Register      Login
Functional Plant Biology Functional Plant Biology Society
Plant function and evolutionary biology
RESEARCH ARTICLE

Certain calcium channel inhibitors exhibit a number of secondary effects on the physiological properties in Nitellopsis obtusa: a voltage clamp approach

Vilmantas Pupkis https://orcid.org/0000-0002-4348-0142 A * , Indre Lapeikaite A , Julius Kavaliauskas A , Kazimierz Trębacz B and Vilma Kisnieriene A
+ Author Affiliations
- Author Affiliations

A Department of Neurobiology and Biophysics, Institute of Biosciences, Life Sciences Center, Vilnius University, 7 Saulėtekio Avenue, 10257 Vilnius, Lithuania.

B Department of Plant Physiology and Biophysics, Institute of Biological Sciences, Faculty of Biology and Biotechnology, Maria Curie-Sklodowska University, 19 Akademicka, 20-033 Lublin, Poland.

* Correspondence to: vilmantas.pupkis@gmc.vu.lt

Handling Editor: Vadim Demidchik

Functional Plant Biology 50(3) 195-205 https://doi.org/10.1071/FP22106
Submitted: 20 May 2022  Accepted: 6 October 2022   Published: 2 November 2022

© 2023 The Author(s) (or their employer(s)). Published by CSIRO Publishing

Abstract

An unsolved problem of contemporary plant electrophysiology is the identity of Ca2+ channels responsible for the initiation of the action potential. We took a pharmacological approach and applied several Ca2+ channel blockers (verapamil, tetrandrine, and NED-19) on a Characean (Nitellopsis obtusa) algae model system. The impact of the selected pharmaceuticals on the parameters of excitation transients of a single cell was analysed employing the two-electrode voltage clamp technique. It was revealed that tetrandrine exerted no effect, while both verapamil and NED-19 prolonged activation and inactivation durations of the excitatory Cl current. NED-19 also significantly depolarised the excitation threshold membrane potential and shifted Ca2+ current reversal potential. Thus, NED-19 most specifically targeted Ca2+ channels. A viability assay paired with observations of cytoplasmic streaming revealed that verapamil affected not only Ca2+ channels but also exhibited non-specific effects, which eventually lead to cell death. Since many potential Ca2+ channel blockers exert additional undesirable non-specific effects, our study underlines the necessity to search for new more specific modulators of plant Ca2+ transport systems.

Keywords: action potential, calcium transport, electrophysiology, NED-19, Nitellopsis obtusa, tetrandrine, verapamil, voltage clamp.


References

Andjus PR (1998) Harvesting the biophysical field of the ‘Green Axon’. Iugoslavica Physiologica et Pharmacologica Acta 34, 1–9.

Beilby MJ (2007) Action potential in charophytes. International Review of Cytology 257, 43–82.
Action potential in charophytes.Crossref | GoogleScholarGoogle Scholar |

Beilby MJ (2019) Chara braunii genome: a new resource for plant electrophysiology. Biophysical Reviews and Letters 11, 235–239.
Chara braunii genome: a new resource for plant electrophysiology.Crossref | GoogleScholarGoogle Scholar |

Beilby MJ, Casanova MT (2014) ‘The physiology of Characean cells.’ (Springer)

Beilby MJ, Al Khazaaly S (2016) Re-modeling Chara action potential: I. From Thiel model of Ca2+ transient to action potential form. AIMS Biophysics 3, 431–449.
Re-modeling Chara action potential: I. From Thiel model of Ca2+ transient to action potential form.Crossref | GoogleScholarGoogle Scholar |

Bhagya N, Chandrashekar KR (2016) Tetrandrine - a molecule of wide bioactivity. Phytochemistry 125, 5–13.
Tetrandrine - a molecule of wide bioactivity.Crossref | GoogleScholarGoogle Scholar |

Biskup B, Gradmann D, Thiel G (1999) Calcium release from InsP3-sensitive internal stores initiates action potential in Chara. FEBS Letters 453, 72–76.
Calcium release from InsP3-sensitive internal stores initiates action potential in Chara.Crossref | GoogleScholarGoogle Scholar |

Brunet T, Arendt D (2016) From damage response to action potentials: early evolution of neural and contractile modules in stem eukaryotes. Philosophical Transactions of the Royal Society B: Biological Sciences 371, 20150043
From damage response to action potentials: early evolution of neural and contractile modules in stem eukaryotes.Crossref | GoogleScholarGoogle Scholar |

Demidchik V, Sokolik A, Yurin V (1997) The effect of Cu2+ on ion transport systems of the plant cell plasmalemma. Plant Physiology 114, 1313–1325.
The effect of Cu2+ on ion transport systems of the plant cell plasmalemma.Crossref | GoogleScholarGoogle Scholar |

De Vriese K, Costa A, Beeckman T, Vanneste S (2018) Pharmacological strategies for manipulating plant Ca2+ signalling. International Journal of Molecular Sciences 19, 1506
Pharmacological strategies for manipulating plant Ca2+ signalling.Crossref | GoogleScholarGoogle Scholar |

Drinyaev VA, Mosin VA, Kruglyak EB, Sterlina TS, Kataev AA, Berestovsky GN, Kokoz YM (2001) Effect of avermectins on Ca2+ -dependent Cl− currents in plasmalemma of Chara corallina cells. The Journal of Membrane Biology 182, 71–79.
Effect of avermectins on Ca2+ -dependent Cl currents in plasmalemma of Chara corallina cells.Crossref | GoogleScholarGoogle Scholar |

Edel KH, Marchadier E, Brownlee C, Kudla J, Hetherington AM (2017) The evolution of calcium-based signalling in plants. Current Biology 27, R667–R679.
The evolution of calcium-based signalling in plants.Crossref | GoogleScholarGoogle Scholar |

Gyenes M, Bulychev AA, Kurella GA (1980) Voltage clamp studies on the slow inward current during the excitation of Nitellopsis obtusa. Journal of Experimental Botany 31, 589–595.
Voltage clamp studies on the slow inward current during the excitation of Nitellopsis obtusa.Crossref | GoogleScholarGoogle Scholar |

Hedrich R (2012) Ion channels in plants. Physiological Reviews 92, 1777–1811.
Ion channels in plants.Crossref | GoogleScholarGoogle Scholar |

Kintzer AF, Stroud RM (2016) Structure, inhibition and regulation of two-pore channel TPC1 from Arabidopsis thaliana. Nature 531, 258–264.
Structure, inhibition and regulation of two-pore channel TPC1 from Arabidopsis thaliana.Crossref | GoogleScholarGoogle Scholar |

Kisnieriene V, Lapeikaite I, Sevriukova O, Ruksenas O (2016) The effects of Ni2+ on electrical signaling of Nitellopsis obtusa cells. Journal of Plant Research 129, 551–558.
The effects of Ni2+ on electrical signaling of Nitellopsis obtusa cells.Crossref | GoogleScholarGoogle Scholar |

Kisnieriene V, Lapeikaite I, Pupkis V (2018) Electrical signalling in Nitellopsis obtusa: potential biomarkers of biologically active compounds. Functional Plant Biology 45, 132–142.
Electrical signalling in Nitellopsis obtusa: potential biomarkers of biologically active compounds.Crossref | GoogleScholarGoogle Scholar |

Kisnieriene V, Lapeikaite I, Pupkis V, Beilby MJ (2019) Modeling the action potential in Characeae Nitellopsis obtusa: effect of saline stress. Frontiers in Plant Science 10, 82
Modeling the action potential in Characeae Nitellopsis obtusa: effect of saline stress.Crossref | GoogleScholarGoogle Scholar |

Koselski M, Pupkis V, Hashimoto K, Lapeikaite I, Hanaka A, Wasko P, Plukaite E, Kuchitsu K, Kisnieriene V, Trebacz K (2021) Impact of mammalian two-pore channel inhibitors on long-distance electrical signals in the Characean macroalga Nitellopsis obtusa and the early terrestrial liverwort Marchantia polymorpha. Plants 10, 647
Impact of mammalian two-pore channel inhibitors on long-distance electrical signals in the Characean macroalga Nitellopsis obtusa and the early terrestrial liverwort Marchantia polymorpha.Crossref | GoogleScholarGoogle Scholar |

Kourie JI (1994) Transient Cl− and K+ currents during the action potential in Charainflata (effects of external sorbitol, cations, and ion channel blockers). Plant Physiology 106, 651–660.
Transient Cl and K+ currents during the action potential in Charainflata (effects of external sorbitol, cations, and ion channel blockers).Crossref | GoogleScholarGoogle Scholar |

Krol E, Dziubińska H, Trebacz K (2004) Low-temperature-induced transmembrane potential changes in mesophyll cells of Arabidopsis thaliana, Helianthus annuus and Vicia faba. Physiologia Plantarum 120, 265–270.
Low-temperature-induced transmembrane potential changes in mesophyll cells of Arabidopsis thaliana, Helianthus annuus and Vicia faba.Crossref | GoogleScholarGoogle Scholar |

Lapeikaite I, Dragunaite U, Pupkis V, Ruksenas O, Kisnieriene V (2019) Asparagine alters action potential parameters in single plant cell. Protoplasma 256, 511–519.
Asparagine alters action potential parameters in single plant cell.Crossref | GoogleScholarGoogle Scholar |

Lapeikaite I, Pupkis V, Neniskis V, Ruksenas O, Kisnieriene V (2020) Glutamate and NMDA affect cell excitability and action potential dynamics of single cell of macrophyte Nitellopsis obtusa. Functional Plant Biology 47, 1032–1040.
Glutamate and NMDA affect cell excitability and action potential dynamics of single cell of macrophyte Nitellopsis obtusa.Crossref | GoogleScholarGoogle Scholar |

Lühring H, Nguyen VD, Schmidt L, Röse USR (2007) Caterpillar regurgitant induces pore formation in plant membranes. FEBS Letters 581, 5361–5370.
Caterpillar regurgitant induces pore formation in plant membranes.Crossref | GoogleScholarGoogle Scholar |

Lunevsky VZ, Zherelova OM, Vostrikov IY, Berestovsky GN (1983) Excitation of Characeae cell membranes as a result of activation of calcium and chloride channels. The Journal of Membrane Biology 72, 43–58.
Excitation of Characeae cell membranes as a result of activation of calcium and chloride channels.Crossref | GoogleScholarGoogle Scholar |

Nishiyama T, Sakayama H, de Vries J, Buschmann H, Saint-Marcoux D, Ullrich KK, Haas FB, Vanderstraeten L, Becker D, Lang D, Vosolsobě S, Rombauts S, Wilhelmsson PKI, Janitza P, Kern R, Heyl A, Rümpler F, Villalobos LIAC, Clay JM, Skokan R, Toyoda A, Suzuki Y, Kagoshima H, Schijlen E, Tajeshwar N, Catarino B, Hetherington AJ, Saltykova A, Bonnot C, Breuninger H, Symeonidi A, Radhakrishnan G V, Van Nieuwerburgh F, Deforce D, Chang C, Karol KG, Hedrich R, Ulvskov P, Glöckner G, Delwiche CF, Petrášek J, Van de Peer Y, Friml J, Beilby M, Dolan L, Kohara Y, Sugano S, Fujiyama A, Delaux P-M, Quint M, Theißen G, Hagemann M, Harholt J, Dunand C, Zachgo S, Langdale J, Maumus F, Van Der Straeten D, Gould SB, Rensing SA (2018) The Chara genome: secondary complexity and implications for plant terrestrialization. Cell 174, 448–464.e24.
The Chara genome: secondary complexity and implications for plant terrestrialization.Crossref | GoogleScholarGoogle Scholar |

Novikova EM, Vodeneev VA, Sukhov VS (2017) Mathematical model of action potential in higher plants with account for the involvement of vacuole in the electrical signal generation. Biochemistry (Moscow), Supplement Series A: Membrane and Cell Biology 11, 151–167.
Mathematical model of action potential in higher plants with account for the involvement of vacuole in the electrical signal generation.Crossref | GoogleScholarGoogle Scholar |

Okazaki Y, Ishigami M, Iwasaki N (2002) Temporal relationship between cytosolic free Ca2+ and membrane potential during hypotonic turgor regulation in a brackish water charophyte Lamprothamnium succinctum. Plant and Cell Physiology 43, 1027–1035.
Temporal relationship between cytosolic free Ca2+ and membrane potential during hypotonic turgor regulation in a brackish water charophyte Lamprothamnium succinctum.Crossref | GoogleScholarGoogle Scholar |

Pyatygin SS, Opritov VA, Vodeneev VA (2008) Signaling role of action potential in higher plants. Russian Journal of Plant Physiology 55, 285–291.
Signaling role of action potential in higher plants.Crossref | GoogleScholarGoogle Scholar |

Reid RJ, Smith FA (1992) Regulation of calcium influx in Chara. Plant Physiology 100, 637–643.
Regulation of calcium influx in Chara.Crossref | GoogleScholarGoogle Scholar |

Shiina T, Tazawa M (1987) Demonstration and characterization of Ca2+ channel in tonoplast-free cells if Nitellopsis obtusa. The Journal of Membrane Biology 96, 263–276.
Demonstration and characterization of Ca2+ channel in tonoplast-free cells if Nitellopsis obtusa.Crossref | GoogleScholarGoogle Scholar |

Shimmen T (2007) The sliding theory of cytoplasmic streaming: fifty years of progress. Journal of Plant Research 120, 31–43.
The sliding theory of cytoplasmic streaming: fifty years of progress.Crossref | GoogleScholarGoogle Scholar |

Stolarz M, Trębacz K (2021) Spontaneous rapid leaf movements and action potentials in Mimosa pudica L. Physiologia Plantarum 173, 1882–1888.
Spontaneous rapid leaf movements and action potentials in Mimosa pudica L.Crossref | GoogleScholarGoogle Scholar |

Sukhov V, Sukhova E, Vodeneev V (2019) Long-distance electrical signals as a link between the local action of stressors and the systemic physiological responses in higher plants. Progress in Biophysics and Molecular Biology 146, 63–84.
Long-distance electrical signals as a link between the local action of stressors and the systemic physiological responses in higher plants.Crossref | GoogleScholarGoogle Scholar |

Tester M, MacRobbie EAC (1990) Cytoplasmic calcium affects the gating of potassium channels in the plasma membrane of Chara corallina: a whole-cell study using calcium-channel effectors. Planta 180, 569–581.
Cytoplasmic calcium affects the gating of potassium channels in the plasma membrane of Chara corallina: a whole-cell study using calcium-channel effectors.Crossref | GoogleScholarGoogle Scholar |

Thiel G, Homann U, Plieth C (1997) Ion channel activity during the action potential in Chara: new insights with new techniques. Journal of Experimental Botany 48, 609–622.
Ion channel activity during the action potential in Chara: new insights with new techniques.Crossref | GoogleScholarGoogle Scholar |

Tsutsui I, Ohkawa T-a, Nagai R, Kishimoto U (1987a) Role of calcium ion in the excitability and electrogenic pump activity of the Chara corallina membrane: I. Effects of La3+, verapamil, EGTA, W-7, and TFP on the action potential. The Journal of Membrane Biology 96, 65–73.
Role of calcium ion in the excitability and electrogenic pump activity of the Chara corallina membrane: I. Effects of La3+, verapamil, EGTA, W-7, and TFP on the action potential.Crossref | GoogleScholarGoogle Scholar |

Tsutsui I, Ohkawa T-a, Nagai R, Kishimoto U (1987b) Role of calcium ion in the excitability and electrogenic pump activity of the Chara corallina membrane: II. Effects of La3+, EGTA, and calmodulin antagonists on the current-voltage relation. The Journal of Membrane Biology 96, 75–84.
Role of calcium ion in the excitability and electrogenic pump activity of the Chara corallina membrane: II. Effects of La3+, EGTA, and calmodulin antagonists on the current-voltage relation.Crossref | GoogleScholarGoogle Scholar |

Vodeneev V, Akinchits E, Sukhov V (2015) Variation potential in higher plants: mechanisms of generation and propagation. Plant Signaling & Behavior 10, e1057365
Variation potential in higher plants: mechanisms of generation and propagation.Crossref | GoogleScholarGoogle Scholar |

Wacke M, Thiel G, Hütt M-T (2003) Ca2+ dynamics during membrane excitation of green alga Chara: model simulations and experimental data. Journal of Membrane Biology 191, 179–192.
Ca2+ dynamics during membrane excitation of green alga Chara: model simulations and experimental data.Crossref | GoogleScholarGoogle Scholar |

Williamson RE, Ashley CC (1982) Free Ca2+ and cytoplasmic streaming in the alga Chara. Nature 296, 647–651.
Free Ca2+ and cytoplasmic streaming in the alga Chara.Crossref | GoogleScholarGoogle Scholar |

Zherelova OM (1990) Verapamil-sensitive cation channels in the plasmalemma of perfused Nitellopsis obtusa cells. Comparative Biochemistry and Physiology Part A: Physiology 96, 173–176.
Verapamil-sensitive cation channels in the plasmalemma of perfused Nitellopsis obtusa cells.Crossref | GoogleScholarGoogle Scholar |

Zherelova OM, Grishchenko VM, Chaylakhyan LM (1994) Blockers of Ca2+ channels in the plasmalemma of perfused Characeae cells. Comparative Biochemistry and Physiology Part C: Pharmacology, Toxicology and Endocrinology 107, 475–480.
Blockers of Ca2+ channels in the plasmalemma of perfused Characeae cells.Crossref | GoogleScholarGoogle Scholar |

Zherelova OM, Kataev AA, Grischenko VM, Shtanchaev RS (2016) Haloperidol modulates ion transport in Chara corallina cells. Cell and Tissue Biology 10, 476–485.
Haloperidol modulates ion transport in Chara corallina cells.Crossref | GoogleScholarGoogle Scholar |

Zimmermann MR, Maischak H, Mithöfer A, Boland W, Felle HH (2009) System potentials, a novel electrical long-distance apoplastic signal in plants, induced by wounding. Plant Physiology 149, 1593–1600.
System potentials, a novel electrical long-distance apoplastic signal in plants, induced by wounding.Crossref | GoogleScholarGoogle Scholar |