Watanabe and Cox - 2
Development in Mopalia
INTRODUCTION
The genus Mopalia (Mollusca, Polyplacophora) is
represented by 14 species along the California coast
(Burghardt and Burghardt, 1969). Among the common forms
in central California are Mopalia muscosa (Gould, 1846),
Mopalia lignosa (Gould, 1846), Mopalia ciliata (Sowerby,
1840), and Mopalia hindsii (Reeve, 1847). Though rela-
tively well known taxonomically, little is known of the
spawning behavior and larval development of this genus.
Heath (1905) made brief mention of spawning in M. lignosa
and M. muscosa in the field. Thorpe (1962) studied spawning
in M. lignosa, M. muscosa, M. ciliata, M. hindsii,
M. imporcata (Carpenter in Pilsbry, 1892), and M. porifera
(Pilsbry, 1892). He also made some general observations
on the larval development of M. ciliata. Comparable
information on larval development in other species of
this genus is not available. We chose to study development
in M. muscosa and M. lignosa in part to fill this gap and
because these species are common and known to spawn in the
spring (Heath, 1899; Boolootian, 1964). Our aim was to
determine the main sequence of events in larval development
and its time schedule. The development of M. muscosa was
followed by Cox, while that of M. lignosa was followed by
Watanabe. A strong effort was made to standardize ex-
perimental procedures and presentation of results so that
Development in Mopalia
Watanabe and Cox - 3
comparisons between the two would be valid and clear. The
work was carried out at Hopkins Marine Station of Stanford
University during the months of April-June, 1974.
METHODS AND MATERIALS
Specimens of Mopalia lignosa were collected under low
rocks at Mission Point, Carmel Bay, California. M. muscosa
were collected at the same location and from rocks in the
mid-tide zone of Point Pinos, Pacific Grove, California.
Heath (1905, p. 392) maintains that Katharina tunicata
(Wood, 1815) attains sexual maturity at two years, reaches
a length of 25mm the firstyear and adds 8-11mm the second
year. He states that this is also characteristic of both
M. lignosa and M. muscosa, therefore only larger animals
(50-60mm long) were taken, to insure sexual maturity.
Once in the laboratory, the chitons were maintainéd in
circulating seawater aquaria.
Embryos were reared in 5-inch finger bowls cleaned
with concentrated nitric acid followed by detergent and
thorough rinsing. Fertilized eggs were pipetted from the
aquaria (see Spawning), rinsed in fresh sea water several
times and placed in thin layers in bowls with one-half-inch
of sea water. Both natural sea water filtered through coarse
filter paper and artificial sea water (Instant Ocean Synthetic
Sea Salts) were used. Little difference was found between
the two and only filtered natural sea water was used for
embryos from the late free-swimming stage on. From 0.1 to
Watanabe and Cox - 4
Development in Mopalia
O.3ml of a mixture of 0.5g streptomycin and 300,000 units
of penicillin in one liter of distilled water was added to
the bowls to control bacterial growth. Water in the bowls
was changed by decanting off about one-half the volume of
water and pouring the remainder of the culture, containing
nearly all the larvae, into a clean finger bowl. This was
done every ?4 hours for the first 12 hours of development
and twice daily for the later stages. The finger bowls
were covered with glass plates and maintained partially
immersed in running sea water at 13.5-15.8°0. Around the
time of settling, eroded fragments-of Mytilus shell covered
with a green algal film were added to most of the bowls as
a settling surface.
Observations of the structure and behavior of living
larvae were made in finger bowls under a dissecting microscope
and in wet whole mounts under a compound microscope. For
permanent whole mounts, larvae were fixed in Bouin's fluid
and stained in dilute acidulated Grenacher's borax carmine
(Galigher and Kozloff, 1964). Due to the large amount of
yolk in the larvae, little internal differentiation could be
discerned.
SPAWNING
To obtain gametes, attempts were made to induce
spawning in the laboratory using methods which have produced
positive results in gastropods and other invertebrates. The
Development in Mopalia
Watanabe and Cox - 5
methods tried were the following (M.1.- Mopalia lignosa,
M.m.- Mopalia muscosa. Numerals preceding these indicate
numbers tested):
1. Dilation of gonopores by insertion of glass probe,
followed by placement of animals in standing seawater;
2 M.1., 1 M.m. No spawning in 1 hour.
Ref. Gould (1967) for Urechis.
Electrical stimulation of gonopores (15v Q 50 cycles AC
2.
for 5 sec.) followed by placement in standing seawater;
3 M.1., 2 M.m. No spawning in 1.5 hours.
Ref. Iwata (1950) for Mytilus edulis Linnaeus, 1758.
3. Electrical stimulation of exposed lateral nerve cord
(15v 2 50 cycles AC for 5 sec.) followed by placement in
standing seawater; 1 M.1., 1 M.m. No spawning in 1 hour.
Ref. Harvey (1956) for echinoids.
Injection of 0.2ml 0.5M KCl into perivisceral hemocoel
or through pallial groove followed by placement in stand-
ing seawater; 3 M.1., 3 M.m. Caused strong body contraction
in 3 minutes but no spawning in 4 hours.
Ref. Harvey (1939) for echinoids.
Injection of nerve tissue homogenate into perivisceral
hemocoel, followed by placement in standing seawater;
1 M.m. No spawning in 1 hour.
Ref. Davis, Mpitsos, and Pinneo (1973) for Pleurobranchaea.
Eggs dissected from ovaries and treated for one hour
with Zml O.1N NaOH in 100ml seawater, inseminated with
sperm obtained by dissection; 1 M.1., 1 M.m, Eggs not
fertilizable with active sperm.
Ref. Wolfsohn (1907) for Acmaea.
5.
Development in Mopalia
Watanabe and Cox - 6
7. Several ml sperm solution obtained from dissected male
gonad added to aquaria containing chitons; 7 M.1., 5 M.m.
No spawning in 5 hours. Ref. Heath (1905) that sperm re-
leased by males may cause female spawning.
8.
Water in tank containing chitons allowed to stand and
become stale. Temperature slightly elevated (15°c.):
7 M.1., 7 M.m. Spawning occurred though not consistently.
Ref. Grave (1932) for Chaetopleura apiculata (Say).
Still water at temperatures slightly above ambient
ocean temperature seem to be common conditions for natural
spawning. Ishnochiton magdalenensis (- Stenoplax heathiana
Berry, 1946) was observed spawning in tide pools during
early morning low tides (Heath, 1899, p. 5). Grave (1932)
and Christiansen (1954) obtained spawning in Chaetopleura
apiculata and Lepidopleurus ascellus (Spengler),
respectively, by allowing them to sit in non-circulating sea
water at slightly elevated temperatures for several hours.
While similar conditions were present when Mopalia lignosa
spawned in the lab, these conditions were not sufficient for
consistent release of gametes. Male and female Mopalia
muscosa spawned together on one occasion in a tank with
circulating seawater at a temperature of approximately 13°c.
Isolated females spawned in finger bowls in which the water
had been allowed to stand for a day. Thus, no coherent
pattern of spawning conditions could he established.
Development in Mopalia
Watanabe and Cox - 7
Thorpe (1962) reports that Mopalia muscosa and Mopalia
lignosa may spawn in the lab at times corresponding to
certain phases of the local tidal cycle. We could not
resolve any such correlation.
Detailed observations of spawning behavior were made
only for Mopalia lignosa. In two instances, once in the
late afternoon and once at night, 7 to 10 chitons placed
in a large plastic dishepan partially filled with non-
circulating seawater spawned after several hours. The pan
was tilted so that the water reached only part way up the
inclined bottom. During spawning, the girdle was elevated
anteriorly and postero-laterally. In both sexes the posterior
tip of the girdle margin was raised to form a spout through
which a single stream of gametes was released. The four
males observed spawning were more active than females during
the process. They released intermittent spurts of sperm
in long, thin streams which pooled in masses at the bottom
of the tank and diffused into the water only slowly.
The duration of each continuous sperm release was 3-5
minutes, separated by intervals of 5-15 minutes. Two of
the three females observed spawning were partially out of
water, with only their posterior ends submerged; all remained
stationary. Eggs flowed slowly out in single file,
loosely held together by a thin mucus sheath, and piled
up behind the animal.
Development in Mopalia
Watanabe and Cox - 8
Small disturbances, including a brief removal from
water, interrupted but did not permanently stop spawning.
Thorpe (1962) reported that in his experience only the
close proximity of a strong, hot light would halt spawning:
agitation and inversion of the animals would not.
DEVELOPMENT OF MOPALIA LIGNOSA
A time schedule of development for Mopalia lignosa
is given in Table I. The various stages of development
are shown in Figure 1.
Early Development
The eggs were light green and about 0.2mm in diameter.
They were very yolky and were surrounded by a transparent,
frilly chorion (Figure 1 A). Once fertilization occurred, a
small space appeared between the egg and the chorion (Figure
1 B). Cleavage was typically spiral (Figure 1 C-F).
Gastrulation begins with an invagination of the macro-
meres (Heath, 1899, p. 50). A slight invagination was
observed in several embryos 10-12 hours after fertilization
but was never positively identified as the blastopore.
Beating of the prototrochal cilia began about 12 hours
after fertilization. Just before hatching (19 hours), the
strongly beating cilia stopped for several minutes. When
beating resumed, small portions of the chorion broke away
and the trochophore struggled out anterior end first using
both muscular contractions of the body and ciliary beat (Fig¬
ure 1 H). The hatching process occupied approximately 5
Development in Mopalia
Watanabe and Cox - 9
minutes.
Free Swimming Trochophore
The newly hatched trochophores were broadly egg-shaped
with the protrochal band encircling the body at its greatest
girth. Several long cilia at the anterior end of the body, which
make up the apical tuft, were present at or before hatching
(Figure 1 I). The apical tuft was surrounded by a field of
much shorter cilia. A similar teleotrochal field of short
cilia occurred around the posterior end of the body (Figure
1 I). By the third day after fertilization these fields
had expanded to cover the body of the trochophore. About
this same time the larval eyes appeared as simple pigment
spots on the lateral margins of the body just posterior to
the prototroch. They persisted until well after metamorphosis.
The prototrochal cilia beat such that the body was
continually rotating around its axis in a clockwise direction
when viewed from the anterior end. Sometimes the larvae halt-
ed their rapid swimming and hovered in a head-up position,
maintaining their places in the water column with the slow
beat of the prototrochal cilia. The apical tuft appeared
in part to serve a sensory function, for when å swimming
larva contacted an obstacle with it, it backed away and swam
in another direction.
Mantle and Shell Development
Large epidermal cells, noted by Heath (1899) in
Stenoplax heathiana and Okuda (1947) in Cryptochiton stelleri
Watanabe and Cox - 10
Development in Mopalia
(Middendorff, 1846), appeared scattered over the dorsal body
surface 2 to 3 days after fertilization (Figure 1 K-L).
Shortly after this, a series of alternating ridges and
grooves, transversely oriented, began forming on the dorsal
surface. Five days after fertilization, eight ridges and
seven grooves were present. The large cells became aligned
along the tops of the ridges (Figure 1 M-N). This system
of ridges and grooves marked the site of the developing mantle.
By 4.5 days, the mantle field had become well-delineated by
the mantle fold (Figure 1 M). Short spicules appeared first
at the anterior margin of the mantle and by six days after
fertilization they completely encircled it (Figure 1 Q-R).
Herein, the term "settled" refers to larvae which have
lost the prototrochal cilia and become permanent crawlers.
"Metamorphosed" refers to larvae which have lost the apical
tuft and have shell plates forming.
As the larvae began to settle (5.5 days), the head
became partially covered by the mantle field. The shell
plates first appeared after about 6.5 days. Plates 2-7
(counting the cephalic or head plate as number one), the
first to form, appeared as thin, opaque slivers in the grooves
between successive dorsal ridges(Figure 1 0). The cephalic
plate formed a day later, by which time the apical tuft had
disappeared. The seven plates enlargedanteriorly and post-
eriorly until they overlapped one another (Figure 1 S-U).
Development in Mopalia
Watanabe and Cox - 11
and expanded laterally to completely obscure the body (Figure
1 V-W). A small pallial groove was formed (Figure 1 W), but
no ctenidia were present. These began appearing approximately
8 weeks after fertilization, the posterior-most ones beling
formed first. By this time the larval eyes had disappeared.
The eight (caudal) plate formed approximately 6 weeks after
fertilization.
Foot Development
The foot began to form on the ventral surface of the
body, just posterior to the prototroch and the mouth (Figure
1 M), about 4.5 days after fertilization. Within a day it
was well-differentiated, and a heavily ciliated flap appeared
at its anterior margin (Figure 1 0-P). Whether or not this
structure aided in early feeding was not determined.
Approximately five days after fertilization,the larvae
began to spend increasing amountsof time crawling on the
bottom rather than swimming with their still-present proto-
trochs. This was accompanied by a dorso-ventral flattening
of the body. The crawling larvae often attached with the
posterior end of the foot and waved their free anterior end
from side to side. They used the anterior portion of the
foot to gain firmer attachment when needed. Once the proto-
troch cilia disappeared, the larvae became permanent crawlers
and apparently began feeding, crawling slowly forward while
moving their heads from side to side. Dark material became
visable in the central region of the body.
Development in Mopalia
Watanabe and Cox - 12
Head Development
At hatching, the head region was clearly marked off
by the prototroch. In the free-swimming trochophore, it was
relatively undifferentiated, its most prominent feature being
the apical tuft (Figure 1 I). As the foot and mantle formed,
it became more flattened (Figure 1 R). Our studies did not
determine whether the mantle grew forward over the head, or
if the head itself formed new mantle tissue. At about 6.5
days, a groove appeared along the mid-ventral line of the head.
being wider at the prototroch and tapering anteriorly (Figure
1 P). Its function was not determined. By 18 days after
fertilization,the head had attained essentially its adult
form (Figure 1 W). Feeding motions of the radula were ob-
served under a compound microscope at this time.
By 8 weeks after fertilization, the juvenile chitons
had all the features of an adult: eight shell plates were
present, the girdle margin had grown out from under the plates
and was covered with spicules, ctenidia had begun to form,
and the larval eyes had disappeared.
DEVELOPMENT OF MOPALIA MUSCOSA
A schedule of development and illustrations of the
various stages appear in Table I and Figure 2. Further
details on the ontogeny of form and behavior are given below.
Early Development
The eggs of Mopalia muscosa are either green or golden
brown, 0.29mm in diameter, and are encased in a bristly
Development in Mopalia
Watanabe and Cox - 13
chorion. The vitelline membrane of an unfertilized egg lies
in close proximity to the chorion (Figure 2 A). A few minutes
after fertilization, a gap was visible between the egg and
chorion (Figure 2 B). Cleavage followed the typical spiral
pattern (Figure 2 C-F). Division after the fourth cleavage
and gastrulation were not followed in detail. The first
prototroch cilia were seen at 10 hours and the ciliated ring
appeared complete 12 hours after fertilization (Figure 2 G).
The cilia beat slowly and feebly at first, but their movements
increased in frequency and amplitude until hatching at 20
hours (Figure 2 H), when their action was sufficiently power-
ful to break away pieces of the chorion as the trochophore
emerged. The trochophore larvae usually hatched with the
apical tuft foremost, but some emerged with part of the
prototroch first.
Free Swimming Trochophore
The trochophores were roughly egg-shaped with the
prototroch a little above the equator (Figure 2 I). The
apical tuft, developed prior to hatching, was surrounded
by a field of smaller somatic cilia. A similar field of
teleotrochal somatic cilia occupied the posterior end (Figure
2 J). These two patches of somatic cilia expanded anteriorly
and posteriorly until they surrounded the body at 5.5 days.
The larval eyes, two red pigmented spots embedded in the
epithelium covering the body, appeared laterally just
Development in Mopalia
Watanabe and Cox - 14
posterior to the prototroch at 3.5 days, and were still
visible at 32 days (Figure 2 J-Y). In swimming,the larvae
rotated continuously in a clockwise direction (as viewed
anteriorly); their bodies oriented vertically with the apical
tuft up when hovering. Swimming of a larva was interrupted
only when it ran into another larva or the side of the bowl,
which caused an asynchronous beat. Ciliary action then stopped
for less than a second and resumed slowly. Larvae usually
swam along the bottom with the apical tuft in front, randomly
moving both horizontally and vertically, but only occasionally
swimming to the surface.
Mantle and Shell Development
The post-trochal region appeared elongated at 3.5
days (Figure 2 J). About 5 days after fertilization,it
began to flatten dorso-ventrally, and large cells appeared,
forming irregular rows on the dorsal surface (Figure 2 K).
This was also reported by Heath (1899) in Stenoplax heathiana
and by Okuda (1947) in Cryptochiton stelleri. A row of
short, pointed spicules then formed anterior to the proto-
troch, and extended almost entirely across the dorsal surface
of the head. A sparse scattering of spicules later appeared
laterally on the periphery of the mantle field, and by 6.5
days, spicules formed a complete ring around the circumference
of the mantle field. At 5.5 days,transverse grooves began
to form dorsally, aligned between the bands of large cells
Development in Mopalia
Watanabe and Cox - 15
(Figure 2 M), and the latter formed more regular rows,except
in the posterior region. The mantle field continued to
expand, and by 11.5 days had extended over the posterodorsal
head region. Mantle growth on the head was accompanied by
the loss of the prototroch (Figure 2 P).
Shell plate formation began around 135 days with the de-
position of seven thin lines of opaque material along the
transverse grooves of the mantle field (Figure 20). During
their early development, the order of size from the largest
to the smallest (counting the cephalic plate as number one)
was 2, 3, (4,1), 5, 6, 7 (Figure 2 0). This order suggests
the sequence of development. Enlargement of the plates con¬
tinued until they came into mutual contact and extended
laterally to the ring of spicules around the mantle field
(Figure 2 R-Y). The mantle field itself extended until the
rest of the larva was completely hidden. The eighth (caudal)
plate did not form until approximately 6 weeks after fert-
ilization. Ctenidia began to form in the pallial groove about
two weeks after this, the posterior-most ones being the first
to develop.
Foot Development
The foot began to form during the free swimming stage
(6.5 days). It gradually extended as a ventral bulge, and
a conspicuous ciliated flap formed on the anterior edge of
the foot on the seventh day (Figure 2 N). By the tenth day,
Development in Mopalia
Watanabe and Cox - 16
the foot was well formed.
Development of the foot was accompanied by a gradual
change in locomotion. The 3-day larvae swam rapidly, pre¬
dominantly near the surface. However, at 6 days,they began
to spend more time on the bottom, occasionally adhering to
it with their posterior ends. Here they swung freely from
the point of attachment or rotated due to prototrochal action
for 1 to 2 seconds; then either they were knocked loose by
movement of water in the bowl or freed themselves by swim-
ming action of the prototroch. Once freed,they resumed swimming.
As the foot developed further, the trochophores spent increas-
ing periods of time on the bottom. At 9 or 10 days,they
attached to the bottom by both the anterior and posterior
ends of the foot and rocked from side to side from these
two points. Their prototrochs were noticeably smaller and
had disappeared by 12.5 days. The larvae were then considered
being
"settled".
crawling
their only means of locomotion.
Head Development
External differentiation of the head was minimal until
which was
the disappearance of the apical tuft at 11.5 days,,about the
same time as first deposCition of the shell plates. As the
mantle field extended, the head became flattened and was soon
covered by the mantle dorsally (Figure 2 V). Radular move-
ments were observed in the mouth at 21 days.
Development in Mopalia
Watanabe and Cox - 17
SETTLING STIMULI
Metamorphosis in both Mopalia lignosa and Mopalia muscosa
was delayed or prevented by the lack of an appropriate
settling substratum. Only two larvae out of several hundred
observed underwent complete metamorphosis on glass.
In Mopalia lignosa, fully metamorphosed juveniles (shell
plates present, no apical tuft, no prototroch cilia) appeared
by seven days after fertilization when water-worn Mytilus
shells with a film of microscopic algae were added 4.5 days
after fertilization. In a bowl left without shells for 14
days, the apical tuft was still present and no shell plates
had appeared,though most of the larvae had lost their proto-
troch cilia and had been crawling 6 days after fertilization.
When Mytilus shells were added on day 14, fully metamorphosed
juveniles were noted within a day.
Similarly, no substratum was added to three bowls contain-
ing seawater and Mopalia muscosa larvae, while pieces of
Mytilus shell with an algal film were added to another three
such bowls at 5 days after fertilization. In the bowls with-
out the shells, only two larvae ever formed dorsal plates.
The rest apparently stopped development, retained the proto-
troch cilia and never metamorphosed; all subsequently died
14 to 17 days after fertilization. Development in the bowls
with Mytilus shell
progressed normally.
The advantage of a substrate-sensitive settling response
is clear. The ability to delay metamorphosis until a suit-
Development in Mopalia
Watanabe and Cox - 18
able substratum is present increases the chances that larvae
will settle under conditions favorable for post-larval
development. Such a settling response has been shown to
exist in Tonicella lineata (Wood, 1815) by Barnes (1972) and
was briefly mentioned by Heath (1899, pp. 62-63) for
Stenoplax heathiana. It has also been found in other molluscs
(Bayne, 1964; Thompson, 1964; Swennen, 1961; and Scheltema,
1961), as well as in other invertebrate groups.
DISCUSSION
Mopalia muscosa and Mopalia lignosa, while similar in
many aspects of development, show some differences, particular¬
ly in the timing of events. They are compared below to each
other, to Mopalia ciliata for which some details are available
(Thorpe, 1962), and to a few more distantly related species.
Development from cleavage through hatching takes place
at similar rates in Mopalia lignosa and M.muscosa, and at
13°C.,the larvae hatch roughly 20 hours after fertilization.
Development of M. ciliata at "normal ocean temperature" is
a little slower, and the larvae hatch 36-42 hours after fert-
ilization (Thorpe, 1962). Structures arising during the free-
swimming stage appear within 2 days of each other in M. lignosa
and M. muscosa; details are not available for M. ciliata.
Settling can be contrasted in all three species: M. lignosa
loses its prototroch cilia at about 5.5 days, while M. ciliata
and M. muscosa lose theirs on the 8th and ilth days, re-
Development in Mopalia
Watanabe and Cox - 19
spectively. For the remainder of development, M. lignosa
adheres to a schedule 4 days ahead of that for M. muscosa.
A major difference in the development of these three
species of Mopalia is in the ontogeny of the plates. Thorpe
(1962) states that "the anterior valves are the first to be
apparent" in M. ciliata. In both M. lignosa and M. muscosa,
the cephalic plate did not form until after the appearance
of the plates posterior to it. The caudal plate in M. ciliata
develops on the 8th day (Thorpe, 1962). In contrast, in both
M. lignosa and M. muscosa the caudal plated doest appear
until the 6th week.
Mopalia lignosa and M. muscosa show sequences of devel-
opmental events similar to those of the few other chitons
studied. The major differences appear in the timing rather
than the nature or order of events. Tonicella lineata,
according to Barnes (1972), hatches at 2 days and the larvae
settle at 3 days. Christiansen (1954) reported that
Lepidopleurus asellus hatched at 20 to 21 hours and settled
at about 10 days. Heath (1899) reported a period of 7 days
between fertilizationand hatching in Ischnochiton magdalenensis
(- Stenoplax heathiana). Okuda (1947) noted that
Cryptochiton stelleri did not hatch until 70 hours or more
after fertilization. The larvae emerged with external
features which did not develop in M. lignosa and M. muscosa
until the later free-swimming stages. Settling in Cryptochiton
began 12 to 20 hours after hatching and liberation from
the jelly mass.
Development in Mopalia
Watanabe and Cox - 20
A curious feature is the placement of the larval eye.
In spiralian development, larval eyes usually occur anterior
to the prototroch. The larval eyes of chitons are located
more posteriorly, behind the head region proper. They
apparently still serve a sensory function, since they are
innervated by the pallial nerve cord (Heath, 1904). Heath
(1904) puts forth the following interesting hypothesis:
"Now it is obvious that if the chiton eye
were situated in front of the velum, as in
the annelids, it would be most unfavorably
placed after metamorphosis. Under the cir-
cumstances the most available situation would
be the furrow about the proboscis, where it
would be continually obscured and would be
practically useless even if provided with
tentacles. It seems most reasonable to suppose
that as the structures characteristic of
the chitons appeared in the phylogenetic
development, the eye-spots gradually shifted
their position into the present more favorable
location.
SUMMARY
1. Spawning behavior and external features of the larval
development were studied in the chitons' Mopalia muscosa
and M. lignosa during the months of April-June, 1974, at
Pacific Grove, California.
Development in Mopalia
Watanabe and Cox - 21
2. Specimens of M. lignosa spawned in the lab after several
hours in containers of standing seawater. Males moved
around during spawning while females remained stationary.
3. Electrical stimulation, injection of O.5M KCl or homo-
genate of nervous tissue failed to induce spawning.- Eggs
obtained through dissection were unfertilizable, even after
treatment with 3ml of O.1N NaoH in 100ml seawater.
4. The sequence of events in the development of the two
species is the same, though some differences in timing
exist. First cleavage, second cleavage, third cleavage,
and hatching occurrdat about 1 hr., 1.5 hrs., 2.5 hrs., and
20 hrS.,respectively, in both species.
5. After hatching,the larvae of both species swam freely
for a period. M. lignosa settled about 5.5 days after
fertilization and M. muscosa about 11.5 days after fert-
ilization. During the free-swimming period,the larval
eyes, mantle and foot developed.
6. Larvae were considered settled when the prototroch cilia
were no longer present, and metamorphosed when the shell
plates appeared and the apical tuft was lost. This happened
at 6.5 days for M. lignosa and 13.5 days for M. muscosa.
The shell plates of M. muscosa appeareto develop in the
following order: 2, 3, 1a4, 5, 6, 7, 8. No such sequence
was noted for M. lignosa. The caudal plate in both species
did not form until about 6 weeks after fettilization.
7. Both species seemedto exhibit a substrate-sensitive
Development in Mopalia
Watanabe and Cox - 22
settling response, though further work is needed to verify this.
AWKNOWLEDGMENTS
We would like to express our thanks to Dr. Donald P.
Abbott for his guidance and willingness to assist us in this
project. We would also like to thank the rest of the faculty
and staff of Hopkins Marine Station for their assistance, with
special thanks to Christopher Harrold, Frances Fulton, and
Kristen Westersund.
Development in Mopalia
Watanabe and Cox -23
LITERATURE CITED
Barnes, James Ray
1972. Ecology and reproductive biology of Tonicella lineata (Wood,
1815). Ph.D. Dissertation. Dept. of Zoology, Oregon State University.
161 pp.; 47 figs.
(June 1972)
Bayne, B.L.
1964. Primary and secondary settlement in Mytilus edulisL. (Mollusca).
Journ. Anim. Ecol. 33:513-523;
8 figs.
Boolootian, Richard A.
1964. On growth, feeding and reproduction in the chiton
Mopalia muscosa of Santa Monica Bay. Helgoländer wiss.
Meeresunters. 11:186-199;
3 figs. (Decem¬
ber 1964)
Burghardt, Glenn E. and Laura E. Burghardt
1969. A collector's guide to West Coast chitons.
San Francisco, Cal¬
45 pp.; 4 plts.
if (San Francisco Aquarium Society, Inc.)
Christiansen, Marit Ellen
1954. The life history of Lepidopleurus asellus (Spengler)
Zool. 2:52-72;
(Placophora). Nytt Mag.
26 figs. (August 1954)
Davis, WilliamJ.,GeorgJMpitsos, and J.M. Pinneo
1973. The behavioral hierarchy of the mollusk Pleuro-
branchaea. I. The dominant position of the feeding
behavior. Journ. comp. Physiol. 90:207-224.
Development in Mopalia
Watanabe and Cox - 24
Galigher, Albert Edward and Eugene N. Kozloff
1964. Essentials of practical microtechnique. 484 pp.; illus.
Philadelphia, Penna. (Lea and Febiger)
Gould, Meredith C.
1967. Echiuroid worms: Urechis. Pages 163-171 in Fred H. Wilt and
Norman K. Wessels, eds. Methods in developmental biology. New
York, N.Y. (Thomas Y. Crowell Co.)
Grave, B.H.
1932. Embryology and life history of Chaetopleura apic-
ulata. Journ. Morphol. 54(1):153-160;
7 figs. (December 1932)
Harvey, Ethel Browne
1939. A method of determining the sex of Arbacia, and a new method
of producing twins, triplets and quadruplets. Biol. Bull. 77: 312
(29 August 1939)
Harvey, Ethel Browne
1956. The American Arbacia and other sea urchins. xiv t 298 pp.:
illus. Princeton, N.J. (Princeton Univ. Press
Heath, Harold
1899. The development of Ishnochiton. Zool. Jahrb. Abt. Anat.
12:1-90; 5 plts,: 5
figs. (9 May 1898)
Proc.
1904. The larval eye of chitons.Acad. NaturSci.
Philad.
1904:257-259; 1 fig.
1905.Thebreeding habits of chitons of the California coast.
Zool. Anz. 29:391-393
(September 1905
Iwata, K.S.
1950. A method of determining the sex of sea urchins and
Development in Mopalia
Watanabe and Cox - 25
of obtaining eggs by electrical stimulation. Annot.
zool. jap. 23:39-42
Okuda, Shiro
1947. Notes on the post-larval development of the giant
(Middendorff)
chiton, Cryptochiton stelleri. Journ. Fac. Sci. Hokkaido
Univ., Sapporo, (6)Zool.9267-275: 18 figs.
Scheltema, Rudolf S.
1961. Metamorphosis of the veliger larvae of Nassarius
obsoletus (Gastropoda) in response to bottom sediment.
Biol. Bull. 120292-109;
i fig. (February 1961)
Swennen, C.
1961. Data on distribution, reproduction, and ecology of
the nudibranchiate molluscs of the Netherlands.
Journ. Sea Res. 1:191-240
(April 1961)
Thompson, T.E.
1964. Grazing and the life cycles of British nudibranchs. Pages
275-297 in D.J. Crisp, ed. Grazing in terrestrial and marine en-
vironments. Brit. Ecol. Soc. Symp. No. 4, Oxford, U.K. (Blackwell
Scientific Publications)
Thorpe, Spencer R.,Jr.
1962. A preliminary report on spawning and related
phenomena in California chitons. The Veliger 41202-210;
3 figs.
Wolfsohn, Julian Mast
1907.Thecausation of maturation in the eggs of limpets
Development in Mopalia
Watanabe and Cox - 26
by chemical means. Biol. Bull. 132344-350,
2 figs.
Development in Mopalia
Watanabe and Cox - 27
TABLE CAPTIONS
Table 1. Time schedule of development for Mopalia lignosa
and Mopalia muscosa in hours and days after fertilization.
A vertical bar indicates the exact beginning and end of
an event as observed. A dotted line indicates that the
exact times were not determined. These times are re¬
presentative and do not indicate the limits of individ-
ual variation.
A. First cleavage
J. Larval eyes
B. Second cleavage
K. Mantle field
C. Third cleavage
1. Large cells appear on
D. Fourth cleavage
the dorsal surface
E. Gastrulation
2. Mantle fold develops
F. Prototroch
3. Mantle field extends
1. faint ciliary action
onto head
within the chorion
L. Foot
2. strong ciliary action
1. initial bulge
within chorion
2. ciliated flaps develops
3. swimming activity
anteriorly
4. disappearance of cilia
3. well-developed foot
G. Apical tuft
M. Spicules
H. Hatching
1. acrcss dorsal surface
I. Somatic cilia
of head only
1. only at anterior and
2. all around mantle field
posterior ends
N. Shell plates
Z. distributed over body
1. CacO deposits first seen
2. seventh plate forms
3. plates expand and over¬
lap one another
VELOPMENTIN MOPACA
WATANABE
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Development in Mopalia
Watanabe and Cox - 28
FIGURE CAPTIONS
Figure 1. Development of Mopalia lignosa.
A. Unfertilized egg within chorion. B. Fertilized egg show-
ing the perivitelline space. C. First cleavage (1 hr.).
D. Second cleavage (1.5 hrs). E. Third cleavage (2.5 hrs).
F. Fourth cleavage (3.5 hrs.). G. Prototrochal cilia begin
beating within chorion (12 hrs.). H. Hatching (19 hrs.). I. New-
ly hatched trochophore (about 1 day). J. Free-swimming troch-
ophore. Eye spots appear (3-3.5 days). K,L. Dorsal ridges
and foot forming (dorsal and lateral views, respectively; about
4 days). M. Mantle fold evident (lateral view; 4.5 days).
N-P. Settling stage (dorsal, lateral, ventral views,respectively):
eight dorsal ridges and ciliated flap of foot present (about
5 days). Q-R. Settled larva (dorsal and lateral views, re-
spectively); shell plates 2-7 begin forming (6.5 days). S-U.
Shell plates expand and apical tuft lost (dorsal, dorsal, lat-
eral views, respectively; 7-8 days). V-W. Juvenile chiton
(dorsal and ventral views, respectively; 24 days).
Figure 2. Development of Mopalia muscosa
A. Unfertilized egg within chorion. B. Fertilized egg,
showing perivitelline space. C. First cleavage (ihr.).
D. Second cleavage (1.5 hrs.. E. Third cleavage (2.5 hrs).
F. Fourth cleavage (3.5 hrs.). G. Prototroch complete
(12 hrs.). H. Hatching (20hrs). I. Early trochophore
showing anterior and posterior fields of somatic cilia (1
Development in Mopalia
Watanabe and Cox - 29
day). J. Post-trochal region elongated; eye spots (3.5
days). K. Post-trochal region flattened dorsoventrally
(5.5 days). L. Spicules on head and beginning of cell
alignment on dorsal surface of mantle field (5.5 days).
M. Grooves on dorsal surface of mantle field, foot beginning
to bulge (6 days). N. Ciliated flap on anterior foot (7
days). 0. Foot more differentiated (7 days). P. Settled
larva, loss of prototroch (11.5 days). 0. Shell plate
deposition, loss of apical tuft (13.5 days). R-S. Mantle
field begins to extend onto head (14 days). T-U. Shell
plates enlarging (15 days). V. Shell plates in mutual
contact (17 days). W. Foot well developed (17 days).
X. Juvenile chiton in curled position, 7th plate hidden
from view (21 days). Y. Ventral view, head and mouth well
formed, but neither eighth plate or first pair of ctenidia
have appeared.
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