Glutamate activates cation currents in the plasma membrane of Arabidopsis root cells

Abstract

The effect of glutamate on plant plasma membrane cation transport was studied in roots of Arabidopsis thaliana (L.) Heynh. Patch-clamp experiments using root protoplasts, ²²Na⁺ unidirectional fluxes into intact roots and measurements of cytosolic Ca²⁺ activity using plants expressing cytosolically-targeted aequorin in specific cell types were carried out. It was demonstrated that low-millimolar concentrations of glutamate activate within seconds both Na⁺ and Ca²⁺ currents in patch-clamped protoplasts derived from roots. The probability of observing glutamate-activated currents increased with increasing glutamate concentration (up to 29% at 3 mM); half-maximal activation was seen at 0.2–0.5 mM glutamate. Glutamate-activated currents were voltage-insensitive, ‘instantaneous’ (completely activated within 2–3 ms of a change in voltage) and non-selective for monovalent cations (Na⁺, Cs⁺ and K⁺). They also allowed the permeation of Ca²⁺. Half-maximal Na⁺ currents occurred at 20–30 mM Na⁺. Glutamate-activated currents were sensitive to non-specific blockers of cation channels (quinine, La³⁺, Gd³⁺). Although low-millimolar concentrations of glutamate did not usually stimulate unidirectional influx of ²²Na⁺ into intact roots, they reliably caused an increase in cytosolic Ca²⁺ activity in protoplasts isolated from the roots of aequorin-transformed Arabidopsis plants. The response of cytosolic Ca²⁺ activity revealed a two-phase development, with a rapid large transient increase (lasting minutes) and a prolonged subsequent stage (lasting hours). Use of plants expressing aequorin in specific cell types within the root suggested that the cell types most sensitive to glutamate were in the mature epidermis and cortex. The functional significance of these glutamate-activated currents for both cation uptake into plants and cell signaling remains the subject of speculation, requiring more knowledge about the dynamics of apoplastic glutamate in plants.