Utilization of the Vibrating Probe and Ion-Selective Microelectrode Techniques to Investigate Electrophysiological Responses to Wounding in Pea Roots

This paper examines the ionic composition of wound-induced electricalcurrents in higher plant tissue, using two non-injurious electrophysiologicaltechniques. By simultaneous recording of K⁺, H⁺ , and Ca²⁺ ionfluxes with extracellular ion-selective microelectrodes, wehave determined that a Ca²⁺ influx (2.4 µA cm^–2),a small H⁺ influx (0.17 µA cm^–2) and a large K⁺efflux (16 µA cm^–2) occur immediately after woundingin roots of Pisum sativum L. var. Greenfeast. Using an extracellularvibrating probe at the wound site, net ion currents of 26 µAcm^–2 were measured 5 min after wounding. In a more concentratedbathing medium (1/4 rather than 1/16 strength Hoagland's solution),net ion currents of 59 µA cm^–2 were measured, andthese would appear to be the largest extracellular currentsthat have been measured in plants. We made a quantitative comparisonof the summed ion fluxes with the net ion currents and thisrevealed that ion fluxes, in addition to those measured here,occur after wounding. Key words: Wounding, ion flux, electric current, calcium, potassium     

A calcium influx precedes organogenesis in Graptopetalum

Abstract. An account is given of an investigation of net ionic currents and specific ion fluxes occuring during the initiation of organogenesis in detached leaves of Graptopetalum paraguayense E. Walther, in which a dramatic change in growth polarity is cytomorphologically evident 3–5 d after leaf detachment from the plant. Using the vibrating probe, it was possible to identify a peak of ionic current which is focused over the area of the leaf base where organogenesis is initiated. This net current is largest during the initial 12h after leaf detachment. With ion-selective microelectrodes capable of measuring H⁺, K⁺ and Ca²⁺ ion fluxes simultaneously in the same region of the leaf base, H⁺ and K⁺ fluxes remain relatively steady during the initial 24 h after detachment, while a large lanthanum-sensitive Ca²⁺ influx decreases by 50% from 2 to 12h. By 24h, Ca²⁺ transport is dominated by an efflux. We present evidence from a quantitative comparison of the ion current data collected using these two techniques, that Ca²⁺, H⁺ and K⁺ transport accounts for the major electrogenic ion fluxes during 2 and 12 but not 24 h after leaf detachment. The possibility is addressed that these ion currents, which precede organogenesis, and in particular the predominant Ca²⁺ flux, play a role in the establishment of growth polarity in higher plant tissues.     

PLANT MITOSIS PROMOTING FACTOR DISASSEMBLES THE MICROTUBULE PREPROPHASE BAND AND ACCELERATES PROPHASE PROGRESSION IN TRADESCANTIA

The regulation of mitosis in higher plant cells has been investigated by microinjecting protein kinase from the metaphase-arresting (met1) mutant ofChlamydomonas. Biochemical characterization of this enzyme complex confirms the presence of a p34^cdc2/cyclin B-like kinase. The enzyme was injected into living stamen hair cells ofTradescantia virginianain which microtubules (MTs) were visualized using fluorescent analogue cytochemistry and confocal laser scanning microscopy. Microinjection of this p34^cdc2/cyclin B-like kinase caused rapid disassembly of the preprophase band of MTs but not of interphase-cortical, spindle or phragmoplast MTs. Effects of the enzyme on the cytomorphology of live prophase cells were also monitored using video microscopy. We found that injection of this enzyme accelerated chromatin condensation and nuclear envelope breakdown. This indicates the presence and function in plants of an enzyme that can initiate nuclear division similar to the maturation or mitosis promoting factor (MPF) of animal cells. These studies provide the first direct evidence that the mitotically-active form of plant MPF can drive disassembly of preprophase band MTs, chromosome condensation and initiation of mitosis in plant cells.