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Supporting Online Material
Materials and Methods
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29 May 2007; accepted 22 June 2007
Published online 12 July 2007;
Include this information when citing this paper.
Rapid Erasure of Long-Term
Memory Associations in the Cortex
by an Inhibitor of PKMz
Reut Shema,1Todd Charlton Sacktor,2Yadin Dudai1*
Little is known about the neuronal mechanisms that subserve long-term memory persistence in the
brain. The components of the remodeled synaptic machinery, and how they sustain the new synaptic or
cellwide configuration over time, are yet to be elucidated. In the rat cortex, long-term associative
memories vanished rapidly after local application of an inhibitor of the protein kinase C isoform,
protein kinase M zeta (PKMz). The effect was observed for at least several weeks after encoding and
may be irreversible. In the neocortex, which is assumed to be the repository of multiple types of
long-term memory, persistence of memory is thus dependent on ongoing activity of a protein kinase
long after that memory is considered to have consolidated into a long-term stable form.
tion (LTP) in hippocampus and for sustaining
hippocampus-dependent spatial memory (1). Itis
to store multiple types of long-term memory in
the mammalian brain (2, 3). We set out to deter-
ersistent phosphorylation by the atypical
for maintenance of long-term potentia-
is critical for storage of long-term memory in
We trained rats on conditioned taste aversion
(CTA) (5) using saccharin as the conditioned
stimulus (CS), and 3 days later, microinfused the
selective PKMz pseudosubstrate inhibitor ZIP
(1, 6) bilaterally into the IC. Controls received
vehicle only. We tested one ZIP group 1 week
later and another 1 month later. ZIP in the IC
blocked CTA memory in both groups [one-way
analysis of variance (ANOVA), F(2,16) = 7.61,
P < 0.005] (Fig. 1A). Post hoc comparisons
unveiled no difference between the ZIP groups;
however, each was different from the control
(P < 0.05). The difference persisted in extinc-
tion[repeated-measures ANOVA, group effect,
F(2,16) = 6.17, P < 0.01, test effect, F(2,32) =
8.91, P < 0.001]. The ZIP groups did not differ
from each other, but each was different from
control (P < 0.05).
Although consolidation of memory in the IC
is considered to be over within hours, judged by
loss of vulnerability to amnesic agents (7), we
wondered whether the vulnerability to ZIP re-
flects a longer consolidation process (8). We ad-
ministered ZIP at various times 3 to 25 days after
training, followed by CTA testing. The PKMz
Fig. 1. Erasureoflong-termCTAmemorybyasingleapplicationofthePKMz
inhibitor ZIP into the IC. (A) ZIP was administered 3 days after training, and
memorywas tested 1 week or1 month later.Controls weretested at 1month.
Data are shown for three successive tests, 1 day apart. The dashed line
for the CS may develop over time in naïve or CTA-extinguished rats, but AI
usually does not decline below 20 to 30 even in naïve rats. For statistics, see
text. (B) ZIP was microinfused into the IC at the indicated times after training,
which was a single conditioning session (3 and 7 days groups) or two
successive conditioning sessions, a day apart (25 days group). Memory was
tested 2 hours (3 and 7 days groups) or 1 day (25 days group) later. (C) Rats
weretrained onCTA and tested once3dayslater,followed 1to4 min laterby
microinfusion of ZIP into the IC. Although spontaneous recovery was seen in
the no-test interval between days 4 and 12 in the control group, the ZIP-
reinstatement (LiCl, day 13).
1Department of Neurobiology, The Weizmann Institute of
Science, Rehovot 76100, Israel.2Departments of Physiol-
ogy, Pharmacology, and Neurology, The Robert F. Furchgott
Center for Neural and Behavioral Science, SUNY Downstate
Medical Center, 450 Clarkson Avenue, Brooklyn, NY 11203,
*To whom correspondence should be addressed. E-mail:
VOL 31717 AUGUST 2007