF); and in the final campaign, exposure was to a 60 Hz sinusoidal magnetic field (MF). A total of over 2500 White Leghorn chick embryos were examined. When the results of the campaigns were analyzed separately, a range of responses was observed. Four campaigns (three PMF campaigns and one 60 Hz campaign) exhibited statistically significant increases (P > or = 0.01), ranging from 2-fold to 7-fold, in the abnormality rate in MF-exposed embryos. In the remaining PMF campaign, there was only a slight (roughly 50%), statistically insignificant (P = 0.2) increase in the abnormality rate due to MF exposure. When the morphological abnormality rate of all of the PMF-exposed embryos was compared to that of all of the corresponding control embryos, a statistically significant (P > or = .001) result was obtained, indicating that PMF exposure approximately doubled the abnormality rate. Like-wise, when the abnormality rate of the sinusoid-exposed embryos was compared to the corresponding control embryos, the abnormality rate was increased (approximately tripled). This robust result indicates that weak EMFs can induce morphological abnormalities in developing chick embryos. We have attempted to analyze some of the confounding factors that may have contributed to the lack of response in one of the campaigns. The genetic composition of the breeding stock was altered by the breeder before the start of the nonresponding campaign. We hypothesize that the genetic composition of the breeding stock determines the susceptibility of any given flock to EMF-induced abnormalities and therefore could represent a confounding factor in studies of EMF-induced bioeffects in chick embryos.