AMER. ZOOL., 22:817-826 (1982)
Ascidian Larvae and the Events of Metamorphosis1
RICHARD A. CLONEY
Department of Zoology, University of Washington,
Seattle, Washington 98195
SYNOPSIS. Ascidian larvae settle and metamorphose after only a brief free-swimming
period; they are all lecilhotrophic. Extrinsic factors (chemical and physical) may trigger
metamorphosis but none are known to be essential. The major larval structures may be
classified as transitory larval organs (TLO), prospective juvenile organs (PJO) or larvaljuvenile organs (LJO). TLO are phagocytized or otherwise destroyed at metamorphosis;
the PJO and LJO become the functional parts of the juvenile or oozooid. Metamorphosis
involves some rapid and some slow morphogenetic movements. Variations in larval morphology are reflected in metamorphosis. Some of the events of metamorphosis are fairly
well known in terms of the role of specific cells but we know little about how the various
events are initiated and coordinated. The nervous system is more complex than had been
previously assumed. The nervous system, neuroid conduction and diffusion of one or
more humoral factors are probably all involved in controlling metamorphosis.
gans or tissues (LJO) (Table 1). Organs in
the first group (TLO) function in larval locomotion, sensory input and settlement.
They are fully differentiated in the larva
but are destroyed, or in the case of the fins,
lost during metamorphosis. Organs in the
second group (PJO) may be well or poorly
differentiated in different species. They are
in an arrested state of development in the
larva; they may become functional either
shortly after settlement or following further histogenesis. The last group (LJO) includes
organs or tissues that function in
(Molgula manhattensis) to 11.0 mm (Eudisboth
larval
and post-larval phases of the
toma digitatum). The largest tadpole in
North America is probably Ecleinascidia life cycle. The mesenchyme cells differturbinata (4.5 mm). All ascidian larvae are entiate into blood cells, smooth muscle and
dependent upon stored substances for nu- cardiac tissue but the timing is variable in
trition during their rather brief non-feed- different species. Some larvae have a funcing, free-swimming period of a few min- tional heart.
utes to several days.
The body plan is similar in all larvae but
LARVAE OF SOLITARY ASCIDIANS
there are variations in the histology of larval organs as well as major differences in
All solitary ascidian larvae that have been
the pace or tempo of development of cer- identified are in the families Cionidae, Astain organs. The principal structures com- cidiidae, Corellidae (order Phlebobranprising ascidian larvae may be categorized, chiata), and Pyuridae, Styelidae and Molfor didactic purposes, into three groups: gulidae (order Stolidobranchiata). In all
transitory larval organs (TLO), prospective ju- these families, except molgulids, the larvae
venile organs (PJO) and larval-juvenile or- have three anterior, simple, non-eversible,
adhesive papillae, arranged in a triangular
field. The papillae secrete adhesives that
' From the Symposium on The Developmental Biology are used to effect settlement at the onset
of the Ascidians presented at the Annual Meeting of
ihe American Society of Zoologists, 27-30 December of metamorphosis. Molgulid larvae have no
papillae but the epidermis of the trunk se1981. at Dallas, Texas.
INTRODUCTION
Most solitary ascidians are oviparous and
most compound ascidians are ovoviviparous, although there are some interesting
exceptions (Brewin, 1956; Berrill, 1975;
Cloney and Cavey, 1982). There are a few
species with direct (anural) development in
the families Molgulidae and Styelidae but
all tadpole larvae are easily recognizable as
having a trunk, tail and fins. Larvae of
species in ten families have been described.
They range in length from about 0.6 mm
817
818
RICHARD A. CLONEY
TABLE 1. Didactic classification of ascidian larval organs.
Transitory larval organs
Notochord
Caudal musculature
Visceral ganglion and nerves
Dorsal tubular nerve cord
Papillae
Outer cuticular layer of tunic
Sensory vesicle and sensory organs
Prospective juvenile organs or rudiments
Branchial and atrial siphons
Atrium (single or paired)
Branchial basket, esophagus,
stomach and intestine
Pyloric gland
Heart (some species)
Neurohypophysis or cerebral
ganglion and neural gland
Ampullae
Epicardium
Probuds or precocious blastozooids (compound ascidians)
Larval-juvenile organs
Epidermis
Inner cuticular layer and inner
compartment of tunic
Mesenchyme and derivatives
including blood cells
Heart (some species)
the epidermis of the dorsal and ventral
parts of the tail (Torrence and Cloney,
1982). Primary sensory cells are probably
also present in the papillae (Fig. 3).
The larval tunic is composed of inner
and outer cuticular layers, extracellular fila tube {e.g., Ascidia, Chelyosoma, Corella, Bol- aments, ground substance and, in some
tenia, Pyura, Styela and Molgula). T h e wall species, amoeboid cells from the hemocoel.
of the notochord is bounded externally by It lies outside the epidermis and most of it
a basal lamina and a filamentous sheath is produced by the epidermis. The outer
(Cloney, 1964). The tail contains about 36 cuticular layer forms the tail fins. Recent
uninucleate, striated muscle fibers, ar- investigations (Cloney and Cavey, 1982) of
ranged in three rows along each side of the Corella inflata and Ascidia paratropa have
notochord. The muscle fibers in each row demonstrated that the fins do not form
are joined at their ends by transverse myo- normally when embryos are dechorionatmuscular junctions that resemble verte- ed (removal of chorion, test cells and folbrate cardiac intercalated discs (Cavey and licle cells) before the early tail bud stage.
Cloney, 1976). Adjacent fibers on each side Larvae that develop from dechorionated
of the notochord are also joined by gap embryos are sticky and cannot swim even
junctions and are electrically coupled.
though they are otherwise active and norThe functional larval nervous system in- mal in morphology. When embryos are decludes a visceral ganglion, an associated chorionated at or following the late tail bud
fluid-filled sensory vesicle and a dorsal hol- stage they develop normal fins and are not
low nerve cord which extends into the tail. sticky. When the test is being formed the
The sensory vesicle contains an otocyst test cells are motile and appear to swarm
(statocyte) and the rear wall usually bears over the embryo, causing it to rotate. They
an ocellus (Eakin and Kuda, 1971). The also produce electron dense granules called
nerve cord is composed of a single layer of "ornaments" in most species which adhere
ependymal cells which embrace axons from to the outer cuticular layer. The possibility
the visceral ganglion in two lateral chan- that the test cells influence morphogenesis
nels. The axons form neuromuscular junc- of the larval tunic is being tested.
tions with the dorsal muscle fibers (Cloney
The PJO of all solitary ascidians are rel1978; Torrence and Cloney, 1982).
atively undifferentiated in the larval stage
Sensory nerves extend both from the pa- (Kowalevsky, 1871; Crave, 1944; Cloney,
pillae and from the tail to the visceral gan- 1961). The branchial basket, esophagus,
glion. The sensory nerves of the tail are stomach, siphons, atrium and heart are
known to be formed by primary sensory represented by yolky anlagen. Smooth
neurons whose somata are located within muscle cells are not yet formed within the
cretes adhesives for the same purpose
(Torrence and Cloney, 1981).
The notochord is a stiff rod composed
of about 40 epithelial cells that, with few
exceptions, are arranged in a single layer
around a central core of matrix, forming
819
ASCIDIAN METAMORPHOSIS
EPIDERMIS^
BRANCHIAL
BASKET
ENDOSTYLE
AXIAL COMPLEX
DORSAL
NERVE
CORD
EPIDERMIS
PYLORIC
GLAND
PAPILLARY
NERVE
STOMACH
TUNIC
Fic. 1. Diagrammatic drawing of the larva of Distaplia occidentals. Details of the atrium, tunic and hypophyseal duct are omitted. The length of the trunk is 1 mm; the entire larva, including the fin is 3.2 mm long.
(From Cloney and Torrence, 1982)
siphons or the body wall. Definitive blood
cells, derived from the mesenchyme cells,
are not yet differentiated. The cerebral
ganglion and neural gland are represented
by a rudiment called the neurohypophysis.
Secondary organs of attachment, called
ampullae, are not visible in most solitary
ascidian larvae although they become
prominent epidermal outgrowths shortly
after settlement (Patricolo et al., 1981;
Torrence and Cloney, 1981).
LARVAE OF COMPOUND ASCIDIANS
Compound ascidians are found in the
families Polyclinidae, Polycitoridae, Didemnidae (order Aplousobranchiata), Perophoridae, Cionidae (order Phlebobranchiata), and Styelidae (order Stolidobranchiata). The larvae are usually more complex than those of solitary ascidians.
There are usually three adhesive papillae arranged in a triangular field (two
dorsal and one ventral, e.g., Distaplia, Hypsistozoa, Botryllus) or in the sagittal plane
(e.g., Eudistoma, Aplidium, Diplosoma). A few
species have only two papillae (Euherdmania claviformis and several didemnids).
The notochord is usually tubular as in
solitary ascidians {e.g., Distaplia, Aplidium
and Diplosoma). At the tail bud stage, as in
solitary ascidians, the notochord is composed of a single row of about 40 discoidal
cells; these cells later become reduced in
diameter but longer and begin to secrete a
matrix that accumulates between the cells
in discrete lenticular pockets. Later the cells
become rearranged to form a cylinder
(closed at both ends) that surrounds a core
of matrix. There are a few striking exceptions to this pattern. Ecteinascidia turbinata
and Clavelina huntsmani retain the intercellular lenticular pockets of matrix and never form a tubular notochord. The notochordal cells of Metandrocarpa taylori and
820
RICHARD A. CLONEY
INNER
CUTICULAR
LAYER
SENSORY/
NERVE
BASAL
LAMINA
COMMON
EPIDERMAL
CELL
SENSORY
NEURON
AXON
HEMOCOEL
FIG. 2. Primary sensory cells and sensory nerves of the tail in Diplosoma macdonaldi. These sensory cells and
nerves are found along the dorsal and ventral parts of the tail. They are thought to be mechanoreceptors.
(From Torrence and Cloney, 1982)
some other styelids secrete little or no ma- extends along the dorsal midline and
trix and the notochord is composed of a another extends along the ventral midline.
Each nerve is composed of 50-70 naked
single row of cells.
The larva of Dislaplia occidentalis (Fig. 1) axons. These lie in epidermal grooves and
has about 1,500 uninucleate, striated mus- extend to the visceral ganglion. The cell
cle cells in the tail; Diplosoma macdonaldi has bodies of these neurons occur in pairs
about 1,600 and Aplidium constellation has within the epidermis. Each cell body has a
about 128. These are not myotendonal single sensory cilium that extends into the
junctions as in vertebrate somatic muscle. inner and outer cuticular compartments
The muscle cells are always joined by of the tunic. These neurons are probably
transverse myomuscular junctions and gap mechanoreceptors. Similar cells are found
junctions (Cavey and Cloney, 1972, 1976; in the wall and stalk of each papilla. PriSchiaffino et al, 1974; Burighel^a/., 1977). mary sensory cells with a single cilium and
The larval nervous system is similar to many microvilli are found on the papillary
that of solitary ascidians although it is usu- rims in this species and may be chemoreally more complex. Ultrastructural details ceptors.
of the ocellus and statocyst of Distaplia oc- The caudal epidermis is a simple squacidentalis have been described by Eakin and mous epithelium, usually about 0.5—2.0 jiim
Kuda (1971). Motor axons from the vis- in thickness (e.g., Distaplia, Diplosoma, Apliceral ganglion innervate the dorsal rows of dium). It makes an abrupt bend at the base
muscle cells in most species.
of the tail and is continuous with the epiPrimary sensory cells have been discov- dermis of a large chamber (Fig. 1) in the
ered in the tail (Fig. 2) and papillae (Fig. posterior part of the trunk. This chamber
3) of Diplosoma macdonaldi (Torrence and receives the axial complex of the tail (musCloney, 1982). In the tail, one sensory nerve cle, nerve cord and notochord) during tail
ASCIDIAN METAMORPHOSIS
SENSORY CELL
MCROvlLU
PAHLLAR WALL
10 fim
PRIMARY SENSORY
CELLS
CIUUM
PAPiLLAR
STALK
821
In most compound ascidians, except
styelids, the tail is rotated 90° counter
clockwise during development. The dorsal
side of the tail, defined by the nerve cord,
is on the left relative to the trunk. The fins
are horizontal in the frontal plane relative
to the trunk. In compound ascidians the
PJO are usually in an advanced state of
differentiation as exemplified by the larva
of Distaplia occidentalis (Fig. 1). As in the
adult zooids, this larva has four rows of
stigmata on each side of the branchial bas\PRIMARY SENSORY CELL
ket. The branchial basket, siphons, esophCirriCULAR LAYERS
^ANCHOR CELL)
agus, stomach, intestine, pyloric gland,
FIG. 3. Diagrammatic longitudinal section of an ad- atrium, smooth muscle of the body wall,
hesive papilla of Diplosoma macdonaldi. Sensory cellsand blood cells are well differentiated. A
are stippled. Primary sensory cells in the rim of the fully differentiated heart is located below
cup (anchor cells) have microvilli that extend between the pharynx; it pumps continuously and
the inner and outer cuticular layers of the tunic. Other sensory cells have cilia but no microvilli. This species periodically the direction of beat is rewill settle instantly when the papillae contact an ap- versed. A cerebral ganglion and ciliated
propriate substrate. (From Cloney and Torrence, hypophyseal duct are located dorsal to the
1982)
visceral ganglion. The hypophyseal duct
joins the branchial basket, forming a ciliated funnel. A clearly defined neural gland
resorption. The epidermis of the trunk has not yet been identified in Distaplia alfollows the contours of the viscera and sen- though it has been described in other larsory vesicle. Anteriorly it forms ampullae vae (Levine, 1962).
(with a variety of shapes in different
SETTLEMENT AND METAMORPHOSIS
species) and the adhesive papillae.
Larvae are covered by two cuticular layThe events of metamorphosis transform
ers of tunic formed by the epidermis with the non-feeding, mobile larva into a filter
contributions from the test cells. The outer feeding, fixed juvenile. Metamorphosis inlayer forms the tail fins and is cast off at volves numerous rapid morphogenetic
metamorphosis; the inner layer then be- movements and physiological changes that
comes the outermost surface of the oo- are initiated at the moment of settlement.
zooid shortly after settlement. In all species
The differentiation of prospective juvethe tunic contains matrix and filaments nile organs may occur mainly before setwithin the inner and outer compartments tlement (most compound ascidians) or
formed by the cuticular layers. In all com- mainly after settlement (solitary ascidians).
pound ascidian larvae there are free cells Some events of metamorphosis may be
in the tunic; many are amoeboid and some completed in seconds or minutes (e.g.,
resemble blood cells in the hemocoel. In papillary eversion and tail resorption) while
polycitorids and didemnids the inner com- others may take many hours {e.g., rotation,
partment is usually filled with conspicuous, ampullar outgrowth, phagocytosis of axial
transparent, vacuolated cells (bladder cells), complex). The major events of metamorreminiscent of vertebrate unilocular fat phosis are listed in Table 2. For the ascidcells as seen in routine paraffin sections, ians, as a whole, the list more or less debut they do not contain fat. These cells tend fines metamorphosis but there are many
to expand and streamline the contours of variations in different species. Papillary
the trunk. Symbiotic spiriliform bacteria are eversion is obviously restricted to species
abundant in the tunic of Distaplia occiden- with eversible papillae but papillary retractalis and Diplosoma macdonaldi; some di- tion occurs in all papillate species.
demnids bear external symbiotic algae.
Settlement begins with the attachment of
822
RICHARD A. CLONEY
species (Cloney, 1978). Nothing is known
about how specific inducers trigger meta1. Secretion of adhesives by the papillae or the
morphosis. It is a difficult problem to solve
epidermis of the trunk
because of variations in species and indi2. Eversion and retraction of papillae
viduals of the same species.
3. Resorption of the tail
4. Loss of the outer cuticular layer of the larval
Patricolo et al. (1981) have studied the
tunic
effects of L-thyroxine on the metamor5. Emigration of blood cells or pigmented cells
phosis of Ascidia malaca. Thyroxine speeds
(many species)
6. Rotation of organs through an arc of about 90°; up the onset of metamorphosis in this
species. These investigators suggest that a
expansion of branchial basket; elongation of
oozooid or juvenile
group of free cells of endodermal origin,
7. Expansion, elongation or reciprocation of am"button cells," located in the ventral hemopullae; reorientation of test vesicles; expansion
coel of the trunk of Ascidia, might produce
of the tunic
a thyroxine-like hormone that stimulates
8. Retraction of the sensory vesicle
metamorphosis, but there is no evidence
9. Phagocytosis of visceral ganglion, sensory organs and cells of axial complex
that these larvae or any other ascidian
10. Release of organ rudiments from an arrested
larvae actually produce thyroxine. Their
state of development
hypothesis raises the questions: How are
the button cells stimulated? What is the
role of the larval nervous system in settlea larva to a solid substratum, another ani- ment?
The responses of ascidian larvae to light
mal, a plant or even an air-water interface
in a culture vessel. In many compound as- and gravity have been reviewed by Crisp
cidians the papillae evert rapidly and ex- and Ghobashy (1971). The larvae of Diplopose adhesives; at the same time adhesives soma listerianum, studied by these investimay be secreted (Cloney, 1978). In solitary gators, are negatively geotactic and posiascidians and some compound ascidians tively phototactic as soon as they emerge
{e.g., Clavelina, Botryllus) the papillae do not from parent colonies. They alternately
evert but simply secrete adhesives. When swim and rest just before settlement. At
these events occur, the other events of this time they avoid light and prefer to setmetamorphosis follow in a sequence char- tle on dark or shaded surfaces. These laracteristic of the species even if the larvae vae, like most other ascidian larvae, are
are not attached. Metamorphosis may oc- stimulated to swim by fluctuating levels of
cur on slides or in culture vessels without light, especially when the illumination is
actual settlement. The release of adhesives interrupted by shadows. Fluctuating intenor papillary eversion, whether or not it re- sities of light also significantly shorten the
sults in attachment, is the first event in mean free-swimming period of D. listerianum. In continuous darkness or continuous
metamorphosis.
Most larvae settle spontaneously over a light the mean swimming period is longer.
Larvae of Diplosoma macdonaldi behave
period of minutes or hours in laboratory
culture vessels, but the percentage may be similarly. They also display a remarkable
low with some species. Metamorphosis of reaction to a specific type of polyethylene.
various species can be triggered with low They usually settle immediately the first
concentrations of vital dyes, some amino time their adhesive papillae contact the
acids, copper and other metallic ions, io- plastic directly. No other substrate has been
dine, thyroxine, extracts of larval and adult found that elicits this response (Cloney,
tissues, exposure to hypotonic sea water 1978). Primary sensory cells in the papillae
and sea water in which larvae have previ- of this species (Cloney and Torrence, 1982)
ously metamorphosed (see review by are assumed to be chemoreceptors that are
Lynch, 1961). Dimethylsulfoxide, acetyl- stimulated by close contact with the subcholine, mechanical stimulation, favorable stratum (Fig. 3). A reflex arc involving the
lighting conditions, and rarely, a specific visceral ganglion and motor fibers may
substrate are also effective with some trigger papillary eversion.
TABLE 2. Events of metamorphosis in ascidians.
ASCIDIAN METAMORPHOSIS
823
Young and Braithwaite (1980) have
demonstrated that the larvae of Chelyosoma
production have a strong preference for settlement on conspecific adults and upon the
substratum adjacent to conspecific juveniles.
Several species (Distaplia occidentals, Diplosoma macdonaldi, Eudistoma ritteri) can be
stimulated to metamorphose by pinching
the tail or parts of the trunk. This suggests
that extrinsic factors (specific ions or molecules) are not essential for metamorphosis in these species even thought they may
trigger it.
Papillary eversion in compound ascidians
Among the compound ascidians there
are at least four types of non-eversible papillae and five types of eversible papillae
(Cloney, 1978). There are striking differences in papillary structure as well as in the
mechanisms of eversion. The cup-shaped
FIG. 4. Contractile properties of the caudal epidermis. When the tail is removed with forceps from the
trunk (along the dotted line) immediately after the
beginning of tail resorption, the epidermis contracts
and forms a compact mass at the tip of the axial complex. The isolated epidermis is also contractile. The
axial complex loses turgor, but does not shorten, fold
or coil. The epidermis will not contract if the tail is
excised before the onset of metamorphosis. (The outer cuticular layer is usually removed before the tail is
pulled from the trunk.) (From Cloney, 1978)
papillae of Distaplia occidentalis have been
studied most intensively (Cloney, 1977, cavity containing the axial complex is then
1979). At the onset of metamorphosis each reduced in volume by contractions of the
papilla everts and transforms into a hy- trunk epidermis, and the caudal epidermis
perboloidal shape within about 30 sec. invaginates. The axial complex is slowly
The transformation is caused by the con- broken apart and engulfed by phagocytes
traction of about 260 myoepithelial cells in over a period of about 3 days.
the wall of the cup. Immediately following
The caudal epidermis can contract ineversion the papillae begin to retract. dependently (Fig. 4) if it is removed from
Eversion of the papillae is not inhibited by the tail immediately after the onset of tail
cytochalasin B but the process of retraction resorption, but it does not manifest conis reversibly inhibited. The scyphate pa- tractile properties if it is removed from the
pillae of Diplosoma macdonaldi (Fig. 3) evert larva (Cloney, 1966, 1972, 1978). In the
by a similar mechanism.
larva the caudal epidermal cells are squamous; during contraction they become
Mechanisms of metamorphosis in
compressed apically and extended basally.
different taxa
They rapidly transform into flask-shaped
Tail resorption in Distaplia occidentalis cells. In early stages of contraction arrays
begins about two minutes after papillary of microfilaments (actin) become aligned
eversion. The epidermis separates from the parallel to the axis of contraction in the
underlying axial complex (notochord, apical cytoplasm of the caudal epidermal
muscle, and nerve cord) and begins to con- cells. As the epidermal cells contract this
tract. The notochordal cells pull apart and layer of microfilaments increases in thickround up, causing the notochord to be- ness. At the same time the surfaces of the
come limp owing to leakage of the matrix. cells become highly folded (Cloney, 1966,
The axial complex moves steadily into the 1972). The caudal epidermis clearly proposterior compartment of the trunk and vides the driving force in tail resorption.
forms into a right hand helix. Within about Cytochalasin B arrests contraction in about
7 min, at 20°C, the caudal epidermis is re- 90 seconds and disorganizes the arrays of
duced to a cone-shaped mass at the pos- microfilaments. If the cytochalasin B is
terior end of the trunk. The posterior body washed away, the microfilaments become
824
RICHARD A. CLONEY
.contractile epidermis
axial complex
Ecteinascidia
Form 2
Boltenia
Form 4
:ontractile notochordal
cells
FIG. 5. Diagrammatic representation of five forms
of tail resorption. The solid black parts represent
contractile tissue. The stippled parts represent the
axial complex (Forms 1-3) or muscle cells (Forms 4
and 5). Microfilaments become aligned in the apices
of the caudal epidermal cells in Forms 1 and 2. In
Form 3 they are in the basal cytoplasm. In Forms 4
and 5 the microfilaments become aligned in the basal
cytoplasm of the notochordal cells. (From Cloney,
1978)
Fig. 5) begins with an abrupt rupture of
the anterior end of the notochord. The notochordal cells and matrix flow out of the
notochordal sheath into the posterior trunk
(Cloney, 1961, 1969; Numakunai et al.,
1964). The epidermis thickens during tail
resorption but remains in close contact with
the muscle cells. No subepidermal space is
formed. During tail resorption the proximal notochordal cells develop conspicuous
folds at their basal surfaces bordering the
notochordal sheath. The folds are always
associated with prominent longitudinal arrays of microfilaments. The changes observed in the proximal cells gradually extend to the more distal cells (Cloney, 1969).
The ultrastructural changes in the notochordal cells in many ways resemble those
of the epidermal cells of Forms 1, 2, and
3.
The mechanism of rupturing the notochordal sheath has not been determined,
and the inference that the notochordal cells
are contractile is based solely on ultrastructural work and cinemicrography. I have
been unable to isolate the notochord and
show that it has independent contractile
properties. The fact that the epidermal cells
do not form aligned arrays of microfilaments during tail resorption in this group
is consistent with the hypothesis that the
driving force in tail resorption is generated
by the notochordal cells.
Resorption of the tail in Molgula occiden-
realigned and contraction resumes (Cloney, 1972; Lash et al, 1973).
The pattern of tail resorption in all of
the aplousobranchs that have been exam- talis and M. manhattensis (Form 5, Fig. 5)
ined is similar to that of Distaplia. This pat- also seems to involve contraction of the notern is called Form 1 (Fig. 5). The process tochordal cells, but the notochordal cells
of tail resorption in phlebobranchs is al- do not flow out of the sheath as the tail
ways slower than in aplousobranchs but the shortens. Instead the whole notochord
caudal epidermal cells form apical, aligned transforms from a long tubular structure
microfilaments during tail resorption and into a short ovoid mass. In the process the
the overall mechanism is very similar (Form matrix escapes but the cells remain joined
2, Fig. 5). In Botryllus schlosseri the caudal to each other. As in Form 4, microfilepidermis is also contractile but the micro- aments become aligned in the basal cytofilaments become aligned in the basal cyto- plasm of the notochordal cells and the basplasm of the epidermal cells instead of in al surfaces of the cells become folded.
the apical cytoplasm (Lash et al, 1973;
In all forms of tail resorption the axial
Schiaffino et al., 1974). When the tail complex is eventually destroyed by phagoshortens the axial complex does not coil or cytes; none of the cells from the axial
fold; the cells become dissociated (Form 3, complex have been shown to have any prospective developmental significance. PostFig- 5).
Tail resorption in pyurids and solitary larval development proceeds normally even
ascidians of the family Styelidae (Form 4, if the tail is removed from a larva, but the
ASCIDIAN METAMORPHOSIS
825
entirely epidermal structures with glandular cells that produce adhesives. In compound ascidians they are often conspicuLoss of fins
ous processes behind the papillae. They
The outer cuticular layer of the tunic that may be digitate, capitate, bulbous, clavate
forms the fins is usually cast off shortly af- or ring-shaped in form. Test vesicles in
ter the completion of tail resorption. In Aplidium are derived from ampulla-like
species with very sticky surfaces the fins processes. Ampullae are usually not visible
may remain attached to the juvenile for in solitary ascidian larvae but grow out from
many days.
the trunk epidermis shortly after settlement and are later retracted. Within a few
Emigration of blood cells
minutes after settlement the bulbous amShortly after the onset of metamorpho- pullae of Distaplia occidentalis begin to exsis, blood cells of the hemocoel of Aplidium pand and form four distended processes
constellatum, in a brief burst of activity, em- that help to affix and support the oozooid.
igrate across the epidermis into the tunic The larvae of Diplosoma macdonaldi have
(Cloney and Grimm, 1970). These cells be- digitate ampullae that extend radially from
come incorporated into vacuoles on the the attached oozooid and spread the area
basal surface of epidermal cells and are lat- of contact with the substratum. The activer exteriorized at the apical surface of the ity of these ampullae is striking because
epidermal cells. The pathway has there- they alternately extend and retract before been called transcellular.
neath the tunic.
In Distaplia occidentalis, cells that are al- The ampullae of Molgula occidentalis have
ready within the tunic become highly mo- been examined in considerable detail
bile shortly after tail resorption. In Diplo- (Torrence and Cloney, 1981). They grow
soma macdonaldi cells containing white out following settlement and form a firm
pigment granules are present in the tunic attachment to the substratum. Repetitive,
close to the epidermis. They are not motile peristaltic contractions pass from the base
in the larva, but during tail resorption they to the distal end of each ampulla. These
suddently become activated and move out- contractions are mediated by a layer of miward to the surface of the tunic. The crofilaments in the basal cytoplasm of the
change in position of these cells alters the ampullar cells. There are no nerve fibers
organism's appearance from a dark to a in the ampullae; ampullae continue to conlighter color. Ciona intestinalis, Ascidia cal- tract with an intrinsic rhythm even when
losa and many other species undergo isolated. Coordination between cells of the
changes in appearance that may be attrib- epithelium is believed to be facilitated by
uted to light scattering properties of cells neuroid conduction via gap junctions.
that enter the tunic during metamorphosis. Activation of the blood cells must be
caused by a diffusible substance, perhaps Destruction of transitional
from the central nervous system.
larval organs
potential nutrients represented by the axial complex are lost.
Rotation of the trunk
During metamorphosis there is usually
a rotation of the siphons and viscera
through an arc of about 90°. The timing is
quite variable in different species; Distaplia
occidentalis requires about 2 days. Very little is known about the mechanisms involved.
Ampullae
The ampullae of juvenile ascidians are
secondary organs of attachment. They are
Within a few minutes following onset of
metamorphosis the sensory vesicle is withdrawn. In Diplosoma macdonaldi this is a
rapid event that probably involves contraction of the epidermal cells over the vesicle.
The axial complex, larval sensory organs
and visceral ganglion are destroyed by
phagocytes over a period of several days
in many ascidians. The problems of what
stimulates phagocytic activity and how the
phagocytes recognize the TLO are unresolved.
826
RICHARD A. CLONEY
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metamorphosis in bryozoan and ascidian larvae.
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cidian metamorphosis. Z. Zellforsch. 100:31—53.
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Involution of the caudal musculature during
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REFERENCES