Ion currents involved in early Nod-factor response in Medicago sativa
root hairs.
Young
root hairs and protoplasts of Medicago sativa. The seeds were germinated
according to the protocol described in Methods. The plantlets were left
for 6 h in a vertical position for root hairs to grow. A, root hairs;
B, protoplasts released from young root hair tips, after 3 min incubation
with enzyme cocktail. Bar scales = 10 µm.
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Root hairs are a primary site for nutrient absorption and for initiation of signaling
processes linked to variations of the root environment: plant-microbe interactions
or abiotic changes. In many of these cases, the earliest detectable response is the
modification of plasma membrane transports, detected through alteration of the electrical
membrane potential. In spite of this, root hairs have not been extensively used in
electrophysiological research so far. Problems with cell shape and current coupling are
often prohibitive for microelectrode voltage clamp on intact root hairs. In the present
study, these difficulties have been overcome and the ion channel currents are described
for young root hairs from alfalfa seedlings (Medicago sativa cv Sitel).
Electrophysiological and pharmacological studies indicated an inward rectifying K+
time-dependent current. This current was sensitive to tetraethylammonium and Cs+ (10 mM
each). Two others currents never shown in root hairs were described: an outward rectifying
time-dependent K+ current, inhibited by tetraethylammonium and Cs+ (10 mM each) allowing
K+ efflux under strong depolarizations and an instantaneous inward current identified as
an anion current, inhibited by 4-acetamido-4'-isothiocyanatostilbene-2,2', disulfonic acid
and anthracene-9-carboxylic acid (100 µM each). These results should contribute to the
understanding of root hair development and of signaling processes in M. sativa root hairs.
Nod-factor [NodRm-IV(Ac,S)], isolated from the bacterium Rhizobium meliloti, induces a
well known depolarization in Medicago sativa (cv Sitel) root hairs (Kurkdjian Plant
Physiol. 107: 783-790, 1995). The analysis of this membrane response using the
discontinuous single electrode voltage clamp technique (dSEVC) shows that anion channel,
K+ channel and H+-ATPase pump currents are involved in young growing root hairs. The early
Nod-factor-induced depolarization is due to the increase of the inward ion current and to
the inhibition of the H+-pump. It involved an instantaneous inward anion current (IIAC)
and/or a time dependent inward K+ current (IRKC). These two ion currents are then
down-regulated while the H+-pump is stimulated allowing the long term rectification of the
membrane potential (Em). Our results support the idea that the regulation of inward
current plays a primary role in the Nod-factor induced electrical response, the nature of
ions carried by these currents depending on the activated anion and/or K+ channels at the
plasma membrane.
