Effects of Dorsal and Ventral Vertical Lobe Electrolytic Lesions on Spatial
Learning and Locomotor Activity in Sepia officinalis
Nicolas Graindorge, Christelle Alves, Anne-Sophie Darmaillacq, Raymond Chichery, Ludovic Dickel, and
Ce ´cile Bellanger
Universite ´ de Caen Basse-Normandie
This study aims to analyze the effects of electrolytic lesion, restricted to either the ventral or the dorsal
parts of the vertical lobe (VL), on the behavior of cuttlefish (Sepia officinalis). Two behavioral tests were
performed on sham-operated and lesioned cuttlefish: assessment of locomotor activity in an open field
and determination of spatial learning abilities in a T maze. The results showed that ventral lesions of the
VL led to marked impairment in the acquisition of spatial learning, whereas dorsal lesions of the VL
increased locomotor activity in the open field and impaired long-term retention of spatial learning. This
study establishes for the first time the existence of distinct functions in the ventral and the dorsal parts
of the VL in cephalopods.
Keywords: cephalopods, electrolytic lesion, vertical lobe, locomotor activity, spatial learning
Over the last decades, researchers have undertaken extensive
investigations to gain a functional understanding of brain struc-
tures related to behavior. Part of these studies has focused on
integrated functions, such as learning and memory. From an evo-
lutionary and comparative standpoint, understanding the neural
basis of different types of learning and memory across a wide
range of taxa is essential. To this purpose, numerous neuroetho-
logical studies have been performed in invertebrates, particularly
arthropods (for a review, see Strausfeld, Hansen, Li, Gomez, & Ito,
1998) and mollusks (e.g., Aplysia; for a review, see Kandel, 2003).
Given their relatively simple nervous systems, they provide pow-
erful model systems for the study of neuronal substrates and
cellular mechanisms underlying learning and memory processes.
squids, and octopuses) exhibit very high behavioral flexibility
from predatory to defensive behavior as well as interindividual
communication (for a review, see Hanlon & Messenger, 1996).
They also display good memory ability in a wide range of learning:
passive avoidance (Agin, Chichery, Dickel, & Chichery, 2006;
Messenger, 1971), taste aversion (Darmaillacq, Dickel, Chichery,
Agin, & Chichery, 2004), discriminative learning (for reviews, see
Boal, 1996; G. D. Sanders, 1975), reversal learning (Mackintosh &
Mackintosh, 1964), spatial learning (Boal, Dunham, Williams, &
Hanlon, 2000; Karson, Boal, & Hanlon, 2003; Mather, 1991),
observational learning (Fiorito & Scotto, 1992), and imprinting
(Darmaillacq, Chichery, & Dickel, in press; Darmaillacq, Chich-
ery, Shashar, & Dickel, 2006). These behavioral abilities are
controlled by a highly developed and centralized nervous system,
which consists of about 100–200 million neurons (for a review,
see Budelmann, 1994). The gut passes through this central nervous
system and divides it into supra- and suboesophageal masses, lying
between two large optic lobes. Cephalopods are valuable alterna-
tive models situated between the inferior vertebrates and simpler
invertebrates, like Aplysia, and appear to be pertinent models for
comparative neurobiological studies of learning and memory.
Numerous studies in cephalopods have established the involve-
ment of the vertical lobe complex in cognitive processes (for
reviews, see G. D. Sanders, 1975; Young, 1991). This complex,
situated in the extreme dorsal part of the supraoesophageal mass,
consists of three lobes closely interconnected: the vertical (VL),
superior frontal, and subvertical lobes (see Figure 1). At the front
of the vertical lobe complex, the superior frontal lobe shows two
distinct parts: the anterior superior frontal lobe, which receives
inputs from the VL, and the posterior superior frontal lobe, which
sends all its axons to the VL. The subvertical lobe is positioned
behind the superior frontal lobe and below the VL. This structure
receives axons from and sends axons to the two other lobes of the
vertical lobe complex. The superior frontal and subvertical lobes
are centers of sensory multiconvergence, sending pretreated visual
and tactile information to the VL. Numerous neuroethological
experiments in Octopus vulgaris have shown impairment in its
ability to acquire and retain associative tasks when the VL has
been removed (for a review, see G. D. Sanders, 1975). Young
(1979) has described in detail the VL in Loligo, but the general
Nicolas Graindorge, Christelle Alves, Anne-Sophie Darmaillacq, Ray-
mond Chichery, Ludovic Dickel, and Ce ´cile Bellanger, Laboratoire de
Physiologie du Comportement des Ce ´phalopodes, Universite ´ de Caen
Basse-Normandie, Caen Cedex, France, and Centre de Recherche en En-
vironnement Co ˆtier, Universite ´ de Caen Basse-Normandie, Luc-sur-Mer,
This work was supported by a grant from the Ministe `re de
l’Enseignement Supe ´rieur et de la Recherche to Christelle Alves. We thank
J. Lejeune and J. L. Durand for statistical advice, J. Boal for valuable
suggestions concerning the article, J. Harris and C. Harris for helping to
correct the English, the Centre de Recherche en Environnement Co ˆtier staff
for their technical assistance, and the Institut National des Sciences de
l’Univers du Centre National de la Recherche Scientifique for providing
Correspondence concerning this article should be addressed to Ce ´cile
Bellanger, Laboratoire de Physiologie du Comportement des Ce ´phalopo-
des, Universite ´ de Caen Basse-Normandie, 14032 Caen, Cedex, France.
2006, Vol. 120, No. 5, 1151–1158
Copyright 2006 by the American Psychological Association
organization is strikingly similar in cuttlefish. This dome-shaped
structure consists of cell layers (cortex) surrounding a neuropil
consisting of networks of fibers (Young, 1979). Young described
two parts in the VL: the central VL corresponding to the whole
middle part of the dome and, surrounding it, the peripheral VL.
The cortices of these two parts are characterized, respectively, by
a fine cell layer and a thick cell wall. In contrast, the neuropils of
these two parts appear widely continuous. The cortex of the
peripheral VL is mainly composed of two types of cells: large
scattered cells with axons passing down through the neuropil to
converge and end in the subvertical and anterior superior frontal
lobes, and very numerous amacrine cells with axons restricted to
the VL. In the dorsal part of the VL, a large tract of fibers coming
from the posterior superior frontal lobe is situated just below the
cortex. The VL also contains smaller cells scattered throughout the
neuropil with axons ending in the subvertical lobe. Cells lying
along the ventral part of the VL are similar to the amacrine cells of
the dorsal cortex. Thus, in summary, the dorsal part of the VL
receives the superior frontal lobe tract, whereas the ventral part of
the VL, near the subvertical lobe, is characterized by the presence
of the VL-subvertical lobe tracts (inputs and outputs) and the
VL-superior frontal lobe tract (see Figure 1).
In cuttlefish, only one study has shown that long-term memory
recall of an associative learning is impaired after the ablation of the
VL (F. K. Sanders & Young, 1940). However, the ablation dam-
aged the lobes surrounding the VL, and this study was conducted
on a very small sample (only 2 individuals) without a control (i.e.,
sham-operated cuttlefish). Taking into account the anatomical
structure of the VL and the localization of the main connections
within the VL, we applied ventral and dorsal lesions of the VL to
study the possible existence of functional dissociation within the
VL. Previous studies have already used electrolytic lesions in
cuttlefish (Chichery & Chichery, 1987). This method of lesion was
chosen to ensure good survival rates and good reproduction of the
size of the lesion. We used two original behavioral tests currently
used in vertebrates in lesioned and sham-operated cuttlefish: as-
sessment of locomotor activity in an open field and of spatial
learning abilities in a T maze (Alves, Chichery, Boal, & Dickel, in
press). This test appears more appropriate for subjects caught in
the wild than the one described by Karson et al. (2003). Indeed,
preliminary experiments seemed to show that the reinforcement
used in Karson’s test is more efficient with cuttlefish born and bred
Subadult cuttlefish (Sepia officinalis; 130–160 mm dorsal mantle length,
N ? 21) were trawled in September 2004 in the vicinity of Luc-sur-Mer,
France, and housed individually in glass tanks (80 ? 50 ? 50 cm) with
circulating seawater at 18 ? 1 °C. The home tanks were maintained on a
12-hr light–dark cycle (lights on at 8:30 am). They were fed shrimp
(Crangon crangon) and crabs (Carcinus maenas) ad libitum. After at least
1 week of acclimatization to laboratory conditions, two groups of cuttlefish
received an electrolytic lesion of the VL: ventral VL lesioned cuttlefish
(n ? 5, Ev group) and dorsal VL lesioned cuttlefish (n ? 6, Ed group). The
control group was made up of sham-operated cuttlefish (n ? 10, S group).
After surgery, the cuttlefish were allowed a 4-day recovery period before
the start of behavioral testing. Eight days after surgery, all of the cuttlefish
were anesthetized and killed, and their supraoesophageal mass was pro-
cessed for histological examination. The time schedule of the experiments
is summarized in Figure 2.
Surgery: VL Electrolytic Lesions
S. officinalis were anesthetized through placement for 90 s in seawater
containing ethanol (2%) with magnesium chloride (17.5 g/L) to prevent
any muscular contraction during surgery. The cuttlefish were held in a
frame during the operation so that we could obtain approximately the same
angular position of the head. This procedure minimizes the variation of the
angular penetration of the electrode in the supraoesophageal mass. An
incision was made above the cranial cartilage. The position of the sagittal
plane and the position of the VL were localized with external morpholog-
ical characteristics of the cranium (see Chichery & Chichery, 1987). A
unipolar enamel-coated stainless electrode 0.12 mm in diameter, uninsu-
lated at the tip for approximately 1 mm, was fixed through a small oval
plate of inert plastic. The electrode was inserted through the cartilage into
the VL. The length of the electrode below the plate (between 5 and 7 mm)
was adjusted according to the dorsal or ventral lesion required (ventral VL,
Ev group, or dorsal VL, Ed group) and the size of the cuttlefish. The lesion
was made by passing a current of 1 mA for 60 s through the electrode.
Sham operations consisted of reproduction of all the steps of surgery with
the exception that no current was delivered through the electrode (S group).
The electrode was then removed and the incision closed up with Histoacryl
(B. Braun Medical, Boulogne, France). Surgery time was about 10 min.
After completion of surgery, cuttlefish were replaced in their home tank,
where they recovered normal activity (lying on the bottom of their tank)
within the 5 min following surgery.
Behavioral tests were performed in experimenter-blind conditions.
Locomotor Activity Test: Open Field
The open field apparatus consisted of a white PVC tank, 80 cm square
and 15 cm deep, filled to the top with seawater and lit by a 300-W halogen
lamp located 1 m above the surface of the water. Twenty-four hours before
surgery (OF1; see Figure 2), each cuttlefish was gently removed from its
home tank and placed in the center of the open field, of which it had no
Sepia officinalis with the localization of the main neural pathways in the
vertical complex. VL ? vertical lobe complex; FS ? superior frontal lobe;
FSpost ? posterior superior frontal lobe; FSant ? anterior superior frontal
lobe; SV ? subvertical lobe.
Schematic representation of the supraoesophageal mass of
GRAINDORGE ET AL.
previous experience. Locomotor activity of the subject in this new envi-
ronment was videotaped for 15 min with a digital video camera (3CCD
DCR-TRV950E, Sony, Paris, France). Then, the cuttlefish was replaced in
its home tank. This test was performed for a second time 96 hr after surgery
(OF2; see Figure 2). For analysis of locomotor activity, a transparent grid
overlay was matched with the image of the video recordings on a TV
screen. The number of squares crossed by each cuttlefish was counted for
each open field test (OF1 and OF2), and the equivalent traveled distances
were calculated. To normalize variation between individual cuttlefish, we
compared the locomotor activity of each cuttlefish in OF2 with its own
locomotor activity in OF1. Thus, OF2 traveled distance was expressed as
a percentage of OF1 traveled distance for each individual. The open field
experiments were conducted between 9 am and 12 pm for all cuttlefish.
Spatial Learning: T Maze
cm and 30 cm high, with internal divisions forming a T-shaped maze (see
Figure 3). The stem of the T, 40 ? 40 cm, served as the start box; the
identical arms of the T (80 ? 30 cm) led to the goal compartments Ca and
Cb, situated at either side of the stem. Both goal compartments were
darkened, each with a sliding PVC top, and the bottom was completely
covered with sand 1 cm deep. The tank was filled to a depth of 20 cm with
seawater maintained at a temperature of 18 ? 1 °C. Sliding doors allowed
The apparatus consisted of a white plastic tank, 200 ? 70
the cuttlefish to be confined either in the start box or in one of the goal
compartments. A 300-W halogen lamp was positioned 1 m above the
surface of the water. The testing room contained various visual extramaze
cues (e.g., water-pipes, sets of shelves).
Preliminary observation of cuttlefish kept in
laboratory tanks has indicated that they usually avoid open, lit areas when
they cannot bury. In this experiment, only one goal compartment (Ca or
Cb) of the T maze was opened throughout training, and the cuttlefish had
to learn how to escape from the light by finding the entrance to this dark
and sandy goal compartment. Five days after surgery, cuttlefish were given
three training sessions of five trials each, with 1 hr between consecutive
sessions. Between trials, the water was stirred to avoid any use of olfactory
cues by the cuttlefish in finding the goal compartment and to reduce water
heating. During the first trial of the first session, both goal compartments
were closed to determine each cuttlefish’s side-turning preference. During
the remaining trials, only the compartment situated at the end of the arm
not chosen during the first trial was open (goal compartment).
During training trials, each cuttlefish was placed in the start box for 15 s
before its sliding door was removed. The cuttlefish was given a maximum
of 10 min to reach the end of one arm. Each trial allowed only a single
choice of direction (right or left arm). If the cuttlefish entered the goal
compartment, the sliding door was closed to prevent it from returning to the
maze. The cuttlefish was then allowed to remain on the sandy bottom, in
the dark, for 15 min. After that, it was gently replaced in the start box. If
the subject entered the incorrect arm and consequently failed to reach the
goal compartment it was immediately removed and replaced in the start
box. The error criterion was the movement of any part of the cuttlefish
beyond the virtual line at the far end of the incorrect arm (see Figure 3).
Between two consecutive sessions, the cuttlefish was maintained in the
goal compartment. After the three training sessions in the experimental
apparatus, the cuttlefish was gently replaced in its home tank. Twenty-four
hours after the end of the three training sessions, an additional session of
five trials was performed for assessment of long-term retention of this task
(see Figure 2). The performance of each cuttlefish was expressed as a
percentage of correct choices within each session of five trials.
After completion of behavioral testing, all of the cuttlefish were anes-
thetized (cf. section Surgery: VL Electrolytic Lesions) and quickly killed by
decapitation. The supraoesophageal masses were removed, fixed for 24 hr
in 0.2 M phosphate buffer 4% paraformaldehyde, and transferred into a 0.2
M phosphate buffer 20% sucrose solution for 12 hr. They were then
quickly frozen and stored at ?80 °C until used for histology. Each
2 - 1
T maze task
24-hr retention test
Schematic representation of the experimental time schedule. OF ? open field.
spatial learning experiment (not drawn to scale). Ca, Cb ? goal compart-
ments; S ? start box.
Schematic representation of the T maze apparatus used for the
VERTICAL LOBE ELECTROLYTIC LESIONS IN SEPIA
complete supraoesophageal mass was sliced in 20 ?m sagittal sections on
a freezing microtome. Sections were stained with hematoxylin eosin.
Localization and extent of the lesion were studied with a computer
coupled to a Leitz-Aristoplan microscope (Leica, Wetzlar, Germany),
using the program Biocom Visiolab 200 (Biocom, Les Ulis, France). To
determine the extent of the lesions, we sampled one section in five, starting
from the beginning of the lesion, which corresponds to an average of ten
sections per cuttlefish. In addition, the average volume of the VL was
calculated over the whole structure following the same procedure, which
corresponds to an average of sixteen sections per cuttlefish. To make
precise comparisons of VL volumes, we defined the exact extent of the VL
as follows: anteriorly, from the appearance of the neuropil in the antero-
lateral part of the VL, and posteriorly, as far as the disappearance of the
neuropil (for details, see Dickel, Chichery, & Chichery, 1997). The coef-
ficient of variation in VL volume estimation was 0.31% (determined from
ten measurements per cuttlefish).
All data were analyzed with nonparametric statistical tests, computed
with StatXact (Cytel Inc., Cambridge, MA). For comparisons of locomotor
activity within each group (before and after surgery), we used exact
permutation tests for paired samples (Siegel & Castellan, 1988). To com-
pare the variation of locomotor activity after surgery between the three
groups, we used exact permutation tests for unpaired samples. p values
were adjusted to an overall ? ? .05 level by use of the sequential
Bonferroni method as described by Holm (1979). For each group, percent-
ages of correct choices in the last session of acquisition and in the 24-hr
retention test of spatial learning were compared with their respective
percentage of correct choices in the first session of acquisition by use of
exact permutation tests for paired samples.
Fine microscopic histological observations of the VL of S group
cuttlefish indicated that the small diameter of the electrodes did not
visibly damage the nervous tissue. The lesioned area was charac-
terized by a substantial necrosed tissue surrounded by intense cell
proliferation. The schematic localization of lesions in Ed and Ev
groups is shown in Figures 4 and 5.
The Ev group exhibited spherical lesions situated in the ventral
part of the VL neuropil and the ventral cortex of the VL (see
Figures 4a and 5a). In Cuttlefish 1, the lesion extended to the
anterior part of the subvertical lobe and the posterior part of the
superior frontal lobe. The average volume of ventral VL lesions
was 1.80 ? 0.16 mm3(mean ? SEM).
The Ed group exhibited a semispherical lesioned area situated in
the dorsal neuropilar and cortical zones of the VL, along the
anterior-posterior axis (see Figures 4b and 5b). All of these lesions
damaged mainly the peripheral VL. The average volume of dorsal
VL lesions was 0.89 ? 0.14 mm3. No signs of damage were
observed in the lobes surrounding the VL. For guidance, the
average volume of the VL was 30.08 ? 1.53 mm3.
No obvious behavioral impairment was observed in lesioned
cuttlefish in laboratory tanks. All cuttlefish fed within 2 days post
Locomotor Activity Test: Open Field
Four days after surgery (OF2), locomotor activity displayed by
the S and Ev group cuttlefish was not significantly different from
their respective activity 1 day before surgery (OF1; exact permu-
tation test for paired samples, S group: n ? 10, p ? .89; Ev group:
n ? 5, p ? .687). Conversely, the locomotor activity of Ed group
increased significantly between OF1 and OF2 (exact permutation
test for paired samples, Ed group: n ? 6, p ? .031).
Through use of the sequential Bonferroni method (Holm, 1979),
pairwise comparisons of locomotor activity variation between the
three groups (expressed as a percentage of OF1 locomotor activity;
see Figure 6) showed no significant difference between the S and
Ev groups (exact permutation test for unpaired samples, sequential
Bonferroni method, S group: n ? 10, Ev group: n ? 5, p ? .408)
and significant increase of locomotor activity in the Ed group
when compared with the other two groups (exact permutation test
for unpaired samples, sequential Bonferroni method, S group: n ?
10; Ed group: n ? 6, p ? .021; Ev group: n ? 5; Ed group: n ?
6, p ? .004).
Spatial Learning: T Maze
As shown in Figure 7, percentages of correct choices were
significantly higher during Session 3 than during Session 1 in the
S and Ed groups (exact permutation test for paired samples, S
Sepia officinalis with the localization of vertical lobe (VL) lesions in the
ventral VL lesion (Ev) group and the dorsal VL lesion (Ed) group: dorsal
view of the supraoesophageal mass (upper schema) and sagittal section
(lower schema). Sections labeled 1–5 indicate individuals of the Ev group,
and sections labeled 6–11 indicate individuals of the Ed group. VLc ?
central part of the vertical lobe; VLp ? peripheral part of the vertical lobe;
SV ? subvertical lobe; FS ? superior frontal lobe.
Schematic representation of the central nervous system of
GRAINDORGE ET AL.
group: n ? 10, p ? .007; Ed group: n ? 6, p ? .031). Percentages
of correct choices were not significantly different between Session
1 and Session 3 in the Ev group (exact permutation test for paired
samples, Ev group: n ? 5, p ? 1.0). In the 24-hr retention test,
cuttlefish from the S group exhibited percentages of correct
choices significantly higher than during Session 1 (exact permu-
tation test for paired samples, S group: n ? 10, p ? .027).
However, in both Ev and Ed groups percentages of correct choices
were not significantly different between Session 1 and the reten-
tion test (exact permutation test for paired samples, Ev group: n ?
5, p ? .125; Ed group: n ? 6, p ? .906).
The S group did not show any modification of locomotor
activity in the open field or any sign of impairment in either the
ventral VL lesion (Ev) group and the dorsal VL lesion (Ed) group at 8 days post lesion (Hematoxylin eosin
staining, sagittal sections of the central nervous system), and sagittal sections of the central nervous system. Scale
bars ? 200 ?m. B: Schematic reconstructions of typical damage to the VL from Cuttlefish 4 in the Ev group and
Cuttlefish 9 in the Ed group. Shaded areas indicate the extent of a typical lesion on a sagittal section of the central
nervous system. Intersection windows ? 200 ?m. SV ? subvertical lobe; FS ? superior frontal lobe.
A: Photomicrographs of vertical lobe (VL) electrolytic lesions from representative subjects of the
VERTICAL LOBE ELECTROLYTIC LESIONS IN SEPIA
acquisition or the 24-hr retention of spatial learning. Thus, the
behavioral impairments observed in the Ev and Ed groups were not
due to any side effects of anesthesia or surgery. Behavioral results
within the Ev and Ed groups appear widely homogeneous, al-
though the lesions were localized along the anterior-posterior axis
of the VL. Even if the subvertical and the superior frontal lobes
were partially damaged in Cuttlefish 1, behavioral impairments
were comparable to the other cuttlefish of the Ev group. Ventral
lesions of the VL did not induce any modification of locomotor
activity in the open field but led to marked impairments in spatial
learning acquisition. The performance of Ev group cuttlefish in the
24-hr retention test does not therefore lead us to any conclusions
concerning the influence of ventral VL lesions on long-term re-
tention abilities. On the other hand, dorsal lesions of the VL led to
increased locomotor activity in the open field and impaired the
24-hr retention of spatial learning, although no impairment in
acquisition was observed. Thus, the locomotor activity and spatial
learning tests appear particularly relevant in the examination of the
functional implications of the VL.
As previous studies have not shown any motor response to
electrical stimulation of the VL in cuttlefish (for a review, see
Boycott, 1961), this structure has not been considered as a higher
motor center. Our study provides the first evidence of VL impli-
cation in locomotor activity. Because the level of locomotor ac-
tivity in cuttlefish in their home tank was not tested during our
study, we cannot conclude that there was a general increase of
locomotor activity. It remains to be determined, therefore, whether
the VL plays a specific role in modulating locomotor activity level
or in stress. Previous experiments have showed that learning and
memory abilities were impaired after ablation of the VL in Octo-
pus (for a review, see G. D. Sanders, 1975). In those studies, the
associative learning tasks used were largely based on the inhibition
of predatory behavior. If the VL was an inhibitory center, as
suggested by G. D. Sanders (1975), it is possible that an increase
in general activity interfered with learning, making cuttlefish in-
capable of inhibiting their predatory behavior. In our study, cut-
tlefish with acquisition impairments (Ev group) did not display any
increase in locomotor activity, so the results do not appear to be
affected by any change in locomotor activity level.
The VL belongs to the vertical lobe complex, which is organized
by reexcitation loops involving the superior frontal and subvertical
lobes (for a review, see Young, 1991). This complex seems to be
implicated in learning and memory processes in cuttlefish. Studies
of ontogenesis and senescence have shown a marked respective
correlation between the appearance of the VL-subvertical lobe
tracts and the emergence of memory ability (Dickel et al., 1997),
and the degeneration of the VL-subvertical lobe tracts and the
impairment of memory ability (Chichery & Chichery, 1992). An
improvement in learning and long-term retention performance has
also been correlated with the postembryonic growth of the VL
(Dickel, Chichery, & Chichery, 2001; Messenger, 1973). Learning
and memory impairments observed in the Ev and Ed groups are
consistent with the implication of the VL in those processes in
Considering that the increase in locomotor activity was ob-
served only in the Ed group, in which the lesion extents were
smaller than in the Ev group, behavioral impairment does not
appear to be linked to lesion volume. Marked behavioral impair-
ments were associated with a relatively small extent of lesion
relative to that of the VL (ranging from 3% to 6% of the VL
volume). So it seems that ventral or dorsal VL functioning depends
on structural integrity. This could explain why the disruption of
information pathways at different points in the vertical lobe com-
plex network induces impairment at different stages in the pro-
cesses of learning and memory. Even though there is converging
evidence for VL involvement in both acquisition and retention
processes (for a review, see G. D. Sanders, 1975), less is known
about the intrinsic processes that allow this structure to elaborate,
store, and recall memory traces. Nevertheless, the cellular organi-
zation of the ventral and dorsal parts of the VL, and the morpho-
logical organization of afferent and efferent VL connections, could
be responsible for the functional heterogeneity of this structure.
Our study did not allow us to draw any conclusions concerning
functional dissociation between central and peripheral parts of the
VL. Furthermore, because various lesion methods could have
neural consequences outside the primary site of brain damage that
(% of OF1)
(Ed group) vertical lobe lesions on locomotor activity. Locomotor activity
in OF2 (4 days after surgery) is expressed as a percentage of locomotor
activity in OF1 (1 day before surgery). Error bars indicate standard errors
of the mean. The dotted line represents locomotor activity level in OF1
(100%). OF ? open field. *p ? .05 (exact permutation test for unpaired
The effect of sham (S group), ventral (Ev group), and dorsal
(Ed group) vertical lobe lesions on acquisition and retention performance
in spatial learning. Results are expressed as mean percentages of correct
choices (? SEM) over the three training sessions (S1, S2, and S3) and
during the 24-hr retention test (R). *p ? .05 (comparisons to their respec-
tive percentage of correct choices in the first session, exact permutation test
for paired samples).
The effects of sham (S group), ventral (Ev group), and dorsal
GRAINDORGE ET AL.
may not be readily observable (as shown in Glenn, Lehmann,
Mumby, & Woodside, 2005), it will be interesting for the behav-
ioral effects of electrolytic lesions to be compared with other
methods like excitotoxic or aspiration lesion.
In the Ed group, the posterior superior frontal lobe-VL tract was
substantially damaged in all cuttlefish. These lesions could have
disrupted information transfer between the posterior superior fron-
tal lobe and the VL. A previous study indicated that the posterior
superior frontal lobe is activated just after the acquisition of an
associative learning task but not during the 24-hr retention test
(Agin, Chichery, & Chichery, 2001). The authors have suggested
that this activation reflects changes of neuronal activity in this area
during consolidation processes in the memory. Hence, impairment
of 24-hr retention in the Ed group could be explained by the
involvement of both the VL cortex and the posterior superior
frontal lobe during consolidation.
In the Ev group, lesions damaged the ventral neuropil, the
connections between the VL and the subvertical lobe (inputs and
outputs), and the VL-superior frontal lobe tract. In Cuttlefish 1, the
low extent of VL neuropil lesion led to behavioral impairment
comparable with those of other cuttlefish from the Ev group. This
lesion also substantially damaged the anterior part of the subver-
tical lobe. For this reason, behavioral impairments observed in this
group may be due mainly to disruption of the VL-subvertical lobe
tracts and the VL-superior frontal lobe tract as well as, to a lesser
extent, damage to the ventral neuropil.
In further investigations, lesions of the superior frontal and the
subvertical lobes would be appropriate for better understanding of
the implications of the superior frontal lobe-VL tract and VL-
subvertical lobe tracts in the vertical lobe complex. VL subdivi-
sions possibly subserving different behavioral roles may be linked
to their respective connections. Such a functional dissociation in a
highly associative structure has been extensively studied in verte-
brates. Indeed, Moser and Moser (1998) reviewed anatomical and
behavioral evidence for functional differentiation between the dor-
sal (locomotor activity) and ventral (spatial memory) parts of the
hippocampus. Other studies have revealed functional subdivisions
of mushroom bodies in arthropods (for a review, see Strausfeld et
al., 1998; Yang, Armstrong, Vilinsky, Strausfeld, & Kaiser, 1995).
The hippocampus in vertebrates and the VL in cephalopods appear
to be constituted in sequences of matrices sharing the same prop-
erties (for a review, see Young, 1991). Furthermore, Hochner,
Brown, Langella, Shomrat, and Fiorito (2003) have shown that the
VL in octopuses shows two different types of mechanisms for LTP
induction as in different parts of the hippocampus. Then, VL and
hippocampus show structural analogy, similar cellular processes,
and comparable functional implications (locomotor activity level
and memory). That seems to show a striking example of evolu-
tionary convergence. Although each animal model presents its own
specific features, understanding how and where information is
encoded and transmitted in the central nervous system of cepha-
lopods will provide better understanding of the evolution of such
Agin, V., Chichery, R., & Chichery, M. P. (2001). Effects of learning on
cytochrome oxidase activity in cuttlefish brain. NeuroReport, 12, 113–115.
Agin, V., Chichery, R., Dickel, L., & Chichery, M. P. (2006). The “prawn-
in-the-tube” procedure in the cuttlefish: Habituation or passive avoid-
ance? Learning and Memory, 13, 97–101.
Alves, C., Chichery, R., Boal, J. G., & Dickel, L. (in press). Orientation in
the cuttlefish Sepia officinalis: response versus place learning. Animal
Boal, J. G. (1996). A review of simultaneous visual discrimination as a
method of training octopus. Biological Reviews, 71, 157–190.
Boal, J. G., Dunham, A. W., Williams, K. T., & Hanlon, R. T. (2000).
Experimental evidence for spatial learning in octopuses (Octopus bi-
maculoides). Journal of Comparative Psychology, 114, 246–252.
Boycott, B. B. (1961). The functional organization of the brain of the
cuttlefish Sepia officinalis. Proceedings of the Royal Society of London,
Series B, 153, 503–534.
Budelmann, B. U. (1994). Cephalopod sense organs, nerves and the brain:
Adaptations for high performance and life style. Marine and Freshwater
Behaviour and Physiology, 25, 13–33.
Chichery, M. P., & Chichery, R. (1987). The anterior basal lobe and control
of prey-capture in the cuttlefish Sepia officinalis. Physiology and Be-
havior, 40, 329–336.
Chichery, M. P., & Chichery, R. (1992). Behavioural and neurohistological
changes in aging Sepia. Brain Research, 574, 77–84.
Darmaillacq, A. S., Chichery, R., & Dickel, L. (in press). Food imprinting,
new evidence from the cuttlefish, Sepia officinalis. Biology Letters.
Darmaillacq, A. S., Chichery, R., Shashar, N., & Dickel, L. (2006). Early
familiarization overrides innate prey preference in newly hatched Sepia
officinalis cuttlefish. Animal Behaviour, 71, 511–514.
Darmaillacq, A. S., Dickel, L., Chichery, M. P., Agin, V., & Chichery, R.
(2004). Rapid taste aversion learning in adult cuttlefish, Sepia officinalis.
Animal Behaviour, 68, 1291–1298.
Dickel, L., Chichery, M. P., & Chichery, R. (1997). Postembryonic mat-
uration of the vertical lobe complex and early development of predatory
behaviour in the cuttlefish (Sepia officinalis). Neurobiology of Learning
and Memory, 67, 150–160.
Dickel, L., Chichery, M. P., & Chichery, R. (2001). Increase of learning
abilities and maturation of the vertical lobe complex during postembry-
onic development in the cuttlefish, Sepia. Developmental Psychobiol-
ogy, 39, 92–98.
Fiorito, G., & Scotto, P. (1992, April 24). Observational learning in
Octopus vulgaris. Science, 256, 545–546.
Glenn, M. J., Lehmann, H., Mumby, D. G., & Woodside, B. (2005).
Differential Fos expression following aspiration, electrolytic, or excito-
toxic lesions of the perirhinal cortex in rats. Behavioral Neuroscience,
Hanlon, R. T., & Messenger, J. B. (1996). Cephalopod behaviour. Cam-
bridge, England: Cambridge University Press.
Hochner, B., Brown, E. R., Langella, M., Shomrat, T., & Fiorito, G.
(2003). A learning and memory area in the Octopus brain manifests a
vertebrate-like long-term potentiation. Journal of Neurophysiology, 90,
Holm, S. (1979). A simple sequentially rejective multiple test procedure.
Scandinavian Journal of Statistics, 6, 65–70.
Kandel, E. R. (2003). The molecular biology of memory storage: A
dialogue between genes and synapses. Medicine Sciences, 19, 625–633.
Karson, M. A., Boal, J. G., & Hanlon, R. T. (2003). Experimental evidence
for spatial learning in cuttlefish (Sepia officinalis). Journal of Compar-
ative Psychology, 117, 149–155.
Mackintosh, N. J., & Mackintosh, J. (1964). Performance of Octopus over
a series of reversals of a simultaneous discrimination. Animal Behaviour,
Mather, J. A. (1991). Navigation by spatial memory and use of visual
landmarks in octopuses. Journal of Comparative Physiology A, 168,
Messenger, J. B. (1971). Two stage recovery of a response in Sepia.
Nature, 232, 202–203.
VERTICAL LOBE ELECTROLYTIC LESIONS IN SEPIA
Messenger, J. B. (1973). Learning performance and brain structure: A
study in development. Brain Research, 58, 519–523.
Moser, M. B., & Moser, E. I. (1998). Functional differentiation in the
hippocampus. Hippocampus, 8, 608–619.
Sanders, F. K., & Young, J. Z. (1940). Learning and other functions of the
higher nervous centres of Sepia. Journal of Neurophysiology, 3, 501–
Sanders, G. D. (1975). The cephalopods. In W. C. Corning, J. A. Dyal, &
A. O. D. Willows (Eds.), Invertebrate learning (Vol. 3, pp. 1–101). New
York: Plenum Press.
Siegel, S., & Castellan, N. J. (1988). Nonparametric statistics for the
behavioral sciences (2nd ed.). New York: McGraw-Hill.
Strausfeld, N. J., Hansen, L., Li, Y., Gomez, R. S., & Ito, K. (1998).
Evolution, discovery, and interpretations of arthropod mushroom bodies.
Learning and Memory, 5, 11–37.
Yang, M. Y., Armstrong, J. D., Vilinsky, I., Strausfeld, N. J., & Kaiser, K.
(1995). Subdivision of the drosophila mushroom bodies by enhancer-
trap expression patterns. Neuron, 15, 45–54.
Young, J. Z. (1979). The nervous system of Loligo.; Vol. V. The vertical
complex. Philosophical Transactions of the Royal Society of London,
Series B, 285, 311–354.
Young, J. Z. (1991). Computation in the learning system of Cephalopods.
Biological Bulletin, 180, 200–208.
Received December 13, 2005
Revision received May 30, 2006
Accepted June 7, 2006 ?
GRAINDORGE ET AL.