RESEARCH NOTES
290
& Trueman, R.R. (eds). Academic Press, San
Diego, p. 169-184.
14. ARKHIPKIN, A.I. 1989. Ph.D. Thesis, Institute of
Oceanography of the USSR Academy of
Sciences, Moscow, 134 p. (in Russian).
15. JACKSON, G.D. 1993a. Fish. Bull U.S., 91:
260-270.
J. MolL Stud. (1997), 63, 290-293
16. JACKSON, G.D. 1994. Can. J. Fish. Aquat Sci.,
51: 2612-2625.
17. LAPTIKHOVSKY, V.V. 1991. BioL nauki, 1991 (3):
37-48 (in Russian).
18. JACKSON, G.D. 1993b. Can. J. Fish. Aquat. Sci.,
50: 2365-2374.
© T7ie Malacological Society of London 1997
Changes in mantle muscle structure associated with growth and reproduction in
the tropical squid Photololigo sp. (Cephalopoda: Loliginidae)
Natalie Moltschaniwskyj
Department of Marine Biology, James Cook University of North Queensland, Townsville, Queensland,
Australia 4811
Squid mantle muscle tissue is predominantly made
up of circular smooth muscle fibres, separated into
blocks by thin (•• 30 u.m) bands of radial muscle
fibres.1 These smooth muscle fibres have helical
myofibrils and are typically small; < 10 jun
diameter.2 In many cephalopods circular muscle
fibres are present in two structural states; mitochondria-rich and mitochondria-poor, analogous to
fast and slow twitch muscle fibres in vertebrates.3
The bulk (~ 80%) of the mantle muscle tissue is
made up of circular, mitochondria-poor muscle
fibres. Growth of the mantle muscle occurs by the
production of new muscle fibres and growth of existing fibres/ Changes in the size structure of mantle
muscle fibres and mantle muscle blocks are very
extensive during growth of the animal/1-3
Processes of senescence and reproduction can
change the structure and appearance of the mantle
muscle tissue. There is evidence that the mantle
muscle tissue changes both in composition6-7 and
structure19 during egg production; although there are
exceptions.10-" One of the most dramatic changes in
the muscle tissue has been recorded in senescent
cephalopods, in which the muscle fibres break down
completely leaving only collagenfibres.12-13Low lipid
levels have been recorded in cephalopods and it is
speculated that energy reserves maybe stored as
protein or carbohydrate" in the muscle tissue, hence
the changes observed.13
In this paper I describe the presence of nodes of
disorganised circular muscle fibres in Photololigo sp.
mantle muscle tissue. I then examined the relationship between the presence and extent of the nodes
and the size and reproductive status of the animals.
Photololigo sp. is an inshore tropical loliginid
squid species found along the central Queensland
coast of Australia. The species used in this study is
the most common in this region and in the past has
been referred to by the specific name chinensis
(Gray, 1849).16-" However, recent taxonomic work
indicates that chinensis is the incorrect specific
name.18 As this is one of two sibling species in the
region, all individuals in this study were identified
as being the same species using allozyme electrophoresis." Photololigo sp. lives for approximately
120 days and growth continues at a constant rate
throughout its life.16
Fifty-one juvenile Photololigo sp. (2.5-49 mm
dorsal mantle length) were caught using automated
light-traps1' in the Townsville (North Queensland,
Australia) region between October and January of
the 1991/92 austral summer. Ninety-eight adult
Photololigo sp. (60-150 mm dorsal mantle length)
were caught in the same area using pair otter trawls
(40 mm mesh) between August and November 1991
and in March 1992.
Dorsal mantle length (mm) of each individual was
recorded before mantle muscle tissue was fixed for
histological analysis. Juveniles were fixed whole 'and
a sample of muscle tissue was removed later. Adults
were killed by chilling and a sample of dorsal mantle
muscle tissue was taken anteriorly, level with the
locking mechanism. All muscle tissue was fixed in a
formalin-acetic acid-calcium chloride solution (10 ml
37% formaldehyde, 5 ml glacial acetic acid, 1.3 g
calcium chloride (dihydrate); distilled water to 100
ml). Fixed tissue was transferred to 70% ethanol 48
hrs before processing in paraffin wax. Muscle tissue
was dehydrated through an ascending isopropanol
series, cleared in chloroform and infiltrated with
paraffin wax (Paramat). Tissue blocks were sectioned at 7 jim, decerated in xylene and hydrated
through a descending ethanol series. Histological
sections were stained with Mallory-Heidenhain
trichrome stain. Sections of muscle tissue were
examined at 160x and 400x. Muscle tissue was
sectioned longitudinally, so circular muscle fibres, ie.
those muscle fibres that encircle the muscle mass,
were cut transversely and radial muscle fibres longitudinally.4
RESEARCH NOTES
The muscle tissue in cephalopods is organised so
that the muscle fibres run roughly parallel to one
another, along the same plane (Fig. la). However, in
the mantle muscle of 69 adults not aH of the muscle
fibres were orientated along the circular axis of the
body. Instead, there appeared to be a breakdown in
the organisation of both the circular and radial
muscle fibres (Fig. lb). This disorganisation of the
muscle fibres presented itself as nodes or discrete
areas in the mantle muscle tissue (Fig. lc). When the
nodes were present in low numbers they were concentrated near the internal or external edges of the
mantle muscle. When the nodes were large and
extensive the breakdown in organisation extended
throughout the mantle muscle. The extent of the
disorganisation ranged from a few scattered nodes
through to most of the muscle fibres in the mantle
being disorganised. The state of disorganisation
(muscle status) was rated on a quantitative scale;
State 0 not present, State 1 < 10% of muscle tissue
occupied by disorganisation, State 2 10-30%, State 3
31-80%, and State 4 81-100%.
Disorganisation of mantle muscle fibres was
absent in juvenile squid mantle muscle tissue. Therefore only data from adults was statistically analysed.
Multiway frequency analysis20 was used to examine
the relationship among the factors; size, reproductive
maturity (from immature, I to mature, V) and
muscle state of the squid. The amount of disorganisation was dependent on the size of the individual
and level of reproductive maturity (Table 1). The
non-significant Likelihood Ratio in Table 1 suggests
that the main effects (size and reproductive maturity) provided a good fit to the data. This meant that
the factors size and reproductive maturity were independent of one another. Therefore, the relationship
between each of the main effects and muscle status
could be examined separately.
Disorganisation was not present in individuals less
than 50 mm in dorsal mantle length. However, as
individuals grew there was an increasing chance of
having some disorganisation present (Fig. 2). The
proportion of individuals not having any nodes
present decreased from 37% in the 60-89 mm size
class to 14% in the largest size class (> 109 mm).
However, larger individuals did not necessarily have
more disorganisation present (Fig. 2). All of the size
classes had individuals with extensive disorganisation present. Therefore, the presence of disorganisation in the mantle muscle tissue was size dependent,
but the extent of disorganisation was not.
Muscle status was dependent on reproductive
status. Of the immature and maturing individuals
(Stage I, II & III) examined 55% had nodes of disorganised tissue present (Fig. 3). More than 80% of
Stage IV and V individuals, who by this stage have
dedicated considerable resources to reproduction,
had disorganised muscle fibres present (Fig. 3).
However, there was no evidence that these very
reproductive individuals had more nodes present in
the mantle muscle tissue.
The presence of what appears to be disorganised
circular muscle fibres in adult squid has not yet been
291
Figure 1.
a) Normal circular mantle muscle fibres from the
anterior part of the mantle. The circular (c)
muscle fibres are cut transversely and the radial
(r) muscle fibres cut longitudinally. This section
was stained with Mallory-Heidenhain trichrome
stain (Scale bar = 50 \im)
b) A node of disorganised circular muscle fibres.
(Scale bar = 50 u,m)
c) Nodes seen in approximately 10% of the mantle
muscle tissue. (Scale bar = 500 jim)
recorded in the literature. This is possibly because
past studies have examined relatively few individuals, or it was considered an artefact of tissue processing. Examination of the fibres with the light
microscope indicated that they were intact (Fig. lb)
292
RESEARCH NOTES
Table 1. Results of a multiway frequency analysis examining the muscle state of each individual and the relationship between size and
reproductive status.
Source
df
Reproductive state 2
Mantle length
3
Likelihood ratio
3
Chi-square Probability
13.69
34.97
6.15
0.0011
0.0000
0.1047
and not in the process of cellular breakdown,
although this needs to be confirmed with electron
microscopy. Handling of all muscle tissue samples
was identical and muscle tissue was not frozen. All
specimens were dead when tissue was removed and
fixed, so there is no suggestion that dead material is
fixing differently from live tissue. The very patchy
occurrence of disorganisation in some of the individuals further suggests that this is not a fixation effect;
for example fixative failing to penetrate the tissue. If
fixative had failed to penetrate the tissue, then any
effect on muscle fibres would have been in the
central region. However, early stages of disorganisation were only evident along the internal and external margins of the mantle muscle. If fixative had
affected fibres near the surfaces, then muscle tissue
of juvenile squid should have been similarly, if not
more affected, given the thinness of their mantle
muscle. Although the presence of disorganised
muscle fibre arrangement cannot be definitively
separated from fixative and processing artefacts, the
very obvious absence from juvenile muscle tissue
suggests that it is a real phenomenon. The presence
of the nodes has been more recently found in adult
Sepia elliptic and Idiosepius pygmaeus*
All the individuals used in this study appeared
to be externally healthy because no lesions were
present on the body surface. No obviously senescent
squid were captured, and there is no information
60-69 mm
n=28
<
1
1
2
3
MUSCLE STATE
Figure 2. Size frequency distribution of adults in
each of the mantle muscle state classes.
2
3
MUSCLE STATE
Figure 3. Percentage frequency of individuals in
each classification of muscle status by reproductive
stage.
RESEARCH NOTES
about the nature of muscle tissue changes in
Photololigo sp. during senescence. Dying squid (eg.
Moroteuthis ingens) can undergo very dramatic
changes in muscle tissue in which the muscle fibres
are completely absent and just the collagen structure
remains.13 Likewise, the muscle fibres of senescent
Octopus vulgaris break down leaving spaces in the
tissue.12 This phenomenon was not present in any of
the Photololigo sp. tissue samples examined. Furthermore, the relatively poor relationship between
size, reproductive status and amount of disorganisation suggests that this phenomenon may not be
directly due to either senescence or reproductive
activities.
The presence of disorganised muscle fibres in the
mantle muscle may affect the swimming abilities of
the squid. Circular muscle fibres are responsible for
providing the power stroke in the flight response, by
forcing water out of the siphon.1 The radial fibres
provide the force to restore the mantle back to the
resting shape.1 It may be envisaged that co-ordination of the muscle fibres to provide the force for the
jet propulsion may be impaired by a breakdown in
the organisation of the fibres.
J. Yeatman and C.C. Lu assisted with the identification of adult and juvenile Photololigo sp. specimens. PJ. Doherty allowed me access to juvenile
cephalopods caught using light-traps. B. Molony,
J.H. Choat, G.D. Jackson, L. Winsor. B. Kier and
the reviewers provided constructive comments and
discussion. This work was supported by a Merit
Research Grant from James Cook University and
was carried out whilst the author was a Commonwealth Scholar at James Cook University.
293
3. MOMMSEN, T.P., BALLANTYNE, J-, MACDONALD,
D., C-OSUNE, J. & HOCHACHKA, P.W. 1981.
Proc NatL Acad. ScL USA, 78: 3274-3278.
4. MOLTSCHANIWSKYJ, N.A. 1993. Can. J. Fish.
AquaL Sci., 51: 830-835.
5. PECL, G. 1994. BSc(Hons.) Thesis, James Cook
University of North Queensland.
6. O'DOR, R.K. & WELLS, M J. 1978. / . exp. Bioi,
77:15-31.
7. POLLERO, RJ. & IRIBARNE, O.O. 1988. Comp.
Biochem. PhysioL, 90B: 317-320.
8. MOLTSCHANIWSKYJ, N.A. 1995. Mar. Biot, V2A:
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9. HATFIELD, E.M.C., RODHOUSE, P.G. & BARBER,
D.L. 1992. /. mar. bioL Ass. U.K., 72: 281-291.
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© The Malacological Society of London 1997
/. Moll. Stud. (1997), 63,293-296
Effect of temperature on embryonic development of two freshwater pulmonates,
Planorbarius comeus (L.) and Planorbis planorbis (L.)
K. Costil
Laboratoire de Zoologie et Ecophysiologie (U.A. INRA & U.M.R. du C.N.R.S. 6553), University de Rennes 1,
Campus de Beaulieu, Av. du Ciniral Leclerc, 35042 Rennes Cedex, France
In freshwater Pulmonates, the embryonic development is direct and takes place in eggs. Eggs, which
comprise a zygote surrounded by perivitelline fluid
and membrane, are embedded in jelly and enclosed
in a common egg capsule. The juveniles leave the
egg capsule using their radula to gnaw the surrounding membranes. Planorbarius comeus and Planorbis
planorbis are freshwater snails commonly found in
Brittany (France) where they belong to relatively
rich communities.1 In our region, P. planorbis shows
an annual life cycle with two generations per year,
whereas the P. corneus cycle tends to be longer
and more variable according to year.2 In experimental populations of these species, we already showed