15. Benesch, S., Polo, S., Lai, F.P., Anderson, K.I.,
Stradal, T.E., Wehland, J., and Rottner, K.
(2005). N-WASP deficiency impairs EGF
internalization and actin assembly at
clathrin-coated pits. J. Cell Sci. 118,
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Contribution of cytoskeleton to the
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Huganir, R.L. (1999). Clustering of AMPA
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Institute for Biochemistry I,
07743 Jena, Germany.
Sleep: The Ebb and Flow of Memory
A new study has shown that successful imprinting in domestic chicks depends
on post-training sleep; individual neurons were found to enter, leave and then
rejoin neural networks, and may constitute the memory trace of the imprinted
The last decade has seen a dramatic
increase in our understanding of
consolidation, moving the concept of a
role for sleep in memory consolidation
from a generally discredited (or at best
ignored) idea to a largely accepted
tenet among both memory and sleep
researchers. While the number of
players in this field remains small,
it is growing rapidly, and researchers
are now approaching the topic
with a remarkably broad array
of tools (Table 1). These range
from strictly behavioral studies in
humans — demonstrating, for
example, selective memory
enhancement across a night of sleep
 — to studies of the role of sleep in
cortical plasticity in the cat visual
cortex . Claire Jackson and
colleagues have now reported in
Current Biology  that imprinting in
domestic chicks is dependent on
post-training sleep; their work further
shows how the contribution of
individual neurons to the memory of
the imprinting stimulus develops over
the 24 hours following imprinting.
One of the most elusive questions
that remain in this field is what actually
happens to memories during sleep?
What makes this question so difficult is
our ignorance concerning the nature of
memory in general. At the cellular and
molecular level, no one has seen
a memory yet, or has strong evidence
for the exact form memories take or
how they are produced, let alone how
post-encoding processing leads to the
stabilization of memories or how they
are subsequently integrated into larger
networks of associated memories. So
it is something of a delight to find a
paper that looks at the time course of
changes in individual neurons as
a memory is first encoded and then
To analyze how neurons become
selectively responsive to a particular
sensory stimulus, and how subsequent
sleep stabilizes this selectivity,
Jackson et al.  studied imprinting in
domestic chicks. It is known that
neurons in the intermediate and medial
mesopallium of the chick brain store
imprinting information . After a
Table 1. Studies of sleep and memory:a sample of the range of topics related to sleep and memory reported in the literature, with citations for select topics.
HumansRodents and catsBirdsFruit flies
Modulation of sleep stage
distribution by prior learning
Sleep stage requirements for
EEG-waveform correlates of
Regional brain activation
correlates of learning [11,12]
Modulation of sleep stage
distribution by prior learning
Sleep stage requirements for
EEG-waveform correlates of
Reactivation of pairs of
hippocampal place cells 
Reactivation of temporal
patterns of place cells
increases in sleep 
Single cells reactivation of
birdsong neurons 
Recruitment of individual
neurons over time following
Developmental plasticity in
single cat visual cortex
Genetic screens for sleep and
wake active genes
deficits in schizophrenia 
Genetic mutant with both
reduced sleep and impaired
Regulation of sleep by
mushroom bodies 
of molecular markers of
synaptic strength 
young chick has been exposed to an
imprinting stimulus, a subset of these
neurons fire selectively in response
to the learned imprinting stimulus
(they are consequently referred to as
In an earlier study , this same
group reported that, for a given
imprinting stimulus, w7% of neurons
in the intermediate and medial
mesopallium were selectively
responsive even prior to imprinting,
and that this proportion increased to
13–14% after one or two hours of
imprinting. Most striking, however,
was the finding that, after an additional
20 hours without any further exposure
to the imprinting stimulus, the percent
of IS neurons increased further, to
22% of those analyzed (Figure 1A),
an increase reminiscent of the
improvements in performance seen
with sleep-dependent memory
consolidation (for review, see ).
The power of this chick preparation
is that single cells can be monitored
over long periods of time, and the
development of their status as IS
neurons followed. When such an
analysis was performed, the seemingly
straightforward monotonic increase
in the number of responding neurons
over time gave way to a much more
complex ebb and flow of cells into
and out of responsiveness. Thus, of the
16 IS neurons identified at baseline,
only one remained responsive after
an hour, and none was still responsive
after two hours. Further, of the 32 IS
neurons identified after one hour of
imprinting (including 31 that had not
been responsive at baseline), only
a third remained responsive after
an additional hour of imprinting
(Figure 1A). Unfortunately, recordings
from individual neurons could not be
maintained long enough to track these
cells forward to the final time point
20 hours later, when the number of IS
neurons had increased by half again.
This study left two unanswered
questions that the new paper by
Jackson et al.  has resolved. First,
might this final off-line increase in
IS neurons seen at 25 hours indeed
reflect sleep-dependent memory
consolidation? And second, had the
additional IS cells seen at 25 hours also
been selectively responsive at earlier
times? To answer these questions,
individual IMM neurons were tested
after one or two hours of imprinting,
as in the earlier study, but then were
followed for an additional 15 hours,
allowing additional testing after 7.5 and
15 hours. Furthermore, two groups
of chicks were studied; in an
attempt to identify a critical period
of sleep-dependent consolidation of
imprinting memories, one group of
chicks was deprived of sleep during
the first 7.5 hour post-imprinting
interval and a second during the
subsequent 7.5 hours period.
There were several striking results.
found 7.5 hour post-imprinting, whether
chicks had been allowed to sleep or
were sleep-deprived (Figure 1B). Thus,
at first blush it would appear that sleep
during this period was not important.
But, in fact, the opposite was true. After
the second 7.5 hour interval, those
chicks which had been allowed to sleep
during the first 7.5 hour interval (but
sleep deprived during the second
interval) showed a large increase in
IS neurons, while those who were
sleep-deprived during the first interval
showed a decrease in IS neurons. Thus,
the increase seen during the second
period was contingent on the chicks
sleeping during the first interval! (The
possibility of the increase being due to
sleep deprivation during the second
interval is eliminated by the earlier
findings of a similar increase with no
The second striking result was how
this absolute increase in the sleep-first
group came about. By the end of the
first 7.5 hour post-imprinting period,
all of the IS neurons identified after
the first hour of training had become
unresponsive (Figure 1C). In contrast,
all of those recruited during the second
hour of training remained responsive,
along with almost half of the previously
unresponsive neurons. Finally, after
the second 7.5 hour post-imprinting
period, two-thirds of the neurons
originally responsive after the first
hour of imprinting regained their
responsiveness (Figure 1C)!
These findings, relevant not just
to sleep-dependent memory
consolidation, but probably to all
consolidation processes, suggest that
the neuronal changes that underlie
such consolidation can ebb and flow
over periods of hours to at least a day,
during which time populations of
neurons can shift from responsive to
unresponsive and back. The functional
significance of such fluctuations will
IS neurons (%)IS neurons (%)
T T01 T2S1S2
Figure 1. Change in IS neurons over time.
(A) Number of responding cells before imprinting (0 hour), after 1 or 2 hours of imprinting, and at 25 hours after no further imprinting. Black bar,
IS neurons prior to imprinting; green bars, previously responsive neurons; red bars, previously unresponsive neurons; blue bar, prior history
unknown (based on ). (B) Percent responding cells before (T0, based on ) and after 1 or 2 hours of imprinting (T1, T2), and 7.5 or 15 hours
after the end of imprinting (S1, S2). Green squares, chicks who slept during the first 7.5 hour post-imprinting period; red triangles, chicks who
slept during the second 7.5 hour post-imprinting period. (C) Initial and subsequent IS neurons in sleep first condition: Number of IS neurons
identified at each time (same as in B). Green bars, IS neurons also active after 1 hour of imprinting (T1); red bars, IS neurons not also active
after 1 hour of imprinting (based on ).
Current Biology Vol 18 No 10
remain a frustrating unknown until
considerably more research is done.
1. Stickgold, R., Whidbee, D., Schirmer, B.,
Patel, V., and Hobson, J.A. (2000). Visual
discrimination task improvement: A multi-step
process occurring during sleep. J. Cogn.
Neurosci. 12, 246–254.
2. Frank, M.G., Issa, N.P., and Stryker, M.P.
(2001). Sleep enhances plasticity in the
developing visual cortex. Neuron 30, 275–287.
3. Jackson, C., McCabe, B.J., Nicol, A.U.,
Grout, A.S., Brown, M.W., and Horn, G. (2008).
Dynamics of a memory trace: effects of sleep
on consolidation. Curr. Biol. 18, 393–400.
of memory. Nat. Rev. Neurosci. 5, 108–120.
5. Horn, G., Nicol, A.U., and Brown, M.W. (2001).
Tracking memory’s trace. Proc. Natl. Acad. Sci.
USA 98, 5282–5287.
6. Stickgold, R. (2005). Sleep-dependent memory
consolidation. Nature 437, 1272–1278.
7. Ganguly-Fitzgerald, I., Donlea, J., and
Shaw, P.J. (2006). Waking experience affects
sleep need in Drosophila. Science 313,
8. Smith, C. (1995). Sleep states and memory
processes. Behav. Brain Res. 69, 137–145.
9. Nishida, M., and Walker, M.P. (2007). Daytime
naps, motor memory consolidation and
regionally specific sleep spindles. PLoS ONE 2,
10. Huber, R., Ghilardi, M.F., Massimini, M., and
Tononi, G. (2004). Local sleep and learning.
Nature 430, 78–81.
11. Peigneux, P., Laureys, S., Fuchs, S.,
Collette, F., Perrin, F., Reggers, J., Phillips, C.,
Degueldre, C., Del Fiore, G., Aerts, J., et al.
(2004). Are spatial memories strengthened in
the human hippocampus during slow wave
sleep? Neuron 44, 535–545.
12. Walker, M.P., Stickgold, R., Alsop, D., Gaab, N.,
and Schlaug, G. (2005). Sleep-dependent motor
memory plasticity in the human brain. Neurosci.
13. Wilson, M.A., and McNaughton, B.L. (1994).
Reactivation of hippocampal ensemble
memories during sleep. Science 265, 676–679.
14. Dave, A.S., and Margoliash, D. (2000). Song
replay during sleep and computational rules for
sensorimotor vocal learning. Science 290,
15. Manoach, D.S., Cain, M.S., Vangel, M.G.,
Khurana, A., Goff, D.C., and Stickgold, R.
(2004). A failure of sleep-dependent
procedural learning in chronic, medicated
schizophrenia. Biol. Psychiat. 56, 951–956.
16. Bushey, D., Huber, R., Tononi, G., and Cirelli, C.
(2007). Drosophila Hyperkinetic mutants have
reduced sleep and impaired memory. J.
Neurosci. 27, 5384–5393.
17. Vyazovskiy, V.V., Cirelli, C., Pfister-
Genskow, M., Faraguna, U., and Tononi, G.
(2008). Molecular and electrophysiological
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Department of Psychiatry, Harvard Medical
School and Center for Sleep and Cognition,
Beth Israel Deaconess Medical Center,
Boston, Massachusetts, USA.
Sexual Dimorphism: Can You Smell
A powerful new technique for visualizing neurons in the fly brain has uncovered
fine neuroanatomical differences between the olfactory circuitries of male and
Elizabeth J. Rideout
and Stephen F. Goodwin*
Dramatic behavioral differences exist
between males and females in
Drosophila [1–3]. Although the genetic
basis for the creation of two
morphologically and behaviorally
distinct sexes has been extensively
studied, few anatomical differences in
the brain have been identified which
may explain these dimorphic behaviors
[4–7]. In fact, gross anatomical studies
suggest that male and female brains
are largely similar . Identifying
potential differences requires detailed
analyses of the neural circuits
underlying these sex-specific
behaviors. Currently, identifying and
tracing the projections of single
neurons or populations of neurons
relies upon clonal analysis techniques,
which can be time-consuming due to
the stochastic nature of generating
uniquely labelled neurons [8,9].
Now, Datta et al.  have described
a novel system for visualizing the cell
bodies and projections of single
neurons with high spatial and temporal
resolution, using a photoactivatable
green fluorescent protein (PA-GFP)
. PA-GFP is a stable
photoactivatable variant of GFP that
after irradiation with 413 nm light
shows a 100-fold increase in
fluorescence when excited by 488 nm
light . Significantly, when PA-GFP
is photoactivated in an isolated region
the entire cell. Datta et al.  exploited
this property of PA-GFP to locate cell
bodies of neurons, and to visualize
axonal projections of identified
neurons. Using this elegant technique,
the authors were able to visualize and
identify sexually dimorphic axonal
projections of a specific population
of neurons in the brain.
During courtship, male and female
flies exchange a variety of stimulatory
and inhibitory sensory cues .
Intriguingly, certain auditory and
olfactory cues elicit opposite
behavioral responses from the two
sexes [13,14]; for example,
cis-vaccenyl acetate, a male-specific
pheromone, has been shown to have
an inhibitory effect on male courtship,
whereas in females, cis-vaccenyl
acetate enhances receptivity to
copulation . How can a single
pheromone elicit such different
behavioral responses? Datta et al.
 addressed this question by
investigating whether anatomical
and/or functional differences in
olfactory neural circuitry may explain
this dimorphic behavior (Figure 1).
In flies, the physiological response
to cis-vaccenyl acetate is mediated by
a class of olfactory sensory neurons
expressing the odorant receptor gene
Or67d. All neurons expressing Or67d
innervate a single glomerulus, DA1,
in the antennal lobe . Although the
size of the DA1 glomerulus is sexually
dimorphic [5,14,15], Datta et al. 
found no essential differences
between males and females in
either the increase of intracellular
Ca2+concentration or in the
electrophysiological responses in the
DA1 glomerulus following exposure
to cis-vaccenyl acetate, suggesting
that the neurobiological basis for the
sex-specific responses to cis-vaccenyl
acetate must lie elsewhere in the
olfactory neural circuit.
Next, using PA-GFP, they
determined that projection neurons
which project from the DA1 glomerulus
to the lateral horn, a higher olfactory
processing centre, have sexually
dimorphic axonal arbors on the lateral
horn (Figure 1). After identifying these
sexually dimorphic arbors, Datta et al.
 went on to show that this
dimorphism depends on the