Ion channel clustering enhances weak electric field detection by neutrophils: apparent roles of SKF96365-sensitive cation channels and myeloperoxidase trafficking in cellular responses

Abstract

We have tested Galvanovskis and Sandblom's prediction that ion channel clustering enhances weak electric field detection by cells as well as how the elicited signals couple to metabolic alterations. Electric field application was timed to coincide with certain known intracellular chemical oscillators (phase-matched conditions). Polarized, but not spherical, neutrophils labeled with anti-K(v)1.3, FL-DHP, and anti-TRP1, but not anti-T-type Ca(2+) channels, displayed clusters at the lamellipodium. Resonance energy transfer experiments showed that these channel pairs were in close proximity. Dose-field sensitivity studies of channel blockers suggested that K(+) and Ca(2+) channels participate in field detection, as judged by enhanced oscillatory NAD(P)H amplitudes. Further studies suggested that K(+) channel blockers act by reducing the neutrophil's membrane potential. Mibefradil and SKF93635, which block T-type Ca(2+) channels and SOCs, respectively, affected field detection at appropriate doses. Microfluorometry and high-speed imaging of indo-1-labeled neutrophils was used to examine Ca(2+) signaling. Electric fields enhanced Ca(2+) spike amplitude and triggered formation of a second traveling Ca(2+) wave. Mibefradil blocked Ca(2+) spikes and waves. Although 10 microM SKF96365 mimicked mibefradil, 7 microM SKF96365 specifically inhibited electric field-induced Ca(2+) signals, suggesting that one SKF96365-senstive site is influenced by electric fields. Although cells remained morphologically polarized, ion channel clusters at the lamellipodium and electric field sensitivity were inhibited by methyl-beta-cyclodextrin. As a result of phase-matched electric field application in the presence of ion channel clusters, myeloperoxidase (MPO) was found to traffic to the cell surface. As MPO participates in high amplitude metabolic oscillations, this suggests a link between the signaling apparatus and metabolic changes. Furthermore, electric field effects could be blocked by MPO inhibition or removal while certain electric field effects were mimicked by the addition of MPO to untreated cells. Therefore, channel clustering plays an important role in electric field detection and downstream responses of morphologically polarized neutrophils. In addition to providing new mechanistic insights concerning electric field interactions with cells, our work suggests novel methods to remotely manipulate physiological pathways.

TRPC1 channels regulate directionality of migrating cells

“…This gradient is steeper in hTRPC1-HA-overexpressing cells. Although we could not visualize a concentration of the TRPC1 channel at the leading edge of MDCK-F cells as observed in neutrophils [21], we could functionally show that TRPC1 is active at the leading edge and generates a Ca 2+ gradient. This finding corresponds to the fact that only the local blockade of stretch-activated Ca 2+ channels at the leading edge causes a global inhibition of traction forces and cell migration [28].…”

“…Related studies showed that TRPC1 is important for path finding of outgrowing neurites [25,42]. Moreover, TRPC1 has been implicated in electric field detection which affects directed migration of neutrophils [21]. Thus, the fact that TRPC1 is important for directed cell migration becomes apparent in various cell types.…”

Silencing TRPC1 expression inhibits invasion of CNE2 nasopharyngeal tumor cells

“…Since cytoplasmic Ca 2+ has an impact on numerous crucial cellular functions relevant to tumorigenesis and progression, like cell proliferation, apoptosis, differentiation and angiogenesis (4,21), it is not unexpected to find that TRPC1 was involved in the development of various cancers. In this study, we found that inhibition of TRPC1 channels by either RNA interference or 2-APB not only leads to significant attenuated migratory and invasive abilities of an NPC cell line CNE2 but is also accompanied by notable increases of MMP2 and MMP9 expression, indicating a crucial role of (Ca Previous data have proven that TRPC1 is of great importance for cellular polarity and migration directing in neuritis, neutrophils and transformed renal epithelial cells (22)(23)(24). In addition to the above studies, we found that TRPC1 are also implicated in the migration of NPC cells.…”

A Model Proposal for Long-Lasting Electromagnetic Forces-Biological System Interaction: Molecular Fatigue Damages

“…An additional effect for enhancing sensitivity (already addressed in the present review) was also discussed by Kindzelskii and Petty: the coherence and cooperative interaction of receptors to receptors or channels to channels (the distance of individual channels being only about 7 nm) [16]. The coupling may take place via conformational mechanisms or via other coupling (electron tunneling or other quantum effects).…”

“…The coupling may take place via conformational mechanisms or via other coupling (electron tunneling or other quantum effects). All these mechanisms may further improve the sinyal amplification [16,17].…”