Phospholipase C-dependent phosphoinositide breakdown induced by ELF-EMF in Peganum harmala calli

Phospholipase C-dependent phosphoinositide breakdown induced by ELF-EMF in Peganum harmala calli Abstract In the last decades, the interaction of extremely low frequency (ELF) electromagnetic fields (EMF) with living systems has received considerable attention because of relevant impact to public health. The very recent interest concerning the biological effects of weak pulsed EMF on plant organisms and cells is motivated by the possibility of affecting the growth of a variety of useful plants. Such opportunity might well have important economic consequences. Several hypotheses have been proposed to explain the influence of ELF-EMF on cells and most of them share the common assumption that the cell membrane is the primary interaction site. Therefore, the long-term objective of our work is the elucidation of the molecular mechanism(s) by which ELF-EMF initiate a cascade of cytoplasmic and nuclear signalling events in plant cells. In particular, the aim of the present study was to investigate whether, like in animal, also in plant cells the ELF-EMF are able to affect the phosphoinositide (PI) signal transduction pathway, by triggering the PI-PLC activation and the consequent phosphatidylinositol 4,5-bisphosphate (PtdIns 4,5-P2) breakdown. In this work, we present evidence for an effect of the 50 Hz, 1 gauss ELF-EMF on the transient PtdIns 4,5-P2 breakdown of callus cells from Peganum harmala (P. harmala), a plant of biochemical and pharmacological interest belonging to Zygophyllaceae known since the first century A.D. for its effects on the central nervous system. Our results evidenced a statistically significant decrease in PtdIns 4,5-P2 concentrations and a different involvement of the constituting fatty acids in the induced breakdown. Neomycin, 1-o-octadecyl-2-omethyl- rac-glycero-3-phosphocholine (ET-18-OCH3) and 1-(6-{[17b-3-methoxyextra-1,3,5(10)-trien-17-yl]-amino} hexyl)-1H-pyrrole-2,5-dione (U-73122), known to be PLC inhibitors, blocked the EMF-induced PtdIns 4,5-P2 breakdown, suggesting the PI-PLC involvement in the EMF signal cascade. The manipulation of the lipid-based signalling pathway by phosphoinositide-phospholipase C (PI-PLC) inhibitors seems to support the hypothesis that, as in animals, also in plants, the cell membrane is the primary impact site of ELF electromagnetic stimulus and that this interaction could probably involve the activation of PI signal transduction pathway including a heterotrimeric G protein. Because of the growing evidence indicating the reactive oxygen intermediates (ROI) as second messengers in cellular responses, especially in the PI cascade activation triggered by physical agents such as UV-C both in animal and in plant cells, we evaluated the effects of treatment with ascorbic acid (Asc) on EMFdependent PtdIns 4,5-P2 breakdown. Our results do not support the hypothesis for ROI formation as an earliest event in the cellular responses to electromagnetic impact.