Effects of Internal K
+
and ABA on the Voltage- and Time-dependence of the Outward
K
+
-rectifier in Vicia Guard Cells
F. Lemtiri-Chlieh
Department of Plant Sciences, University of Cambridge, Downing Street, Cambridge CB2 3EA
Received: 1 December 1995/Revised: 8 May 1996
Abstract. One of the main effects of abscisic acid (ABA)
is to induce net loss of potassium salts from guard cells
enabling the stomata to close. K
+
is released from the
vacuole into the cytosol and then to the extracellular
space. The effects of increasing cytosolic K
+
on the volt-
age- and time-dependence of the outwardly rectifying
K
+
-current (I
K,out
) in guard cell protoplasts (GCP) was
examined in the whole-cell configuration of the patch-
clamp technique. The same quantitative analysis was
performed in the presence of ABA at different internal
K
+
concentrations ([K
+
]
i
). Varying [K
+
]
i
in the patch
pipette from 100 to 270 mM increased the magnitude of
I
K,out
in a nonlinear manner and caused a negative shift in
the midpoint (V
0.5
) of its steady-state activation curve.
External addition of ABA (10–20 M) also increased the
magnitude of I
K,out
at all [K
+
]
i
, but caused a shift in V
0.5
of the steady-state activation curve only in those GCP
loaded with 150 mM internal K
+
or less. Indeed, V
0.5
did
not shift upon addition of ABA when the [K
+
]
i
was
above 150 mM and up to 270 mM, i.e., the shift in V
0.5
caused by ABA depended on the [K
+
]
i
. Both increase in
[K
+
]
i
and external addition of ABA, decreased (by ≈
20%) the activation time constant (
n
) of I
K,out
. The
small decrease in
n
, in both cases, was found to be
independent of the membrane voltage. The results indi-
cate that ABA mimics the effect of increasing cytoplas-
mic K
+
, and suggest that ABA may increase I
K,out
and
alter V
0.5
of its steady-state activation curve via an en-
hancement in cytosolic K
+
. This report describes for the
first time the effects of [K
+
]
i
on the voltage- and time-
dependence of I
K,out
in guard cells. It also provides an
explanation for the quantitative (total membrane current)
and qualitative (current kinetics) differences found be-
tween intact guard cells and their protoplasts.
Key words: Outward K
+
current — Abscisic acid —
Internal K
+
concentration — Voltage-dependence —
Time-dependence — Guard cell protoplast — Vicia faba
Introduction
The plant hormone abscisic acid (ABA), produced dur-
ing conditions of water stress, plays an important role in
regulating stomatal movement. ABA inhibits stomatal
opening and promotes stomatal closing, thereby reducing
transpirational water loss. The mechanisms by which
ABA promotes stomatal closing have been extensively
studied during the last decade (for reviews, see MacRob-
bie, 1991; Blatt, 1991; Blatt & Thiel, 1993; Assmann,
1993; Ward, Pei & Schroeder, 1995). It is believed that
this effect is achieved through a net loss from the guard
cells of osmotically active solutes, mainly in the form of
potassium salts.
On the question of the origin of this K
+
, earlier tracer
flux studies, measuring rate of loss of
86
Rb
+
from iso-
lated guard cells of Commelina communis. L., have
shown that the amount of tracer lost during the ABA
transient is greater than the cytoplasmic content at the
time of the change (MacRobbie, 1981). Further flux
work (MacRobbie, 1990), has also shown a biphasic
transient response of
86
Rb
+
efflux from intact guard cells
after short periods of applications of ABA. The second
(or slow) component of this biphasic response was at-
tributed to the release of
86
Rb
+
from the vacuole into the
cytoplasm, following activation of vacuolar K
+
channels.
The evidence for such speculation was the absence of the
slow component of the efflux transient in tissue loaded
for only a short period of time, with little or no tracer in
the vacuole. In a recent report, Ward and Schroeder
(1994) showed evidence for the existence in the tono-
plast of voltage-independent K
+
-selective channels
(VK). These VK channels have a conductance of 70 pS Correspondence to: F. Lemtiri-Chlieh
J. Membrane Biol. 153, 105–116 (1996) The Journal of
Membrane
Biology
© Springer-Verlag New York Inc. 1996