Download Topography of Modular Subunits in the Mushroom Bodies of the

THE JOURNAL OF COMPARATIVE NEUROLOGY 399:153–161 (1998)
Topography of Modular Subunits in the
Mushroom Bodies of the Cockroach
MAKOTO MIZUNAMI,1* MASAYUKI IWASAKI,2 RYUICHI OKADA,1
AND MICHIKO NISHIKAWA2
1Laboratory of Neuro-Cybernetics, Research Institute for Electronic Science,
Hokkaido University, Sapporo 060–0812, Japan
2Faculty of Science, Fukuoka University, Fukuoka 814–0180, Japan
ABSTRACT
The mushroom body (MB), a conspicuous neuropil structure in the insect brain, is
implicated in associative memory and in some aspects of motor control. Intrinsic neurons of
the MB (Kenyon cells) extend dendrites into the calyx, and their axons run through the
pedunculus and then bifurcate to form the a and the b lobes. At the pedunculus and the lobes,
Kenyon cells make synaptic connections with dendrites of extrinsic (output) neurons.
Previously, we reported that the a lobe of the cockroach MB consists of repetitive modular
subunits (Mizunami et al. [1997] Neurosci. Lett. 229:153–156). Each subunit is composed of a
dark layer and a light layer, and the layers are refereed to as slabs. Each slab is composed of
axons of a specific subset of Kenyon cells. In the present study, we examined serial sections of
reduced silver preparations and found that each dark and light slab continues throughout the
length of the pedunculus and the a and b lobes. We also found that Golgi-impregnated Kenyon
cells often exhibit a characteristic grouping, forming a thin sheet interlaced by dozens or
hundreds of axons. The sheet is much thinner than the slab, and each sheet remains within a
particular slab throughout the length of the pedunculus and the lobes. Thus, the sheet is a
component forming the slab. In the pedunculus and the b lobe, a class of Golgi-impregnated
extrinsic neurons exhibit segmented dendritelike arbors that interact with every other slab,
i.e., either with only dark or light slabs. Because each neuron of this class interacts with each
particular set of dark or light slabs, we conclude that the slabs are units for transmitting
output signals from the MB. J. Comp. Neurol. 399:153–161, 1998. r 1998 Wiley-Liss, Inc.
Indexing terms: insect; brain; neuroanatomy; associative memory; higher center
The insect brain contains a pair of prominent neuropils
referred to as mushroom bodies (MBs). In the honey bee,
the establishment of olfactory memory is related to normal
functioning of MBs during the consolidation period following olfactory associative learning (Erber et al., 1980). In
the fly, mutants with chemical or structural deficits in MBs
exhibit poor learning performance (Heisenberg et al.,
1985; Nighorn et al., 1991; de Belle and Heisenberg, 1994;
Davis, 1996). In the cockroach, surgical ablation of MBs
resulted in poor performance in navigation based on visual
place memory, and observations of the activity of its
extrinsic neurons of the lobes while the cockroach is
walking freely suggest that the MB is involved in the
control, and possibly the planning, of locomotory actions
(Mizunami et al., 1993).
The MB neuropil has four distinct parts, i.e., calyces,
pedunculus, and the a and b lobes (Fig. 1A). In the calyces,
axon terminals of input neurons, the majority of which
derive from the deutocerebrum (Weiss, 1974; Mobbs, 1982)
and some from the protocerebrum (Nishikawa et al., 1998;
r 1998 WILEY-LISS, INC.
Nishino and Mizunami, 1998; Yamazaki et al., 1998), make
synapses onto dendrites of intrinsic neurons, called Kenyon
cells (Steiger, 1967; Schürmann, 1974; Strausfeld, 1976).
Axons of Kenyon cells project anteroventrally to the pedunculus and then bifurcate to form two lobes: the dorsally
projecting a lobe and the medially projecting b lobe (Fig.
1A). At the pedunculus and lobes, Kenyon cells make
synapses onto dendrites of extrinsic (output) neurons
(Schürmann, 1970; Frontali and Mancini, 1970). Axons of
extrinsic neurons project into different areas of the brain
(Mobbs, 1984; Rybak and Menzel, 1993), including lateral
Grant sponsor: Naitoh Foundation; Grant sponsor: Narishige Zoological
Science Award; Grant sponsor: Ministry of Education, Science, Culture and
Sports of Japan.
*Correspondence to: Dr. Makoto Mizunami, Laboratory of NeuroCybernetics, Research Institute for Electronic Science, Hokkaido University, Sapporo 060-0812, Japan. E-mail: [email protected]
Received 29 December 1997; Revised 6 May 1998; Accepted 10 May 1998
154
M. MIZUNAMI ET AL.
Fig. 1. A: Camera lucida reconstruction of a Golgi-impregnated
Kenyon cell in serial frontal sections of a cockroach brain. The neuron
gives off arborizations in the calyx (C), and its axon runs anteroventrally through the pedunculus (Ped) and then bifurcates; one branch
runs dorsally through the a lobe and the other runs medially through
the b lobe. B: Horizontal section of the a lobe in a reduced silverstained brain. Fifteen dark (D) and 15 light (L) slabs alternate in this
individual. Dark and light slabs are numbered 1–15 from posterior to
anterior (A). M, medial. Scale bars 5 200 µm in A, 20 µm in B.
proto- and deutocerebri from which descending neurons
originate to supply thoracic motor centers (Li and Strausfeld, 1997).
Each MB of the cockroach Periplaneta americana contains 200,000 Kenyon cells (Neder, 1959), the largest
number reported thus far for insect MBs: the reported
number of Kenyon cells in each MB is 170,000 in the honey
bee Apis (Witthöft, 1967), 50,000 in the cricket Acheta
(Schürmann, 1973), 50,000 in the locust Schistocerca
(Leitch and Laurent, 1996), 21,000 in the fly Musca
(Strausfeld, 1976), and 2,500 in the fly Drosophila (Balling
et al., 1987). Our recent study using reduced silver stains
showed that 15 modular subunits, each consisting of a pair
of dark and light slabs, are present in the a lobe of the
cockroach (Mizunami et al., 1997). By comparing reduced
silver and Golgi preparations, we concluded that each
modular subunit is formed from the axons of a specific
subset of Kenyon cells. It was unclear, however, if the
modular organization is restricted to the a lobe or is also
present in other output neuropils. In the present study, we
report that these modular subunits are maintained
throughout the length of the output neuropils, i.e., the
pedunculus and the a and b lobes. In addition, smaller
groupings of Kenyon cells, which we refer to as sheets, are
also maintained throughout the length of the output
neuropils.
MATERIALS AND METHODS
Studies were done on adult male and female cockroaches, Periplaneta americana, from a colony raised in
Hokkaido University. The cockroaches were maintained at
25–27°C and at 12-hour light–dark cycles. For Golgi
impregnations (Strausfeld, 1980), each cockroach was
anesthetized by cooling with ice, and the head was removed and mounted, dorsal side up, on a small dish. A
small piece of cuticle on the front of the head was removed
to expose the brain. The head was immersed in 2.5%
potassium dichromate for 1 hour, and the brain was
dissected out. The isolated brain was then transferred to a
bath containing five parts 2.5% potassium dichromate and
one part 25% glutaraldehyde (Polysciences, Tokyo, Japan)
for 4–7 days in the dark at 4°C. The tissues were subsequently treated by one of two methods: (1) the brains were
immersed in 0.75% silver nitrate for 3–4 days at 4°C; or (2)
the brains were washed in 2.5% potassium dichromate for
1 hour, followed by 4–7 days of incubation in 100 parts of
2.5% potassium dichromate and one part of 1% osmium
tetroxide for 6–10 days at 4°C, and then the brains were
immersed in 0.75% silver nitrate for 4–7 days at 4°C. The
brains were dehydrated through a graded series of ethanol, immersed in propylene oxide, and embedded in soft
Araldite (Nisshin EM, Tokyo, Japan). Serial sections were
cut at 50–110 µm in the three major planes, frontal,
MODULAR SUBUNITS IN THE COCKROACH MUSHROOM BODIES
155
horizontal and sagittal, with respect to the axis of the
head. For reduced silver impregnation, the heads were
isolated and mounted in a small dish, and the brains were
exposed by removing a piece of cuticle. The heads were
immersed in cockroach saline (Yamasaki and Narahashi,
1959) containing 3% paraformaldehyde for 1 hour, and
then the brains were dissected out. The isolated brains
were fixed in a solution containing 4% paraformaldehyde,
5% glacial acetic acid, and 85% ethanol for 2 days, dehydrated, and embedded in paraffin. The original (Bodian,
1936) and a variation (Otsuka, 1962) of the Bodian reduced silver impregnation method were used for 12-µm
sections. For staining with toluidine blue/azure II, the
brains were fixed in ethanol for 4 days and embedded in
Araldite. Serial sections were cut at 12 µm and briefly
stained with a mixture of 1% toluidine blue and 1% azure
II. The specimens were rinsed, dehydrated with a graded
series of ethanol, cleared in xylol, and coverslipped under
Canada balsam. The preparations were observed under
Nomarski interference contrast (Olympus, Tokyo, Japan),
real-time three-dimensional (Edge Scientific Instruments,
Santa Monica, CA), and conventional microscopes. A high
magnification observation at 1,0003 was made by using
an oil-immersion objective lens (1003). Trajectories of
stained cells were traced by using a camera lucida.
RESULTS
Topography of modular subunits in the MB
Observations of reduced silver preparations of the cockroach brain have shown that there are repetitive modular
subunits, each consisting of a pair of dark and light slabs
in the a lobe of the MB (Fig. 1B; Mizunami et al., 1997).
More or less similar slablike striations have been described in some anatomical accounts of the cockroach MB
(Hanström, 1928; Howse, 1975; Weiss, 1981; Li and Strausfeld, 1997), but their detailed morphological features have
not been reported. Comparisons of reduced silver and
Golgi preparations have shown that different slabs are
formed by axons of different subsets of Kenyon cells
(Mizunami et al., 1997). In the cross section of the a lobe,
the slabs at the posterior region are arched, with the
largest curvature at the posterior end, and those at the
anterior region are straight (Fig. 1B).
The number of modular subunits differs among individuals; the typical number seen in about 60% of individuals is
15 and the minimum and maximum numbers observed
among 24 cockroaches are 14 and 19, respectively. The
number of modular subunits in the right and left MBs,
however, is the same for all individuals examined thus far.
No prominent difference was found in the number of
subunits between males and females. The MBs of juveniles
exhibited fewer modular subunits: juveniles with a body
length of 8–10 mm exhibited 9–12 subunits and those with
a body length of 12–18 mm exhibited 13–14 subunits. This
observation indicates an increase in the number of modular subunits during ontogenetic processes.
To describe the modular structures quantitatively, the
averaged width at the center (Fig. 2A) and the area (Fig.
2B) of each of the dark and light slab was measured in
cross sections of the a lobe. We set a specific assumption to
assess borders between neighboring slabs: the borders
were set so that the total area of 15 dark slabs was equal to
that of 15 light slabs. Measurements were made at each of
three locations of the right and left a lobes in serial
Fig. 2. The width (A) and the area (B) in the cross section of slabs,
measured at the a lobe. Dark (D) and light (L) slabs are numbered
1–15 from posterior to anterior. Border lines between neighboring
dark and light slabs are defined by assuming that the total area of 15
dark slabs is equal to that of 15 light slabs. The widths at the center
and the areas were measured at three locations of the left and right a
lobes in serial horizontal sections of a reduced silver-stained brain,
and averages and standard deviations of six measurements are shown.
horizontal sections of a reduced silver-stained brain. Dark
(D) and light (L) slabs are numbered 1–15 from posterior to
anterior. The width of slabs is smallest in the median part
(3–4 µm) and larger in the anterior or posterior parts. The
widths of the slabs at the posterior end (1L) and anterior
end (15D) are approximately 15 µm and 20 µm, respectively. The area in the cross section is the smallest at the
posterior end and largest at the anterior end: the area of
each of the last four slabs at the anterior end (14L–15D) is
about three times that of most posterior slabs (Fig. 2B).
This observation suggests that anterior slabs contain a
much larger number of axons of Kenyon cells.
Figure 3 shows serial horizontal sections of the MB in a
reduced silver-stained brain. Although no prominent slablike laminar structures are visible in the calyx (Fig. 3A),
concentrically arranged dark and light slabs alternate at
the base of each lateral and medial calyx (Fig. 3B). Tracing
of individual Kenyon cell axons in Golgi preparations show
that the concentric arrangement of dark and light slabs
156
Fig. 3. A-E: Serial horizontal sections of the mushroom body of a
reduced silver-stained brain showing topography of modular subunits.
A: In the calyx, no slablike striations are visible. B: Sets of alternating
dark and light slabs are concentrically arranged at the base of medial
and lateral calyces. Two sets of concentric slabs uncurl at the head of
the pedunculus (P) and fuse edgewise to form flattened slabs as they
M. MIZUNAMI ET AL.
descend through the pedunculus (C). M and L are axons of Kenyon
cells derived from medial calyx (MC) and lateral calyx (LC), respectively. Flattened slabs continue throughout the length of the a (A–D)
and b (E) lobes. The anterior side is at the top; the lateral side is at the
right. Scale bar 5 50 µm.
MODULAR SUBUNITS IN THE COCKROACH MUSHROOM BODIES
157
Fig. 4. Two serial horizontal sections, one just dorsal to (A) and the other at (B) the junction between
the pedunculus and lobes of a brain stained with toluidine-blue/azure II, showing that each dark and light
slab of the pedunculus (P) continues to each dark and light slab of the a lobe. The anterior side is at the
top. Scale bar 5 50 µm.
reflects a concentric arrangement of axons of Kenyon cells
in the calyx (Mizunami et al., 1998). Concentric slabs from
each calyx uncurl at the head of the pedunculus and fuse
edgewise to form single flattened slabs as they descend
through the pedunculus. Arrays of flattened slabs are
maintained throughout the pedunculus and a and b lobes.
A similar pattern of striation is visible in plastic sections
stained with toluidine blue/azure II (Fig. 4) or osmiumethyl gallate, in paraffin sections stained with hematoxylin–eosin, and by Nomarski interference contrast observation of unstained sections. We attempted to examine the
cause of the different staining intensity between dark and
light slabs by a high magnification observation (1,0003) of
these preparations, but no convincing conclusions were
attained, apparently due to the limitation of the resolution
of light microscopy.
Observations of serial horizontal sections showed that
each dark or light slab of the pedunculus is continuous
with a corresponding dark or light slab in the a lobe (Fig. 4)
and the b lobe (not shown); thus, each dark or light slab
keeps its dark or light appearance over the entire extent in
the output neuropil. In Figure 5, the topography of modular subunits is schematically drawn on the basis of observations of serial sections. Each slab continues throughout the
length of the pedunculus and a and b lobes, thereby
indicating that each slab consists of a particular subset of
Kenyon cell axons. Note that bifurcation of slabs reflects
bifurcation of individual axons of Kenyon cells (Fig. 1A).
Extrinsic neurons with segmented dendritic
arborizations
A class of Golgi-stained extrinsic (output) neurons of the
b lobe have segmented dendritelike spiny arborizations,
the interval and the width of segments corresponding to
the width of slabs (Fig. 6A). The segments are maintained
throughout the dendritic domain of the neuron. This
Fig. 5. Schematic representation of the topography of modular
subunits in the pedunculus (Ped) and the a and b lobes, based on
observations of serial horizontal, sagittal, and frontal sections of
reduced silver-stained brains. Bifurcation of each dark and light slab
at the junction between the pedunculus and lobes reflects bifurcation
of axons of Kenyon cells constituting the slabs. A, anterior; P,
posterior; D, dorsal; V, ventral; M, medial; L, lateral.
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M. MIZUNAMI ET AL.
Fig. 6. Examples of Golgi-stained extrinsic neurons whose dendritelike arbors exhibit prominent segmentation, observed in cross sections
of the b lobe (A) and the pedunculus (B). A is a sagittal section and B is
a horizontal section. Axons of Kenyon cells (not shown) run perpendicular to the plane of the sections. A: Dendritic arbors of the extrinsic
neuron exhibit at least nine segments, the width and intervals of
which match those of slabs. B: Dendritic arbors of the neuron exhibit
approximately 14 segments (arrowheads), each of which covers less
than half of the width of each slab. The dorsal side is at the top in A,
and the anterior side is at the top in B. Scale bars 5 30 µm.
suggests that extrinsic neurons of this class receive synaptic inputs from Kenyon cells in every other slab, i.e., in only
dark slabs or light slabs. Four of 17 Golgi-impregnated
extrinsic neurons in the b lobe have segmented dendritic
arborizations. We reported that about 35% of output
neurons in the a lobe exhibit similar segmentation (Mizunami et al., 1997). In the pedunculus, three of 11 Golgiimpregnated extrinsic neurons possess segmented dendritic arbors. In all three neurons, the dendritic
arborizations cover only less than half of the thickness of
each slab, thus interacting with the axons of only less than
half of Kenyon cells that constitute each slab (Fig. 6B).
Different extrinsic neurons with segmented dendrites appear to interact with different sets of dark or light slabs:
dendrites of some neurons cover up to 13–14 slabs, whereas
those of other neurons cover only a few particular slabs.
Thus, we suggest that the slabs represent units to transmit output signals from the MB.
In Figure 8, axons of Kenyon cells consisting of a sheet
are traced to the cell body region and the synaptic neuropil
of the calyx in serial horizontal sections. Their cell bodies
form at least three separate clusters in the anterolateral
(arrow in Fig. 8A), anterior (arrowhead), and anteromedial
(not shown) parts of the cell body region dorsal to the
lateral calyx, and their dendritic arborizations occupy
anterolateral, anterior, and anteromedial parts of the
synaptic neuropil of the lateral calyx, respectively (Fig.
8A,B). Axons of each group form a bundle as they converge
to the center of the calyx, and the three bundles run side by
side at the base of the calyx (Fig. 8B, arrows). Another
axonal bundle derived from the posteromedial part of the
lateral calyx then joins these bundles (not shown); the cell
bodies could not be located. These four axon bundles are
flattened and fuse together to from a thin sheet at the head
of the pedunculus (Fig. 8C). The sheetlike structure is
maintained throughout the length of the a (Fig. 8C–E) and
b (not shown) lobes. The lateral extent of the sheet covers
half of the width of the pedunculus and covers the entire
width of the a and b lobes (Fig. 8C–E), indicating that
axons of lateral and medial calyces are mixed together as
they descend to the lobes. A comparison with serial horizontal sections of reduced silver preparations suggests that
the sheet is located in slab 9D or 10L on its entire extent in
the pedunculus and the a and b lobes. Thus, we consider
the sheets to be elements that constitute the slabs.
Topography of sheets in the MB
Axons of Golgi-impregnated Kenyon cells often exhibit a
characteristic grouping, a thin mesh- or sheetlike structure, which we refer to as a sheet (Fig. 7A,B). The sheet in
the a lobe appears as an array of axons when observed in a
horizontal section (Fig. 7A) and as a meshwork interlaced
by dozens or hundreds of Kenyon cell axons when observed
in a frontal section (Fig. 7B). Figure 7B shows a sheet
wider than 120 µm, indicating that the sheet is extremely
flat and thin. Each axon of an individual Kenyon cell
remains within a particular sheet for its entire extent in
the pedunculus and the a and b lobes. Observations of
sheets in cross sections of the a lobe show that the sheets
are flat only in the anterior and medial parts (Fig. 7B) and
that they are curved in the posterior part, the largest
curvature being at the posterior end. Thus, the form of the
sheets precisely agrees with that of slabs at that location,
except that the sheets are much thinner than slabs. The
number of Kenyon cell axons counted in each sheet differed
from 40 to 280 (n 5 30), the large variance possibly due to
the capricious nature of the Golgi stain.
DISCUSSION
We previously reported that repetitive modular subunits, each consisting of a pair of a dark and a light slab,
are present in the a lobe of the MB of the cockroach
(Mizunami et al., 1997). Each slab represents the axons of
a particular subset of Kenyon cells. We now report that
each slab continues throughout the length of the output
neuropils. Some extrinsic neurons have a characteristic
segmented dendrites: these segments have been shown to
represent interactions with specific slabs. In addition,
slabs are now shown to contain smaller elements, referred
MODULAR SUBUNITS IN THE COCKROACH MUSHROOM BODIES
159
Fig. 7. A: Axons of Golgi-impregnated Kenyon cells observed in a horizontal section of the a lobe, near the junction between the a and b lobes.
Axons of Kenyon cells are arranged in rows. B: Golgi-stained Kenyon cell axons at the a lobe, observed in a frontal section. A thin meshwork, or
sheet, formed by a number of Kenyon axons can be seen. The anterior side is at the top in A, and the dorsal side is at the top in B. Scale bars 5 20
µm.
to as sheets, that arise from smaller subsets of Kenyon
cells. The cause of the different staining intensities between dark and light slabs could not be determined in the
present study; this is the subject of an electron microscopic
study (Iwasaki et al., unpublished observations).
Topography of modular subunits in the MB
At the base of the calyx, alternating dark and light slabs
are arranged concentrically like growth rings of a tree, and
this concentric arrangement is converted into a linear
array of flattened slabs in the pedunculus, as was described by Weiss (1981). The concentric arrangement of
slabs at the base of the calyx reflects concentric arrangements of Kenyon cell axons in the calyx, as is discussed in
an accompanying paper (Mizunami et al., 1998).
Each slab differs in shape, thickness, and area in the
cross section. The area in the cross section of a slab at the
anterior part is three times larger than that of a slab at the
posterior end, suggesting that the anterior slabs consist of
a larger number of Kenyon cells. In an accompanying
paper (Mizunami et al., 1998), we describe how Kenyon
cells can be classified into four types, each of which forms a
respective set of modular subunits. We speculate that each
of the modular subunits and slabs may have specific
morphological features that reflect different functional
roles.
A class of extrinsic (output) neurons of the pedunculus
and b lobe possesses segmented dendritelike arbors that
interact with every other slab, i.e., with only dark or light
slabs. Similar neurons have been observed in the a lobe
(Mizunami et al., 1997), indicating that this class of
extrinsic neurons is distributed throughout output neuropils. Different extrinsic neurons cover different sets of dark
or light slabs, suggesting that the slabs are units for
transmitting output signals from the MB. Extrinsic neurons with segmented dendritic arbors of the pedunculus
cover less than half of the width of each slab. This coverage
appears to correspond to substructures of the slabs (discussed below).
Modular organizations of the MB may not be restricted
to cockroaches. In the honey bee, Mobbs (1982) reported
that there are seven layers in the pedunculus and lobes
and that each layer represents a different morphological
type of Kenyon cell originating from a different concentric
zone of the calycal neuropil. Some of these layers exhibit
Phe-Met-Arg-Phe-NH2 (FMRF amide)-like immunoreactivity (Schürmann and Erber, 1990) or immunoreactivity to
type II cAMP-dependent protein kinase (Müller, 1997).
Dendrites of extrinsic neurons of the a lobe of the honey
bee also exhibit a prominent layered arrangement (Mobbs,
1982; Rybak and Menzel, 1993). In the fruit fly Drosophila,
subdivision of the MB by several different types of Kenyon
cell was found when patterns of gene expression were
examined (Yang et al., 1995) and ablation of a specific type
of Kenyon cell resulted in a defect in sex-specific courtship
behavior, whereas that of other types did not (O’Dell et al.,
1995). Weiss (1981) observed a number of layers in the
pedunculus and lobes in the MB of several species of
Orthoptera by reduced silver stainings. These modular
structures may be a common design of insect MBs.
Topography of sheets in the MB
In Golgi preparations, the axons of Kenyon cells are
often seen to take the form of a very thin sheet in the
pedunculus and the a and b lobes. This arrangement
suggests a nonrandom nature of our Golgi stains: the
probability that axons constituting the same sheet are
concurrently impregnated appears to be high in our Golgi
procedure. Bundles of axons arising from several groups of
Kenyon cells, which may occupy separate areas of the cell
body region and may send dendritic processes to distinct
regions of the calycal neuropil, flatten and fuse together at
Fig. 8. A–F: Serial horizontal sections showing groups of Golgiimpregnated Kenyon cells whose axons form a sheet. Their cell bodies
are located in anterolateral (arrowheads), anterior (arrowheads), and
anteromedial (not shown) parts of the cell body region dorsal to the
lateral calyx (LC; A), and their dendrites cover anterolateral, anterior,
and anteromedial portions of the synaptic neuropil of the lateral calyx
(A,B). The axons of these cell groups form a bundle (arrows in A,B) and
are arranged side by side at the base of the calyx (B). As the axonal
bundles descend to the head of the pedunculus (P), they are flattened
and fuse together to form a thin sheet (arrows in C–E). The sheet
continues throughout the length of the a (C–E) and b lobes (not
shown). The anterior side is at the top; the medial side is at the right.
Scale bar 5 50 µm.
MODULAR SUBUNITS IN THE COCKROACH MUSHROOM BODIES
the head of the pedunculus. The sheet is much thinner
than the slab, and each sheet appears to remain in a
particular slab for its entire extent in the output neuropils,
suggesting that the sheet is a component forming the slab.
It remains unclear as to whether all Kenyon cells are
organized into sheets or some Kenyon cells form sheets but
others run solitary without forming a sheet. If the former
is the case, the sheet can be regarded as a smaller modular
substructure of the MB. A variation in the periodic pattern
of modules has been noted (Li and Strausfeld, 1997) and
will be explained by a different organization of substructures (Iwasaki et al., unpublished observations).
The functional significance of the sheetlike structure
remains to be clarified. One possibility is that Kenyon cells
constituting each sheet make synaptic connections with
each other. Frontali and Mancini (1970) noted in their
electron microscopic study that Kenyon cells make synapses with each other in the a lobe of the cockroach. In the
locust, Kenyon cells exhibit synchronous and oscillatory
activity when olfactory stimulation is applied to the antenna. Reciprocal synaptic interactions among Kenyon
cells may play a role in producing this synchronized
electrical activity (Laurent and Naraghi, 1994). It would
be interesting to investigate whether each subset of Kenyon
cells constituting each sheet exhibits similar synchronous
activity in the cockroach.
ACKNOWLEDGMENTS
We thank Dr. N.J. Strausfeld for helpful discussion,
especially for pointing out the possibility that slabs represent radial arrangements in the calyces. The advice helped
us to interpret our results correctly. M. Ohara provided
helpful comments.
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