Conceived and designed the experiments: DF. Performed the experiments: DF. Analyzed the data: DF. Contributed reagents/materials/analysis tools: DF. Wrote the paper: DF.
The author has declared that no competing interests exist.
Information transfer is a fundamental of life. A few studies have reported that cells use photons (from an endogenous source) as information carriers. This study finds that cells can have an influence on other cells even when separated with a glass barrier, thereby disabling molecule diffusion through the cell-containing medium. As there is still very little known about the potential of photons for intercellular communication this study is designed to test for non-molecule-based triggering of two fundamental properties of life: cell division and energy uptake. The study was performed with a cellular organism, the ciliate
Information transfer is a life principle. On a cellular level we generally assume that molecules are carriers of information, yet there is evidence for non-molecular information transfer due to endogenous coherent light
It was the paucity of more detailed knowledge on biophotons as a means for electromagnetic information transmission that motivated this study - to examine whether cell populations of the ciliate
In a series of experiments, population growth and the feeding rate of
All experiments were performed with the ciliate
Populations of
The graph shows the wavelengths where transmission through glass differs from transmission through quartz.
I performed three major experiments to test – indirectly – for effects of endogenous light on the growth and feeding rate of
Arrows pointing at the inner cuvette show the molecule barrier. Arrows going through the inner cuvette refer to photon transmission.
In the first experiment (1a) mutual influence between the two separated populations was tested, i.e. both populations were considered as sender and receiver. In the second experiment (2), inner populations were taken as senders and their effect on the outer receiving populations measured. In the third experiment, outer populations were taken as senders and their effect on inner receiving populations were measured (refer to the summary table:
Exp |
Testing on | Cells in∶out | Sender>receiver | Material | Effects |
1a | Cell division | 5∶100 | Inside>outside | Glass | Increase |
5∶100 | Inside>outside | Quartz | No | ||
5∶100 | Inside<outside | Glass | No | ||
5∶100 | Inside<outside | Quartz | Decrease |
||
2 | Cell division | 5∶25 | Inside>outside | Glass | No |
5∶25 | Inside>outside | Quartz | Decrease |
||
3 | Energy uptake | 15(20)∶15(20) | Inside<outside | Glass | Decrease |
15(20)∶300(400) | Inside<outside | Glass | Increase | ||
15(20)∶15(20) | Inside<outside | Quartz | Increase | ||
15(20)∶300(400) | Inside<outside | Quartz | Decrease |
The effects on receiver populations depend on the number of receiving and/or sender cells in the inner and outer cuvette (the so called units) as well as on the separating material (quartz or glass). In experiment 1a and 2 cell division (growth) was assessed 48 hrs after the mutual exposure of cell populations. In experiment 3 energy uptake (vacuole formation) of cells was assessed 3 hrs after the mutual exposure.
For experiment 1a see also
The significance of these effects follows from a
The aim of this experiment was to measure the effect of dense populations (initial size: 100 cells) in the outer cuvettes on the growth of small populations (initial size: 5 cells) in the inner cuvettes and
Twenty-eight blocks were assayed in 14 experimental sessions that were performed at different days. In each session two blocks were randomly placed on a four by five grid, where the units were optically separated from each other by a black carton. The grid itself was placed in a cardboard box. The populations were kept for 48 hours at constant room temperature, which was of 27°C in 10 sessions, 25°C, in one session, 23°C in one session and 22°C in two sessions. During the 48 hour period of growth a second cardboard box covered the one with the grid and was wrapped with a double-layered sheet of black cloth. This prevented external light from influencing the populations.
After 48 hours, each population was distributed into four sections of a glass well. The number of individuals per population was assessed with a hand counter. The mean of two or, if the difference between two counts was high, of three counts served as data points.
A potential problem with experiment 1a was that
This experiment assayed the effect of a small inner population on the growth of an outer population that was smaller than in experiment 1 (25 individuals). It tested, furthermore, for all possible combinations of the two cuvette materials. A block consisted, therefore, of 8 units: the inner cuvette made of glass or quartz (two possibilities), containing no or 5 cells (2 possibilities), combined with the outer glass or quartz cuvette (two possibilities). An experimental session consisted of two such blocks (i.e. 16 units in the grid) and was repeated 5 times at different days. The exposure period (under conditions as in experiment 1) was 48 hours before counting the number of individuals from each of the cuvettes.
Rather than considering the growth of a population, this experiment assayed the feeding rate of individuals, assessed as the number of vacuoles within an individual's cytoplasm. This demands the fixation of individual cells, a method described elsewhere
The main experiment is methodologically distinct from experiments 1 and 2 in two ways. First, it has a blind (random) design and second, the cells were not individually picked but population densities were assessed and fractions taken to obtain the desired cell densities. The effect of outer populations on inner populations was tested.
Two experiments were performed. In the first experiment 15 cells were in the inner cuvette and 15 or 300 cells in the outer cuvette. Furthermore, the material was for each unit the same, both inner and outer cuvette were either of glass or of quartz. This led to 4 units each of which was replicated four times within the experiment. The experimental block consisted, therefore in 16 units that were placed in the grid (as mentioned above). The second experiment differed from the first one slightly in cell numbers: in the inner cuvette there were always 20 cells, while there were 20 or 400 cells in the outer cuvette. Otherwise everything was kept as in the first experiment.
In both experiments the mutual exposure lasted for 3 hours. During exposure the paired populations were kept in a box as in experiments 1 and 2 on growth. When taking out the ciliates for fixation, the box was repeatedly opened and closed. Consequently, the later a unit was taken out of the box the more this unit would experience light from the laboratory illumination (In the second experiment this exposure to external light was kept at a minimum: the box was opened in a dark room with a few standby-lights of incubators only). For the analysis, however, the four replicates per treatment group were separated into the first two replicates (of each treatment that were taken from the random distribution in the box) and the last two replicates.
All experiments were tested with an ANOVA (or t-test for contrast analysis in experiments 1 and 2) using the statistical package JMP
The main results of effects of neighboring cell populations through glass are summarized in a table (
Both populations, the smaller in the inner cuvette and the larger in the outer cuvette, experienced strong effects from the day(s) of experimentation and from the cuvette material as well as from interactions between the two. More importantly, both day(s) of experimentation and also material interacted significantly with the mutual exposure of cell populations, i.e. the treatment (
The graph shows in the upper row the sizes of the larger outer populations and in the lower row those of the smaller inner populations. The filled squares refer to treatments using glass cuvettes, the open squares to those with quartz cuvettes. The x-axis shows the three treatment groups: paired are the combined (outer and inner) populations; C1 refers to the controls using medium instead of a second population and C2 is the control with demineralised water.
Source | DF | SS | F-ratio | P>F |
Day effects (day) | 13 | 7.495 | 22.511 | <0.0001**** |
Material (mat) | 1 | 1.291 | 50.389 | <0.0001**** |
Day×mat | 13 | 1.615 | 4.849 | <0.0001**** |
Treatment (treat) |
2 | 0.036 | 0.708 | 0.496 |
Day×treat | 26 | 1.154 | 1.733 | 0.032 |
Mat×treat |
2 | 0.228 | 4.447 | 0.015 |
Day×mat×treat | 26 | 0.948 | 1.424 | 0.116 |
(ANOVA: DF = degree(s) of freedom; SS = sum of squares).
Treatment refers to the presence or absence of a neighbouring population.
The important result is the interaction of treatment with separating material.
The (large) populations in the outer cuvette grew significantly better in glass than in quartz (independent from neighbours; statistics not shown; confer
Source | DF | SS | F-ratio | P>F |
Day effects (day) | 13 | 21.735 | 37.827 | <0.0001**** |
Material (mat) | 1 | 0.323 | 7.300 | 0.008*** |
Day×mat | 13 | 1.296 | 2.255 | 0.014 |
Treatment (treat) |
2 | 0.113 | 1.282 | 0.283 |
Day×treat | 26 | 1.151 | 1.001 | 0.048 |
Mat×treat |
2 | 0.292 | 3.303 | 0.042 |
Day×mat×treat | 26 | 1.431 | 1.245 | 0.225 |
(ANOVA: DF = degree(s) of freedom; SS = sum of squares).
Treatment refers to the presence or absence of a neighbouring population.
The important result is the interaction of treatment with separating material.
An
Testing for effects of fresh medium
This experiment on the effects of material and of the presence of cells in the inner cuvette on growth of cells in the outer cuvette revealed very strong material effects. When the outer cuvettes were of glass, growth of the outer population was significantly better than compared to outer populations in quartz cuvettes. The material of the inner cuvette did not affect this growth of the outer populations and there was no interaction between the materials of the inner and outer cuvettes. Also, the presence of cells in the inner cuvette (irrespective of material) did not affect growth either (
The graph shows the combined effects of material and presence or absence of cells in the inner cuvette on cell growth in the outer population. Filled squares refer to separation by glass and open squares to separation by quartz.
Source | DF | SS | F-ratio | p>F |
Day effects | 4 | 18.73 | 90.27 | >0.0001**** |
Material of cuvette outside (mat-out) | 1 | 0.76 | 14.74 | 0.0003*** |
Material of cuvette inside (mat-in) | 1 | 0.06 | 1.16 | 0.2848 |
Cells inside (cells-in) | 1 | 0.11 | 2.03 | 0.1591 |
Mat-out×mat-in | 1 | 0.08 | 1.60 | 0.2105 |
Mat-out×cells-in | 1 | 0.01 | 0.16 | 0.6861 |
Mat-in×cells-in |
1 | 0.31 | 5.93 | 0.0175* |
Mat-out×mat-in×cells-in | 1 | 0.17 | 3.22 | 0.0773 |
(ANOVA: DF = degree(s) of freedom; SS = sum of squares).
The important result is the interaction of material with (presence or absence of) cells.
An
The upper graph represents the significantly better growing paired populations separated by glass while the lower graph refers to the reduced growth of paired populations separated by quartz (compare
Pooling the two experiments and taking the subset of the first two replicates (i.e., those taken first out of the black box), it turned out that the number of vacuoles produced by the ciliates was significantly higher for the ciliates in glass cuvettes. There was no overall treatment effect. However, the interaction of material and treatment showed a highly significant effect on vacuole formation (
The population size in the outer cuvette reflected either the size of the population in the inner cuvette (15 or 20) or a 20-fold of it (300 or 400). Filled squares refer to separation with glass, open squares to separation with quartz.
Source | DF | SS | F-ratio | p>F |
Material | 1 | 2.078 | 5.475 | 0.021 |
Treatment |
1 | 0.039 | 0.104 | 0.748 |
Material×treatment |
1 | 10.357 | 27.284 | <0.0001**** |
(ANOVA: DF = degree(s) of freedom; SS = sum of squares).
Treatment refers to many or few cells in the outer cuvette.
The important result is the interaction of material with cell-number outside.
In the present study, three major experiments confirmed that separated populations of the ciliate
Comparing these results with corresponding studies on onion roots
Are these interactions based on biophotons? Clearly this was not measured and to my knowledge is not currently measurable, for the electromagnetic spatial dimension of cells is so far not reachable either for precise assessment or application. But there is strong indirect evidence for biophotons as information carriers coming from the separation of cell populations through the use of quartz or glass (a filter for UV-light below 340 nm, see above), which produces different results. If only frequencies above 340 nm were used, we should expect to find no differences between populations separated by glass or quartz, because both materials allow the transmission of these frequencies. Yet there were such differences. Likewise, if only frequencies below 340 nm were to be used, an effect on growth for populations separated by glass should not be observed, but there were effects. Consequently, one can deduce that at least two frequencies are in use, one above 340 nm and the other one below 340 nm (note that below 340 nm, transmission through glass can still occur but in decreased percentages as shown in
There was a material effect on the outer populations with glass resulting in better growth or energy uptake as compared to quartz. It is not within the scope of this study to address why this was the case. More important, however, is the fact that these pure material effects do not explain the neighbourhood effects, for they were repeatedly opposed to each other.
The evidence for electromagnetic information transfer is strong, and it is difficult to think of alternative mechanisms that could have produced similar results. One possibility is that molecules in a gaseous state left the cuvettes and influenced neighbouring populations. But this appears improbable because these molecules would have to diffuse only into one neighbouring cuvette (i.e. the right one). Note that for an inner cuvette the outer cuvette is indeed the nearest one, while for an outer cuvette, the nearest cuvette is the one standing next in the grid, i.e., the larger cuvette of another unit. This is so because of the differing heights of the smaller inner cuvette (45 mm) and the larger outer cuvette (40 mm). Furthermore, due to the random design, the treatments differ between neighbouring units in the grid. In addition, such molecules would have to be specific to the material of the cuvette and to the number of cells they originated from. If they existed, however, they would most probably produce a mixture in the microclimate (recall that the mutual exposure lasted for 48 hrs in exp. 1a and 2) that would be more or less the same for all cuvettes into which they might diffuse, and consequently lead to results that are independent from the treatments. As the results were very distinct between treatment groups, it is unlikely that such molecules were present.
Another alternative explanation for effects from the neighbouring cells would be heat production, i.e. infrared waves caused by cell metabolism. Once again however, there is the problem of omni-directional results: cells in the neighbouring cuvette would experience both enhancing and reducing effects on growth. Furthermore, the cells were exposed at 27°C and it seems doubtful that the small cells could heat up the water in the neighbouring cuvette due to metabolism. Finally, within the single frequency (above 340 nm) the results should not show differences between quartz and glass cuvettes (see argument above), yet, they do.
The goal of this study was to look for the potential of endogenous photons to act as triggering signals: under the conditions of the experiment an information transfer was indeed discerned. It is very probably due to photons emitted by cells, hence biophotons. Since the cells can communicate between populations separated by glass as described in this study, one may deduce that the cells do also communicate within a population, i.e. between cells. Cells, in addition to being a world of molecules, are also a world of electromagnetic fields that play major roles in morphogenesis of multicellular organisms
Cells can influence each other without using a molecular signal for the purpose: this means that not all cellular processes are necessarily based on a molecule-receptor recognition. The non-molecular signals are most probably photons. If so, cells use more than one frequency for information transfer and mutual influence. The effects are manifold, acting positively or negatively on cell growth, correlated growth and energy uptake. Since there are already existing reports of the induction of chemical reactions through glass
The author is greatly thankful to Marcel Tanner, head of the Swiss Tropical Institute, for granting required laboratory space, to Fritz-Albert Popp for repeated motivation, to Oliver Kaltz for discussing the performance of advanced experiments at the CNRS in Paris (France). Special thanks go to Roland Steiner from the Institute of Physics in Basel (Switzerland) for the measurements of the cuvette material and most of all to Master Zhang for producing excellent cuvettes in Beijing (China). A great thank to Andrea Mayer for advice in statistics, to Frank Hirth for discussing the experimental design and to Jacob Koella, Jean Nordmann and Nigel Powell for important comments on drafts of the manuscript.