Effects of low frequency rectangular electric pulses on Trichoplax (Placozoa)

Main Article Content

A. V. Kuznetsov

leading researcher, D. Sc.

http://orcid.org/0000-0002-0015-7994

https://elibrary.ru/author_items.asp?id=1043295

O. N. Kuleshova

junior researcher

http://orcid.org/0000-0003-3745-7066

https://elibrary.ru/author_items.asp?id=827844

A. Yu. Pronozin

junior researcher

http://orcid.org/0000-0002-3011-6288

https://elibrary.ru/author_items.asp?id=1038772

O. V. Krivenko

leading researcher, PhD

http://orcid.org/0000-0001-6292-5293

https://elibrary.ru/author_items.asp?id=933995

O. S. Zavyalova

head of department, PhD

http://orcid.org/0000-0003-2539-8927

https://elibrary.ru/author_items.asp?id=853234

Abstract

The effect of extremely low frequency electric and magnetic fields (ELF-EMF) on plants and animals including humans is quite a contentious issue. Little is known about ELF-EMF effect on hydrobionts, too. We studied the effect of square voltage waves of various amplitude, duration, and duty cycle, passed through seawater, on Trichoplax organisms as a possible test laboratory model. Three Placozoa strains, such as Trichoplax adhaerens (H1), Trichoplax sp. (H2), and Hoilungia hongkongensis (H13), were used in experiments. They were picked at the stationary growth phase. Arduino Uno electronics platform was used to generate a sequence of rectangular pulses of given duration and duty cycle with a frequency up to 2 kHz. Average voltage up to 500 mV was regulated by voltage divider circuit. Amlodipine, an inhibitor of calcium channel activity, was used to check the specificity of electrical pulse effect on voltage-gated calcium channels in Trichoplax. Experimental animals were investigated under a stereo microscope and stimulated by current-carrying electrodes placed close to a Trichoplax body. Variations in behavior and morphological characteristics of Trichoplax plate were studied. Stimulating and suppressing effects were identified. Experimental observations were recorded using photo and video techniques. Motion trajectories of individual animals were tracked. Increasing voltage pulses with fixed frequency of 20 Hz caused H2 haplotype individuals to leave “electrode zone” within several minutes at a voltage of 25 mV. They lost mobility in proportion to voltage rise and were paralyzed at a voltage of 500 mV. Therefore, a voltage of 50 mV was used in further experiments. An animal had more chance to move in various directions in experiments with two electrodes located on one side instead of both sides of Trichoplax. Direction of motion was used as a characteristic feature. Trichoplax were observed to migrate to areas with low density of electric field lines, which are far from electrodes or behind them. Animals from old culture were less sensitive to electrical stimulus. H2 strain was more reactive than H1 strain and especially than H13 strain; it demonstrated stronger physiological responses at frequencies of 2 Hz and 2 kHz with a voltage of 50 mV. Motion patterns and animal morphology depended on the duration of rectangular stimulation pulses, their number, amplitude, and frequency. Effects observed varied over a wide range: from direct or stochastic migration of animals to the anode or the cathode or away from it to their immobility, an increase of optical density around and in the middle of Trichoplax plate, and finally to Trichoplax folding and detach from the substrate. Additional experiments on Trichoplax sp. H2 with pulse duration of 35 ms and pulse delay of 1 ms to 10 s showed that the fraction of paralyzed animals increased up to 80 % with minimum delay. Nevertheless, in the presence of amlodipine with a concentration of 25 nM, almost all Trichoplax remained fast-moving for several minutes despite exposure to voltage waves. Experimental animals showed a total discoordination of motion and could not leave an “electrode trap”, when amlodipine with a concentration of 250 nM was used. Further, Trichoplax plate became rigid, which appeared in animal shape invariability during motion. Finally, amlodipine with a concentration of 50 μM caused a rapid folding of animal plate-like body into a pan in the ventral-dorsal direction and subsequent dissociation of Trichoplax plate into individual cells. In general, the electrical exposure applied demonstrated a cumulative but a reversible physiological effect, which, as expected, is associated with activity of voltage-gated calcium channels. Amlodipine at high concentration (50 μM) caused Trichoplax disintegration; at moderate concentration (250 nM), it disrupted the propagation of activation waves that led to discoordination of animal motion; at low concentration (25 nM), it prevented an electric shock.

Article Details

Kuznetsov A. V., Kuleshova O. N., Pronozin A. Yu., Krivenko O. V., Zavyalova O. S. Effects of low frequency rectangular electric pulses on Trichoplax (Placozoa). Marine Biological Journal, 2020, vol. 5, no. 2, pp. 50-66. doi: 10.21072/mbj.2020.05.2.05
Keywords:
rectangular electric pulses, Trichoplax, Placozoa, voltage-gated calcium channels
Section
Scientific communications

References

Albertini M. C., Fraternale D., Semprucci F., Cecchini S., Colomba M., Rocchi M. B. L., Sisti D., Di Giacomo B., Mari M., Sabatini L., Cesaroni L., Balsamo M., Guidi L. Bioeffects of Prunus spinosa L. fruit ethanol extract on reproduction and phenotypic plasticity of Trichoplax adhaerens Schulze, 1883 (Placozoa). PeerJ, 2019, vol. 7, article e6789 (22 p.). https://doi.org/10.7717/peerj.6789

Armon S., Bull M. S., Aranda-Diaz A., Prakash M. Ultrafast epithelial contractions provide insights into contraction speed limits and tissue integrity. Proceedings of the National Academy of Science of the USA, 2018, vol. 115, no. 44, pp. E10333–E10341. https://doi.org/10.1073/pnas.1802934115

Bers D. M., Perez-Reyes E. Ca channels in cardiac myocytes: Structure and function in Ca influx and intracellular Ca release. Cardiovascular Research, 1999, vol. 42, iss. 2, pp. 339–360. https://doi.org/10.1016/S0008-6363(99)00038-3

Catterall W. A. Voltage-gated calcium channels. Cold Spring Harbor Perspectives in Biology, 2011, vol. 3, iss. 8, article 003947 (23 p.). https://doi.org/10.1101/cshperspect.a003947

Chandler N. J., Greener I. D., Tellez J. O., Inada S., Musa H., Molenaar P., Difrancesco D., Baruscotti M., Longhi R., Anderson R. H., Billeter R., Sharma V., Sigg D. C., Boyett M. R., Dobrzynski H. Molecular architecture of the human sinus node: Insights into the function of the cardiac pacemaker. Circulation, 2009, vol. 119, no. 12, pp. 1562–1575. https://doi.org/10.1161/CIRCULATIONAHA.108.804369

d’Alessandro J., Mas L., Aubry L., Rieu J. P., Rivière C., Anjard C. Collective regulation of cell motility using an accurate density-sensing system. Journal of the Royal Society Interface, 2018, vol. 15, iss. 140, article 20180006 (11 p.). https://doi.org/10.1098/rsif.2018.0006

DuBuc T. Q., Ryan J. F., Martindale M. Q. “Dorsal-Ventral” genes are part of an ancient axial patterning system: Evidence from Trichoplax adhaerens (Placozoa). Molecular Biology and Evolution, 2019, vol. 36, iss. 5, pp. 966–973. https://doi.org/10.1093/molbev/msz025

Eitel M., Francis W. R., Varoqueaux F., Daraspe J., Osigus H. J., Krebs S., Vargas S., Blum H., Williams G. A., Schierwater B., Wörheide G. Correction: Comparative genomics and the nature of placozoan species. PLoS Biology, 2018, vol. 16, no. 9, article e3000032 (1 p.). https://doi.org/10.1371/journal.pbio.2005359

Eitel M., Francis W. R., Varoqueaux F., Daraspe J., Osigus H. J., Krebs S., Vargas S., Blum H., Williams G. A., Schierwater B., Wörheide G. Comparative genomics and the nature of placozoan species. PLoS Biology, 2018, vol. 16, no. 7, article E2005359 (36 p.). https://doi.org/10.1371/journal.pbio.2005359

Gao R., Zhao S., Jiang X., Sun Y., Zhao S., Gao J., Borleis J., Willard S., Tang M., Cai H., Kamimura Y., Huang Y., Jiang J., Huang Z., Mogilner A., Pan T., Devreotes P. N., Zhao M. A large-scale screen reveals genes that mediate electrotaxis in Dictyostelium discoideum. Science Signaling, 2015, vol. 8, iss. 378, pp. ra50 (10 p.). https://doi.org/10.1126/scisignal.aab0562

Godfraind T. Discovery and development of calcium channel blockers. Frontiers in Pharmacology, 2017, vol. 8, article 286 (25 p.). https://doi.org/10.3389/fphar.2017.00286

Grassi C., D’Ascenzo M., Torsello A., Martinotti G., Wolf F., Cittadini A., Azzena G. B. Effects of 50 Hz electromagnetic fields on voltage-gated Ca2+ channels and their role in modulation of neuroendocrine cell proliferation and death. Cell Calcium, 2004, vol. 35, iss. 4, pp. 307–315. https://doi.org/10.1016/j.ceca.2003.09.001

Heyland A., Croll R., Goodall S., Kranyak J., Wyeth R. Trichoplax adhaerens, an enigmatic basal metazoan with potential. In: Developmental Biology of the Sea Urchin and Other Marine. Invertebrates: Methods and Protocols / D. J. Carroll, S. A. Stricker (Eds). New York : Humana Press, 2014, chap. 4, pp. 45–61. https://doi.org/10.1007/978-1-62703-974-1_4

Iftinca M. C. Neuronal T-type calcium channels: What’s new? Journal of Medicine and Life, 2011, vol. 4, iss. 2, pp. 126–138.

Kamm K., Osigus H. J., Stadler P. F., DeSalle R., Schierwater B. Trichoplax genomes reveal profound admixture and suggest stable wild populations without bisexual reproduction. Scientific Reports, 2018, vol. 8, iss. 1, article 11168 (11 p.). https://doi.org/10.1038/s41598-018-29400-y

Kopecky B. J., Liang R., Bao J. T-type calcium channel blockers as neuroprotective agents. Pflügers Archiv – European Journal of Physiology, 2014, vol. 466, iss. 4, pp. 757–765. https://doi.org/10.1007/s00424-014-1454-x

Kuznetsov A. V., Halaimova A. V., Ufimtseva M. A., Chelebieva E. S. Blocking a chemical communication between Trichoplax organisms leads to their disorderly movement. International Journal of Parallel, Emergent and Distributed Systems, 2020, vol. 35, iss. 4, pp. 473–482. https://doi.org/10.1080/17445760.2020.1753188

Lawrence A. F., Adey W. R. Nonlinear wave mechanisms in interactions between excitable tissue and electromagnetic fields. Neurological Research, 1982, vol. 4, iss. 1–2, pp. 115–153. https://doi.org/10.1080/01616412.1982.11739619

Ledoigt G., Belpomme D. Cancer induction molecular pathways and HF-EMF irradiation. Advances in Biological Chemistry, 2013, vol. 3, pp. 177–186. https://doi.org/10.4236/abc.2013.32023

Linz K. W., Meyer R. Control of L-type calcium current during the action potential of guinea-pig ventricular myocytes. Journal of Physiology, 1998, vol. 513, pt. 2, pp. 425–442. https://doi.org/10.1111/j.1469-7793.1998.425bb.x

Lipscombe D., Andrade A. Calcium channel CaVα1 splice isoforms – Tissue specificity and drug action. Current Molecular Pharmacology, 2015, vol. 8, iss. 1, pp. 22–31. https://doi.org/10.2174/1874467208666150507103215

Marchionni I., Paffi A., Pellegrino M., Liberti M., Apollonio F., Abeti R., Fontana F., D’Inzeo G., Mazzanti M. Comparison between low-level 50 Hz and 900 MHz electromagnetic stimulation on single channel ionic currents and on firing frequency in dorsal root ganglion isolated neurons. Biochimica et Biophysica Acta (BBA) – Biomembranes, 2006, vol. 1758, iss. 5, pp. 597–605. https://doi.org/10.1016/j.bbamem.2006.03.014

Moran Y., Barzilai M. G., Liebeskind B. J., Zakon H. H. Evolution of voltage-gated ion channels at the emergence of Metazoa. Journal of Experimental Biology, 2015, vol. 218, pt. 4, pp. 515–525. https://doi.org/10.1242/jeb.110270

Nanou E., Catterall W. A. Calcium channels, synaptic plasticity, and neuropsychiatric disease. Neuron, 2018, vol. 98, iss. 3, pp. 466–481. https://doi.org/10.1016/j.neuron.2018.03.017

Pall M. L. Electromagnetic fields act via activation of voltage-gated calcium channels to produce beneficial or adverse effects. Journal of Cellular and Molecular Medicine, 2013, vol. 17, iss. 8, pp. 958–965. https://doi.org/10.1111/jcmm.12088

Pall M. L. Microwave frequency electromagnetic fields (EMFs) produce widespread neuropsychiatric effects including depression. Journal of Chemical Neuroanatomy, 2016, vol. 75, pt. B, pp. 43–51. https://doi.org/10.1016/j.jchemneu.2015.08.001

Ritter J. M., Flower R. J., Henderson G., Loke Y. K., MacEwan D., Rang H. P. Rang & Dale’s Pharmacology. 9th edition. Elsevier, 2019, 808 p.

Ruthmann A., Terwelp U. Disaggregation and reaggregation of cells of the primitive metazoon Trichoplax adhaerens. Differentiation, 1979, vol. 13, iss. 3, pp. 185–198. https://doi.org/10.1111/j.1432-0436.1979.tb01581.x

Santini M. T., Rainaldi G., Indovina P. L. Cellular effects of extremely low frequency (ELF) electromagnetic fields. International Journal of Radiation Biology, 2009, vol. 85, iss. 4, pp. 294–313. https://doi.org/10.1080/09553000902781097

Schierwater B. My favorite animal, Trichoplax adhaerens. BioEssays, 2005, vol. 27, iss. 12, pp. 1294–1302. https://doi.org/10.1002/bies.20320

Schulze F. E. Trichoplax adhaerens, nov. gen., nov. spec. Zoologischer Anzeiger, 1883, vol. 6, pp. 92–97.

Senatore A., Raiss H., Le P. Physiology and evolution of voltage-gated calcium channels in early diverging animal phyla: Cnidaria, Placozoa, Porifera, and Ctenophora. Frontiers in Physiology, 2016, vol. 7, article 481. https://doi.org/10.3389/fphys.2016.00481

Shiels H. A., Vornanen M., Farrell A. P. Temperature dependence of cardiac sarcoplasmic reticulum function in rainbow trout myocytes. Journal of Experimental Biology, 2002, pt. 23, pp. 3631–3639.

Shiels H. A., Vornanen M., Farrell A. P. Temperature-dependence of L-type Ca2+ channel current in atrial myocytes from rainbow trout. Journal of Experimental Biology, 2000, pt. 18, pp. 2771–2780.

Simms B. A., Zamponi G. W. Neuronal voltage-gated calcium channels: Structure, function, and dysfunction. Neuron, 2014, vol. 82, iss. 1, pp. 24–45. https://doi.org/10.1016/j.neuron.2014.03.016

Smith C. L., Abdallah S., Wong Y. Y., Le P., Harracksingh A. N., Artinian L., Tamvacakis A. N., Rehder V., Reese T. S., Senatore A. Evolutionary insights into T-type Ca2+ channel structure, function, and ion selectivity from the Trichoplax adhaerens homologue. Journal of General Physiology, 2017, vol. 149, iss. 4, pp. 483–510. https://doi.org/10.1085/jgp.201611683

Smith C. L., Pivovarova N., Reese T. S. Coordinated feeding behavior in Trichoplax, an animal without synapses. PLoS One, 2015, vol. 10, iss. 9, article e0136098 (15 p.). https://doi.org/10.1371/journal.pone.0136098

Smith C. L., Reese T. S., Govezensky T., Barrio R. A. Coherent directed movement toward food modeled in Trichoplax, a ciliated animal lacking a nervous system. Proceedings of the National Academy of Science of the USA, 2019, vol. 116, no. 18, pp. 8901–8908. https://doi.org/10.1073/pnas.1815655116

Smith C. L., Varoqueaux F., Kittelmann M., Azzam R. N., Cooper B., Winters C. A., Eitel M., Fasshauer D., Reese T. S. Novel cell types, neurosecretory cells, and body plan of the early-diverging metazoan Trichoplax adhaerens. Current Biology, 2014, vol. 24, iss. 14, pp. 1565–1572. https://doi.org/10.1016/j.cub.2014.05.046

Srivastava M., Begovic E., Chapman J., Putnam N. H., Hellsten U., Kawashima T., Kuo A., Mitros T., Salamov A., Carpenter M. L., Signorovitch A. Y., Moreno M. A., Kamm K., Grimwood J., Schmutz J., Shapiro H., Grigoriev I. V., Buss L. W., Schierwater B., Dellaporta S. L., Rokhsar D. S. The Trichoplax genome and the nature of placozoans. Nature, 2008, vol. 454, no. 7207, pp. 955–960. https://doi.org/10.1038/nature07191

Sun Z. C., Ge J. L., Guo B., Guo J., Hao M., Wu Y. C., Lin Y. A., La T., Yao P. T., Mei Y. A., Feng Y., Xue L. Extremely low frequency electromagnetic fields facilitate vesicle endocytosis by increasing presynaptic calcium channel expression at a central synapse. Scientific Reports, 2016, vol. 6, article 21774 (11 p.). https://doi.org/10.1038/srep21774

Varoqueaux F., Williams E. A., Grandemange S., Truscello L., Kamm K., Schierwater B., Jékely G., Fasshauer D. High cell diversity and complex peptidergic signaling underlie placozoan behavior. Current Biology, 2018, vol. 28, iss. 21, pp. 3495–3501 (10 p.). https://doi.org/10.1016/j.cub.2018.08.067

Vianale G., Reale M., Amerio P., Stefanachi M., Di Luzio S., Muraro R. Extremely low frequency electromagnetic field enhances human keratinocyte cell growth and decreases proinflammatory chemokine production. British Journal of Dermatology, 2008, vol. 158, iss. 6, pp. 1189–1196. https://doi.org/10.1111/j.1365-2133.2008.08540.x

Warille A. A., Altun G., Elamin A. A., Kaplan A. A., Mohamed H., Yurt K. K., El Elhaj A., Skeptical approaches concerning the effect of exposure to electromagnetic fields on brain hormones and enzyme activities. Journal of Microscopy and Ultrastructure, 2017, vol. 5, iss. 4, pp. 177–184. https://doi.org/10.1016/j.jmau.2017.09.002

Weiss N., Zamponi G. W. Control of low-threshold exocytosis by T-type calcium channels. Biochimica et Biophysica Acta (BBA) – Biomembranes, 2013, vol. 1828, iss. 7, pp. 1579–1586. https://doi.org/10.1016/j.bbamem.2012.07.031

Weiss N., Zamponi G. W. Genetic T-type calcium channelopathies. Journal of Medical Genetics, 2019, vol. 57, iss. 1, pp. 1–10. https://doi.org/10.1136/jmedgenet-2019-106163

Xiang Z., Thompson A. D., Brogan J. T., Schulte M. L., Melancon B. J., Mi D., Lewis L. M., Zou B., Yang L., Morrison R., Santomango T., Byers F., Brewer K., Aldrich J. S., Yu H., Dawson E. S., Li M., McManus O., Jones C. K., Daniels J. S., Hopkins C. R., Xie X. S., Conn P. J., Weaver C. D., Lindsley C. W. The discovery and characterization of ML218: A novel, centrally active T-type calcium channel inhibitor with robust effects in STN neurons and in a rodent model of Parkinson’s disease. ACS Chemical Neuroscience, 2011, vol. 2, iss. 12, pp. 730–742. https://doi.org/10.1021/cn200090z

Zalata A., El-Samanoudy A. Z., Shaalan D., El-Baiomy Y., Mostafa T. In vitro effect of cell phone radiation on motility, DNA fragmentation, and clusterin gene expression in human sperm. International Journal of Fertility and Sterility, 2015, vol. 9, iss. 1, pp. 129–136. https://doi.org/10.22074/ijfs.2015.4217