Number 50 December 2001
In May we moved to the Friday Harbor Labs where Charley worked on germinal
vesicle breakdown in ascidian oocytes. Shortly after getting settled
in, Gretchen rushed off to California for the birth of our first grandchild
(a boy, Alex), and she also identified several hundred ascidians at the
California Academy of Sciences in San Francisco and taught an ascidian
identification workshop there. Back at the labs, Gretchen continued
with taxonomy projects including a description of a new species of Trididemnum
from the Friday Harbor region which she will name after her grandson; she
hopes it will be an inspiration to him to become a biologist like his parents
and grandparents. Charley taught the summer comparative embryology
course at FHL with Mark Martindale from Hawaii. We enjoyed talking to fellow
ascidiologists Billie Swalla and Richard Whittaker who also spent the summer
at the labs. In August we spent a week in Kodiak, Alaska with the
Smithsonian Environmental Research group (SERC) looking for invasive species.
Fourteen species of ascidians were collected but none were considered nonindigenous.
Among the ascidians we collected were specimens of Aplidium coei described
by Ritter on the Harriman Alaska Expedition of 1899. It was a real
thrill to find Ritter’s species just about where he said it was after so
many years. We were also impressed by the huge numbers and size of
the Molgula retortiformis we found on floats. We just returned from the
SERC facility in Edgwater, Maryland where we spent a week giving workshops
on how to identify the hundreds of ascidian samples from their various
sampling sites around the U.S.
Gretchen spends many hours assembling and editing AN twice a year;
she was wondering if it were worth while to continue with it after so many
years. Accordingly, she sent out an inquiry as to the usefulness
of continuing with this effort. Numerous replies were unanimous that
AN continues to be very useful and interesting to many researchers; thus
the newsletter will continue for a few more years at least! We are
grateful for past articles but need your continued input to insure that
AN remains useful and informative. Please send us a meeting abstract,
thesis abstract, work in progress, or copy of your latest publications
for the next issue. There are 118 new publications listed
in this issue, plus numerous additional ones in the Table of Contents for
The Biology of Ascidians included near the end of this newsletter just
before the New Publications.
*Ascidian News is not part of the scientific literature and should not be cited as such.
NEWS AND VIEWS
1. The word ascidian is derived from the Greek askidion, a diminutive of askos, meaning wineskin or bladder. It was Aristotle who said, "For man once a leathern bottle was."
2. About 40 copies of "The Biology of Ascidians" (eds by H. Sawada, H. Yokosawa, and C.C.Lambert), Springer-Verlag Tokyo 2001, are still available in the secretariat of the First International Symposium on the Biology of Ascidians (c/o Department of Biochemistry, Graduate School of Pharmaceutical Sciences, Hokkaido University, Sapporo 060-0812, Japan). Anyone interested in having this proceedings is requested to send a purchase order form, obtainable from the following URL, by FAX. http://www.hokudai.ac.jp/pharma/seika/ISOBA.htm The Table of Contents is included below and can also be found on the ISOBA website.
3. New major monographs on ascidian taxonomy. Drs. Françoise
and Claude Monniot have published Ascidians from the tropical western
Pacific. Zoosystema 23: 201-383 (2001). With so little known about
the ascidians from this large part of the world, this excellent new work
representing many years of collecting and identifications by the Monniots
will be a standard reference for many years. We have used it extensively
already and find it an invaluable addition to the literature.
Dr. Patricia Kott has published the fourth and
last volume of her set of monographs on the Australian ascidians: Kott,
P. 2001. The Australian Ascidiacea part 4, Aplousobranchia (3) Didemnidae.
Mem. Queensland Mus. 47: 1-407. This monumental volume focuses
entirely on the didemnids and is the product of many years of work by Dr.
Kott.
Both volumes contain descriptions of many new species.
4. Web site about Development of Ascidians, submitted by Dr.
Christian
Sardet.
Our new web site has a section on ascidians you may like to consult.
Connect directly at http://www.obs-vlfr/
and click on Biomarcell. We will complete the site in the next few months
and your comments will help us. Christian Sardet, BioMarCell, UMR
7009 CNRS/UPMC, Station Zoologique, Villefranche sur Mer 06230. Email:
sardet@obs-vlfr.fr
5. There will be a joint meeting of the British Soc. for Cell Biology (topic: Cell Regulation through Molecular Machines) and the genetics society and British Soc. for Developmental Biology (topic: Evolution of Developmental Mechanisms) at the Univ. of York, 20-23 March 2002. Further information is available at http://www.bscb.org . There will undoubtedly be some ascidian papers presented and we hope the presenters will send us their abstracts for the spring 2002 AN.
6. Dr. Teruaki Nishikawa contributed the following obituary of his deeply
revered mentor and major professor. We are greatly saddened by the
death of this remarkable man.
Dr. Takasi Tokioka, Professor Emeritus of Kyoto
University, passed away on the 30th of September, 2001, at the age of almost
88. He was a distinguished taxonomist of sessile and pelagic tunicates,
chaetognaths, ctenophorans, crustaceans (Argulus), etc. After his retirement
in 1977, he continued to have good days with his beloved wife and daughter
at his home near the Seto Marine Biological Laboratory, where he worked
since 1962 (as the director from 1975 to 1977), in the town of Shirahama,
Wakayama Pref., Japan. Though suffering from heart disease and poor sight
in his later days, he retained a warm heart, a strong will, and a sharp
brain. Unfortunately, a serious heart attack suddenly stopped his wide-ranging
considerations on animal phylogeny and evolution. At his request, the body
was donated to science, and a simple funeral ceremony was attended by his
close relatives only.
Dr. Tokioka published ca. 220 works from 1936 on, many
appearing in the "Publications of the Seto Marine Biological Laboratory".
His last ones were chapters on ctenophorans and chaetognaths contributed
to a Japanese book, "Dobutsu Keito Bunruigaku [Systematic Zoology], Supplement"
(Nakayama-shoten, 2000), where he gave original phylogenetic considerations
of these two animal groups with a new classification of ctenophorans. He
had a sincere hope to revise the chapters for English publication, but
this was not to be. His tireless scientific activities with deep insights
till his latest days were obviously made possible by his extensive and
detailed biological knowledge, derived from his very long, various taxonomic
and morphological experiences.
As may be well known by readers of "Ascidian News", his significant
contributions to tunicate taxonomy are represented by ca. 60 works on ascidians
and ca. 30 on pelagic tunicates with accurate descriptions and exact (also
very artistic and beautiful) figures. Among others, "Ascidians of Sagami
Bay" (Iwanami-shoten, 1953) and "Pacific Tunicates of the United States
National Museum" (Smithsonian Institution, 1967) are really significant
works. The specimens treated in the latter were examined during his stay
at the museum in 1957, supported by the National Academy of Sciences, USA.
Furthermore, his 1971 paper of "Phylogenetic speculations of the Tunicata"
has been often cited in discussions of chordate phylogeny.
From 1967 to 1969, Dr. Tokioka was one of the leaders
of a grass-roots movement against the reclamation of land from a shallow
inlet in Shirahama, although the result was unsuccessful. Nevertheless,
his consistent interests in and activities for conserving the environment
bore fruit in that an island in Tanabe Bay off Shirahama survived a plan
to turn it into a pleasure park by being purchased by Kyoto University
in 1968. Since then, the island has been protected as a reserve for marine
biological studies, and he began an intertidal census of biota there.
In his private life, he was a famous philatelist, even publishing a
book for stamp collectors probably in 1949. His huge world-wide collection
of stamps, as well as old biological books, will probably be donated to
the Yamaguchi Prefectural Museum, because he loved Yamaguchi where he was
born (in 1913) and lived until entering into Kyoto Imperial University.
He and his family were also famous in Shirahama for keeping many cats once
discarded in front of his home by those who looked for their kind care.
They truly loved the cats by giving them individual names, as he did for
many ascidian species.
Teruaki Nishikawa (The Nagoya University Museum)
WORK IN PROGRESS
1. Gian-Luigi Russo and Elisabetta Tosti (Stazione Zoologica “Anton Dohrn”, Naples, Italy; glrusso@alpha.szn.it) and colleagues M. Tosto, A. Cuomo and I. Castellano have a number of projects in progress regarding the molecular mechanisms regulating meiosis resumption in the ascidian Ciona intestinalis. Now, they have a paper in preparation describing the role of protein kinase CK-2 (formerly known as casein kinase II) at fertilization. In vertebrates, this enzyme is formed by two catalytic subunits (alpha and/or alpha’), and two regulatory subunits, beta. Tosti’s group cloned the CK2 beta subunit in C. intestinalis (Gene Bank accession number AF360544) and characterized the activity of the holoenzyme during meiosis resumption. It is worthwhile to note that in Xenopus oocytes, CK-2 beta subunit interacts and blocks the activity of c-mos, the main component of the cytostatic factor leading to the hypothesis that CK-2 might play an important role in meiosis regulation. The cloning and molecular characterization of CK-2 catalytic subunit, alpha, is in progress and will hopefully be completed by the end of the year.
2. Blood cell vanadium storage in Ascidia ceratodes shows
ecological variations. An X-ray absorption spectroscopic study. Patrick
Franka,b, Robert M. K. Carlsonc, Elaine J. Carlsond
and Keith O. Hodgsona,b
a. Dept. of Chemistry, Stanford Univ., Stanford, CA 94305; b. Stanford
Synchrotron Radiation Laboratory, SLAC, Stanford Univ., Stanford, CA 94309;
c. Chevron Petroleum Technology Co., Richmond, CA 94802; d. Buck Institute,
Novato, CA 94945.
Vanadium storage in the blood cells of the
tunicate Ascidia ceratodes resident in Bodega Bay, California was
assessed using x-ray absorption spectroscopy (XAS). Six blood cell samples,
representing between 1 and 6 animals, were prepared and frozen on the docks
of the Spud Point Marina. About 96% of the blood cell vanadium was
found stored as V(III)-sulfate in aqueous solution, implying a high-acid,
high sulfate environment. No evidence was found for a systematic
change in vanadium storage following admixture of blood cells from different
animals, implying a lack of connection between vanadium distribution and
immune response. A divergent distribution of V(III) complexes between individual
specimens indicated that blood cell vanadium is not tightly regulated.
Comparison of the average vanadium distributions within blood cells from
A.
ceratodes from Monterey Bay, California (~ 200 km south) with those
of Bodega Bay revealed a greater preponderance of V(III) in a pH~0 intracellular
environment in the Bodega Bay ascidians.
3. H. Abdul Jaffar Ali, Ph.D student under Dr. V. Sivakumar, V. O. Chidambaram College, Tuticorin 628 008, India and a junior research fellow working on marine biodiversity and taxonomy of Indian ascidians under Dr. V.K. Meenakshi. I am comparing population distribution, associated flora and fauna, microbial load, heavy metal accumulation, and proximate compositions of Phallusia nigra between the east and west coasts along the southern peninsula of India. I am also analyzing bioactive compounds from P. nigra. Any directions and suggestions are most welcome. zaphar2002@yahoo.co.in
THESIS ABSTRACTS
1. Functional and biochemical characterization of cytostatic factor
(CSF) in the ascidian Ciona intestinalis. Immacolata Castellano,
M.S. thesis, Laboratory for Cell Biology, Stazione Zoologica “Anton Dohrn”,
Naples, Italy. Advisor Dr. Elisabetta Tosti.
In this thesis, we demonstrated the presence of an active
CSF in extracts of C. intestinalis oocytes arrested in metaphase
I, that, when microinjected into blastomers, was able to block their division.
In addition, we presented evidence that a CSF mechanism mediated by protein
kinase Mos is active in ascidian oocytes; in fact, an antibody raised against
Xenopus
Mos, was able to block the ascidian CSF. Our data are partially in agreement
with the proposed role of Mos in oocyte maturation in vertebrate. In fact,
in C. intestinalis a Mos-like protein seems to be expressed constitutively
during the meiotic division as in Xenopus; however, in Vertebrate,
Mos is degraded by the proteasome complex at the metaphase-anaphase II
transition, i.e. after the removal of metaphase block at fertilization.
Same fate is due to MAP kinase activity, another component of CSF. On the
other hand, in C. intestinalis oocytes, Mos expression is still
detected after metaphase I-anaphase I transition, and MAP kinase activity
is maximal after five minutes from fertilization. Finally, we demonstrated
that the metaphase I block in C. intestinalis is independent from
the protein synthesis, similarly to vertebrate, a situation that does not
find similarity among other invertebrates, where the existence of short-lived-proteins
seems to regulate meiotic arrest. This finding further confirms that meiosis
regulation in ascidians resembles the vertebrate one better than the invertebrate
one, in agreement with the position of these organisms in the evolutionary
tree.
2. Reproductive strategies in colonial ascidians: relationships with
other life-history traits and genetic structure. Isabel Tarjuelo. Ph.
D. Thesis. University of Barcelona, X. Turon thesis advisor. isabelt@bio.ub.es
Current theory on the investment in reproduction
by invertebrates has been mainly developed for groups with both lecitotrophic
and planktotrophic larval developmental modes, and this dichotomy explained
a great deal of the diversity of strategies found. Colonial ascidians,
in spite of a uniform (lecitotrophic) developmental mode and a prevalence
of brooding, feature a wide range of fecundities, larval sizes, and reproductive
investment. We wanted to quantify these parameters in a range of species
and to look for correlates of the reproductive traits that may explain
the variability found. We quantified fecundity, reproductive investment
(in terms of larval production), and investment in tunic production in
11 colonial forms. There was a wide range (about one order of magnitude)
of variation in all parameters. Tunic production was related to growth
form, and was higher in encrusting/massive forms than in stoloniferous
forms. It bears no relationship, however, with reproductive investment.
The latter was positively related to the degree development of juvenile
structures in the larva, and negatively to zooid weight, fecundity and
absolute larval weight.
For six of the species, we further studied the biological
cycles, palatability and energy contents of tunic, zooids and larvae, growth
rates, and male reproductive investment. No relationship was found between
adult and larval palatability. Larvae of those species with lowest fecundity
and largest larval sizes were better defended than small larvae. Energy
content and the amount of inorganic material were related to palatability.
Seasonal reproductive cycles, but with varying length and season of the
reproductive period, were observed over three years for the six species.
Species with high growth rate tended to have a short brooding period, low
tunic production, and produced a high number of small, palatable larvae
with a low degree of structural complexity. On the other hand, species
with low growth rates tended to have long brooding periods and produced
a low number of big, complex and well-defended larvae. Total reproductive
investment was higher in the low-fecundity species. We chose two species
representative of these two strategies, Clavelina lepadiformis and
Pseudodistoma
crucigaster, to analyze and compare dispersal capabilities and juvenile
mortality. Our prediction was that both would be higher in the species
with higher fecundity and a more opportunistic strategy (C. lepadiformis).
Dispersal was indirectly estimated through population structure using COI
partial sequences of several populations from the Spanish Mediterranean
littoral. We found a high degree of genetic structure in both species.
In C. lepadiformis this was related to the distinction, possibly
at the species level, between the form inhabiting harbours and the population
in the open shore habitats. In P. crucigaster, genetic differences,
although not so important as to justify a species-level distinction, were
found among color morphs. When comparable populations were studied, however,
the degree of gene flow at a scale of tens of kilometres was ten times
higher for C. lepadiformis. On the other hand, post-settlement mortality
(during the first four weeks of benthic life) was three times higher in
C.
lepadiformis. It is concluded that there is a continuum of reproductive
strategies in colonial ascidians, the extremes of which are the production
of a high number of small larvae vs. the production of a single, complex
larva per zooid. These extremes have important correlates in terms of allocation
to reproduction and defense, growth rates, tunic production, dispersal
capabilities and larval and post-metamorphic mortality.
3. Comparative biology of Clavelina lepadiformis (Ascidiacea)
populations from inside and outside harbours in the western Mediterranean.
Sònia de Caralt, Master’s thesis, University of Barcelona, X. Turon
thesis advisor. socabo@latinmail.com
Clavelina lepadiformis is a colonial ascidian
that inhabits both harbour environments and open sea rocky communities.
Recent investigations have shown that the populations of these habitats
exhibit a marked genetic divergence and a highly restricted genetic flow
between them. However, no morphological differences between the two forms
could be substantiated. In this work we compared the biology and heavy
metal accumulation of populations of these two forms. Our goals were to
investigate whether the genetic isolation is reflected by adaptive divergences
in the two types of habitat, and to ascertain whether the two forms can
survive in each other’s habitat. The abundance and seasonal cycles showed
contrasting trends in the two types of habitat: the populations inside
harbours reach densities of ca. 3000 zooids m-2, and active colonies were
found all year round. In the open littoral habitat, abundance was an order
of magnitude lower and showed a clear seasonal pattern, with disappearance
of zooids during the summer season (aestivation). The reproductive cycles
were also different, as larvae were present in the populations inside harbours
from November through July, with several sexual cycles during this period,
while in the outer habitats larval appearance was restricted to 2-3 months
during winter-spring, with only one gonadal cycle per year. The zooids
of the internal form were significantly bigger than those from the exterior
populations, and so were the larvae produced, but the total reproductive
effort and tunic production (in weight ratios), as well as the fecundity,
were not significantly different between habitats. The populations from
inside harbours accumulate significantly more Cu and Pb than the exterior
populations, and the heavy metal concentrations showed a seasonal cycle
with minima in summer. On the other hand, both varieties accumulate a similar
amount of vanadium, a metal known to play a role in ascidian metabolism.
Both the production of secondary metabolites and the toxicity of polar
extracts were higher in the external populations than in those from the
interior habitats. These results, however, did not correlate with the outcomes
of palatability tests with specialist and generalist predators, where no
preference could be substantiated. An experiment of juvenile transplantation
between habitats showed that newly-settled individuals from the external
habitat can survive in both habitats (with survival figures of 30-50% during
the first four weeks), while juveniles from the internal form survive very
poorly in the outside habitat (ca. 5% survival after 4 weeks). It appears
that both varieties showed marked contrasts in most biological parameters
studied. The variety inhabiting harbours reaches very high densities, produces
several sexual generations per year and seems to withstand heavy metal
pollution without deleterious effects. The variety inhabiting open rocky
littoral shows more restricted growth, aestivation phenomena, and a single
larval generation per year. On the other hand, the internal population
produces less secondary chemistry, which may explain its inability to survive
in the external habitat, where predation pressures are presumably higher.
The lack of genetic flow between these two forms suggests that the differences
may not be mere phenotypic adaptations to the different environments and
may have a genetic basis.
4. New Zealand ascidians: masters of amino acid-derived secondary
metabolites.
Allison Norrie Pearce, Ph.D. thesis, Dept. of Chemistry, The Univ.
of Auckland, Auckland, New Zealand. Thesis advisor, Dr. Brent Copp b.copp@auckland.ac.nz
A survey of 34 New Zealand ascidians was performed for
the purpose of investigating the biological and chemical properties of
ascidians from New Zealand waters. The crude extracts were assayed for
biological activity and were also analyzed by HPLC with photodiode array
capability for comparison of absorption spectra. In this manner biological
profiles of the cytotoxicity and antimicrobial activities of the crude
extracts were obtained, as well as a UV metabolic profile. The (-)-enantiomer
of the known compound 1,2,3-trithiane was isolated from the ascidian Hypsistozoa
fasmeriana, and the (+)-enantiomer from the ascidian Distaplia stylifera,
a member of the same family (Holozoidiae). The (+)-enantiomer was originally
isolated from the New Zealand ascidian Aplidium sp. D and appears
to be unique to New Zealand waters. A new class of compound, the fasmerianamines
A and B, derivatives of 1,2,3-trithiane were isolated from a second collection
of Hypsistozoa fasmeriana. Another new class of metabolite, the
fluorescent, biologically inactive distomadines A and B were discovered
in the ascidian Pseudodistoma aureum. A previously unknown trimethylated
purine, 2,2,7-trimethylguanine as well as other known purine bases were
detected and isolated. Much of the biological activity of the ascidian
extracts could be attributed to the presence of unsaturated long-chain
amino alcohol-type compounds. The presence of the pyridoacridine
class of compound, a large class of aromatic polycyclic alkaloids of worldwide
distribution, was detected in a New Zealand ascidian for the first time.
The pyridoacridone alkaloid ascididemin possesses a wide
range of biological activity and progressed as far as in vivo xenograft
antitumoural assays at the NCI, revealing its value as a lead compound.
The antitumoural activity of ascididemin is attributed to the intercalative
ability of its planar pentacyclic structure. The synthetic strategy for
another natural pyridoacridone, kuanoniamine A was optimized, and the synthetic
compound also showed good human solid tumour selectivity, progressing as
far as in vivo hollow fibre testing at the NCI. Three other synthetic
ring A-modified analogues were prepared. Two, combining either a furan
or a thiophene A ring with a carboxylic acid methyl ester substituent in
position two on ring A, were tested at the NCI and were found to be inactive.
This indicated conclusively that the carboxylic acid methyl ester substituent
modification had a detrimental effect on cytotoxicity. The disruption of
the planarity of the heterocyclic ring system with subsequent inability
to intercalate into DNA might be implicated. An isoxazole ring A modified
analogue was found to be too unstable for evaluation purposes.
5. Three thesis titles submitted by Dr. Masaaki Morisawa, Misaki
Marine Station. morisawa@mmbs.s.u-tokyo.ac.jp
Studies on the ion channels regulating the activation of sperm motility
in the ascidians Ciona intestinalis and C. savignyi.
Hiroko Izumi, Ph. D. thesis, Univ. of Tokyo (1999).
Studies on the cell signaling for the activation of sperm motility
in ascidian. Mamoru Nomura, Ph. D. thesis, Univ. of Tokyo (2000).
The study on the mechanism of metamorphosis in the ascidian, Ciona
savignyi. Yikiko Kimura, MS. Thesis, Univ. of Tokyo(2000).
MEETINGS ABSTRACTS
1. Annual Conference of the Italian Embryology Group, Fano, Italy, 7-9 June 2001
Serotonin localization in Phallusia mammillata larvae and during
first events of metamorphosis. R. Pennati, S. Groppelli, C. Sotgia,
U. Fascio, *M. Pestarino and F. De Bernardi, Dept. of Biology, Univ. of
Milano, Italy. *Dept. of Experimental, Environmental and Applied
Biology, Univ. of Genova, Italy.
Serotonin [5-Hydroxytryptamine (5-HT)] is a neurotransmitter
which plays an important role in a wide range of non-neural processes,
during embryonic development such as egg cleavage, gastrulation cell movements
and morphogenesis of many invertebrate and vertebrate embryos and larvae.
The presence of serotonin has been reported in the adults of a variety
of ascidian species, in which was localized in the enterocromaffin cells
of the gut, in the peripharyngeal band and in the endostyle (Georges, 1985.
Cell Tissue Res. 242: 341-348; Pestarino, 1982. Cell Tissue Res.
226:
231-235) but the origin and the distribution of serotonin-containing cells
during the ontogeny of ascidians are not known.
We used an anti-serotonin antibody under a confocal
microscope to localize endogenous serotonin by immunofluorescence in the
swimming and metamorphosing larvae of the ascidian Phallusia mammillata.
The serotonin-containing cells in the swimming larvae were detected in
the central nervous system. Bright fluorescence is present in 11 pear-shaped
cells surrounding the ocellus, corresponding to the retinal cells of the
photoreceptor complex, in at least one of the elongated cells of each papilla
and in the pericharia of two epidermal neurons of the rostral trunk, which
are present on the axon-like fibers connecting the papillae with the sensory
vesicle. In the tail, serotonin is evident in the pericharia of two groups
of neurons in the ventral and in the dorsal epidermis, connected by immunopositive
fibers. These neurons are often paired but their distribution and their
number varied among the larvae. An immunofluorescent signal is found in
some ventral endodermal cells of the anterior trunk. These cells may contribute
to the formation of the posterior half of the adult endostyle.
During all the swimming larva period the serotonin
is detectable principally in the nervous system as described above, when
larvae stop swimming and tail retraction begins, the immunofluorescent
signal in the cells of photoreceptor complex progressively faints together
with signal in the neuroepithelial cells of the rostral part of the trunk.
At the end of tail retraction, five days after fertilization, when the
ampullae definitively keep the larvae attached to the substrate, serotonin
appears in the cells of the peripharingeal band, of the endostyle and of
the gut.
The serotonergic system may control the complex
larval behaviour in response to light. In fact the larvae of the solitary
ascidians show a positive phototaxis during the dispersal-phase of their
life, but at the time of settlement, most ascidian larvae avoid light and
prefer to settle on dark or shaded surfaces (Cloney, 1982. Amer. Zool.
22:
817-826)
Metamorphosis transforms a swimming larva into a
sessile juvenile. The early photoreceptive function of the serotonergic
system may be lost, together with the reduction of sense organs, and serotonin
becomes restricted to other systems. In fact after metamorphosis we were
not able to detect serotonin in the neural complex of the juveniles. Experiments
carried out treating newly hatched swimming larvae with ritanserin, an
antagonist of 5-HT2 receptor subtype, significantly shorten the swimming
period. A 6 hour treatment with 10µM ritanserin induces attachment
to the substrate of 70 % of larvae. After 24 hours of treatment all larvae
are attached to the substrate by the papillae, while 80% of control larvae
are still swimming. This suggests a photoreceptive or a photoneuroendocrine
function of serotonin for ascidian larvae similar to that observed in amphioxus
and in vertebrates (Terio, 1964. Atti Soc. Peloritana Sc. Fis., Mat. e
Naturali 10: 111-125; Nunez et al., 1981. J. Histochem. Cytochem.
29:
1336-1346).
2. Congress of the Italian Society of Neuroscience, Torino, Italy, 8-11 September 2001.
Localization and possible role of serotonin during the development
of the ascidian Phallusia mammillata. R. Pennati, S. Groppelli,
C. Sotgia and F. De Bernardi, Dept. of Biology, Univ. of Milano, Italy.
It is known that the neurotransmitter serotonin
[5-hydroxytryptamine (5-HT)] plays an important role in a wide range of
non-neural processes. Its activity is mediated by multiple receptors subtypes.
5-HT is present in early stages of sea urchin embryo development (Buznikov
et al., 1964, 1972) and it has been demonstrated that it has a key role
during the morphogenesis of chicken and mouse embryo (Gustafson and Toneby,
1970, Moiseiwitsch and Lauder, 1995). By immunofluorescence with an anti-serotonin
antibody,we demonstrated the presence and localization of 5-HT in brain
and in some neurons of the larval tail of Phallusia mamillata.
To test the effects of serotonin on ascidian
development, we treated embryos with different pharmacological substances:
fluoxetine, an inhibitor of 5-HT reuptake, and three antagonist for different
serotonin receptor subtypes. The treatments were performed at 10 micromolar
concentration, begun before cleavage or at late gastrula stage, and were
terminated at swimming larva stage. Malformations induced by the treatments
were analyzed successively (i) at the end of the treatment, (ii) 7 days
after fertilization, corresponding to the beginning of metamorphosis, and
(iii) 15 days after fertilization, corresponding to a completely metamorphosed
juvenile. The early treatment of fluoxetine caused anomalies of the segmentation
and malformations , consisting mainly in roundish cephalenteron and short
tail, in 50% of larvae developed from treated embryos. Few larvae metamorphosed
and 15 days after fertilization juveniles showed reduction of the neural
ganglion size.
Treatments with serotonin antagonists were more
efficient from gastrula stage onwards. WAY, an antagonist for the 5-HT1A
receptor subtype, caused extreme reduction of brain vesicle and of sensory
organs; ondansetron, an antagonist for the 5-HT3 receptor subtype, induced
high incidence of malformations of cephalenteron and short tails in later
treatment. Juveniles developed almost normally. Ritanserin, a substance
with a high affinity for the 5-HT2 receptor subtype, caused mainly truncation
of the anterior end of larvae cephalenteron. The juveniles developed from
these larvae had an abnormal cardio-circulatory system: no blood circulation,
the heart contractions were no-rhythmic, and the blood cells were less
numerous than in controls. We conclude that serotonin play an important
role in the morphogenesis of the ascidian. An appropriate level of serotonin
is necessary for a correct segmentation, since increased concentration
caused by the inhibitor of 5-HT reuptake, fluoxetine, severely perturbed
this early phase of embryo development. During morphogenesis the action
of the serotonin is mediated by different receptor subtypes, among which
5-HT1A, 5-HT2 and 5-HT3 appear to play a key role in modulating the multiple
morphogenetic effects of serotonin.
3. 4th Workshop "A. Dohrn" New Perspective in Tunicate Biology, Ischia, Italy, 29 September - 2 October 2001.
Sperm chemotaxis in ascidians. M. Morisawa and M. Yoshida, Misaki
Marine Biol. Station, Graduate School of Sci., Univ. of Tokyo, 1024 Koajiro,
Misaki, Miura, Kanagawa, 238-0225 Japan.
Spermatozoa of the ascidians Ciona are immotile
in seawater. Motility is activated around the egg, and then the activated
sperm are attracted towards the egg, suggesting that the factor for sperm
activation and attraction is released from eggs at fertilization (Yoshida
et al., Dev. Biol., 1993). The factor named as sperm-activating and -attracting
factor (SAAF) in Ciona (Yoshida et al., DGD, 1997) was recently
purified and identified as a steroid with molecular mass of 596 (Yoshida
et al., submitted). Cell signaling for sperm activation has been clarified
(Izumi et al., Dev. Biol., 1999; Nomura et al., DGD, 2000). Ca2+ introduced
into the sperm cell through Ca channel activates calmodulin-calmodulin
kinase system to hyperpolarize the plasma membrane, and the change in membrane
potential synthesizes cAMP via activation of adenylyl cyclase. The
cAMP phosphorylates dynein light chain and a 26kDa protein, and the phosphorylation
triggers the final step for the activation of sperm motility.
Sperm chemotaxis does not require cAMP, and Ca is the
only prerequisite second messenger (Yoshida et al., DGD, 1997), although
cell signaling cascade underlying the sperm chemotaxis in ascidians as
well as other species is unclear. Nifedipine, Flunarizine, omega-conotoxin
had no effects on chemotaxis of Ciona sperm, but SK&F96365,
Ni2+ and Zn2+, blockers of store-operated Ca2+ channel completely inhibited
chemotactic behavior of sperm. Furthermore, depletion of internal Ca2+
store by thapsigargin, caused increase in intracellular Ca2+. These
suggest that store-operated calcium channel exists in the ascidian sperm
and participates in the chemotactic behavior of the sperm.
Quantitative analysis of sperm trajectories measuring
four parameters: distance and orientation between SAAF and sperm head,
radius of curvature of sperm trajectory; and sperm velocity and computer
simulation proposed model shows that the change of SAAF concentration controls
chemotactic behavior of ascidian sperm.
4. 11th Intl. Symp. On Environmental Pollution and its Impact on Life in the Mediterranean Region, 6-10 Oct. 2001.
Influence of TBT on the activity of the detoxicant enzymes, GST and
GPX, from haemocytes of the colonial ascidian Botryllus schlosseri.
F. Cima, D. Dominici, L. Ballarin and P. Burighel, Dept. of Biology, Univ.
of Padova, Italy.
In the last decade, some authors have hypothesized that
a considerable part of the immunotoxic effects described for tributyltin
(TBT) in mammals was due to Ca2+-independent mechanisms involving changes
in sulphydryl groups of both proteins and reduced glutathione (GSH).
Among the detoxicant enzymes, which play a multiple role in the metabolism
of many xenobiotics of invertebrates and vertebrates, there are glutathione-transferase
(GST) and glutathione-peroxidase (GPX). GST catalyzes the conjugation
reactions of GSH with electrophilic xenobiotics or their derivatives.
It has been described in the antioxidant defense systems mainly of the
gill of bivalves and as a mediator of xenobiotic detoxification in the
liver and kidney of marine fish. In mammals, the activation of GST
has been proposed to occur after TBT treatment, and the presence of mercapturic
acid supports this hypothesis. However, this derivative was never
found in molluscs, suggesting that invertebrates possess a different detoxicant
system of organotins.
GPX is an antioxidant enzyme, which protects from the
effects of reactive oxygen species production, requiring GSH consumption.
In marine fish, since the inhibition of GPX by various organotin compounds
had been recently described, this enzyme can be used as an effective biomarker
of these pollutants. High concentrations of GPX were found in the
blood of many invertebrates. The GPX activity detected inside the
gill of bivalves is associated with GST and is catalyzed by this enzyme.
In cultured haemocytes of the colonial ascidian Botryllus
schlosseri, we have observed by immunocytochemistry that positivity
to both GST and Se-dependent GPX antibodies disappeared after incubation
for 60 min at 25o C, in the presence of 0.1 µM and 1 µM
TBT, respectively. This suggests that conformational changes occur, probably
due to direct interaction of the xenobiotic with these antioxidant enzymes.
Moreover, the spectrophotometric assays on the haemocytic lysate have shown
that TBT was able to significantly inhibit the GST activity (µmoles/min/mg
protein) 2.6 times at 0.01 µM and 4.8 times at 0.1 µM, whereas
the Se-dependent GPX activity was inhibited 1.5 times at 0.05 µM
TBT. Our data suggest that in ascidian blood cells, enzymes involved
in detoxification metabolic processes are present and that GST is the most
important and sensitive among them.
Table of contents for: The Biology of Ascidians, edited by H. Sawada, H. Yokosawa and C. C. Lambert, the symposium volume published 2001 by Springer-Verlag Tokyo resulting from The First Intl. Symposium on the Biology of Ascidians, Hokkaido University, Sapporo, Japan, June 26–30, 2000. If you have not yet purchased a copy, please go to the website http://www.hokudai.ac.jp/pharma/seika/ISOBA.htm. Please request that your institution library also acquire a copy.
1. Fertilization and Egg Activation
Structural and Molecular Investigations on the Egg Coat
in Phallusia mammillata. Thomas G. Honegger and Monika
Füglister 3
Analysis of the Self-sterility in Halocynthia roretzi.
Naoyuki
Murabe and Motonori Hoshi 9
Further Observations on the Molecular Bases of Gamete
Self-discrimination in Ciona intestinalis: Seasonal Variation of
Self-sterility Rate.
Rosaria De Santis, Rita Marino, and Maria Rosaria
Pinto 14
Self-nonself Recognition and Lysin System in Fertilization
of the Ascidian
Halocynthia roretzi. Hitoshi Sawada and Hideyoshi
Yokosawa 18
Fertilization-induced Glycosidase Release and Interspecific
Sperm Competition in Ascidians. Charles C. Lambert 24
Cell Signaling in Ascidian Sperm: Upstream and Downstream
of Internal Calcium Release. Robert A. Koch, Kathleen Allen, Ju Kim,
and Ali Lotfizadeh 30
Sperm-triggered Calcium Oscillations at Fertilization.
Alex
McDougall 36
Diversity of Calcium Channels Involved in Meiosis Resumption
of Ascidian Oocytes. Mireille Albrieux, Christophe Arnoult, Didier Grunwald,
Marie-Jo Moutin, and Michel Villaz 47
Ascidian Sperm Acrosin and Spermosin: Structures and
Roles in Fertilization. Eri Kodama, Tadashi Baba, Hideyoshi Yokosawa,
and Hitoshi Sawada 54
Acrosome Differentiation in Ciona intestinalis Spermatozoa
and Some Speculations on Ascidian Fertilization. Makoto Fukumoto 60
Follicle Cells of Styela plicata Eggs (Ascidiacea). Luisanna
Villa and Eleonora Patricolo 67
High Level of Protein Ubiquitination in Ascidian Sperm.
Kazuo
Inaba 74
Calcium Transients Signal Ooplasmic Segregation through
the Small GTPase rho in Ascidian Eggs. Manabu Yoshida, Yuji Horiuchi,
and Masaaki Morisawa 81
Cell Signalings for Activation of Motility and Chemotaxis
in the Sperm of Ciona. Masaaki Morisawa, Hiroko Izumi, Manabu
Yoshida, and Yoshitaka Oka 86
Roles of MLCK and PI3 Kinase on Deformation and Ooplasmic
Segregation at Fertilization in the Egg of Ciona savignyi. Noburu
Sensui, Manabu Yoshida, and Masaaki Morisawa 92
Identification of Phallusia mammillata Egg ?-N-Acetylhexosaminidase
with a Potential Role in Prevention of Polyspermy. Markus Eisenhut and
Thomas G. Honegger 97
The “Complex” Ascidiosperm of Aplousobranchs. GianBruno
Martinucci, Mila Della Barbera, Francesco Boldrin, and Paolo Burighel
102
2. Reproductive Biology and Neuroscience
The Origin of Germ Cells in Ciona intestinalis.
Katsumi
Takamura 109
Oocyte Maturation and Self-sterility by Treatment with
Ovary Extracts of the Ascidian, Halocynthia roretzi. Takaharu
Numakunai 117
Induction of Gamete Release by Gonadotropin-Releasing
Hormone (GnRH) in Ciona intestinalis. Kiyoshi Terakado 125
Light Regulated GnRH Neurons in Biological Clock for
Reproduction in the Ascidian, Halocynthia roretzi. Motoyuki Tsuda,
Mahito Ohkuma, Masashi Nakagawa, and Yasuo Katagiri 131
Tailbud Embryogenesis and the Development of the Neurohypophysis
in the Ascidian Ciona intestinalis. Alison G. Cole and Ian A.
Meinertzhagen 137
The Peripheral Nervous System of an Ascidian Revealed
by AChE Activity. Lucia Manni, Marina Sorrentino, Giovanna Zaniolo,
and Paolo Burighel 142
Heterotrimeric G Protein ??and ??Subunit Genes of the
Ascidian, Halocynthia roretzi. Tatsuo Iwasa, Kazue Kanehara,
Ayako Watari, Mahito Ohkuma, Masashi Nakagawa, and Motoyuki Tsuda 147
Photoresponse and Habituation of Swimming Behavior of
Ascidian Larvae, Ciona intestinalis. Motoyuki Tsuda, Isao Kawakami,
Takayuki Miyamoto, Masashi Nakagawa, Shuhei Shiraishi, and Muneki Gouda
153
Multiple cis-Regulatory Regions Control Neuronal Gene
Expression of Synaptotagmin in Ascidian Embryos. Jun Matsumoto, You
Katsuyama, and Yasushi Okamura 158
3. Development, Differentiation, and Evolution
Maternal Genetic Information Stored in Fertilized Eggs
of the Ascidian, Halocynthia roretzi. Kazuhiro W. Makabe, Takeshi
Kawashima, Shuichi Kawashima, Yasunori Sasakura, Hisayoshi Ishikawa, Hiroshi
Kawamura, Minoru Kanehisa, Takahito Nishikata, and Hiroki Nishida 165
RNA-binding Proteins in Ascidian Development. Takahito
Nishikata, Michiko R. Wada, and Kimio J. Tanaka 178
Functional Analysis of Ciona intestinalis Y-Box
Protein. Kimio J. Tanaka and Takahito Nishikata 186
Ci-sna cis-Regulation of Ascidian Tail Muscle Genes.
Albert
Erives and Michael Levine 193
T-box Genes and the Development of Axial Tissues in Ciona
intestinalis. Anna Di Gregorio and Michael Levine 202
Cloning and Embryonic Expression of HrzicN, a Zic Family
Gene of the Ascidian Halocynthia roretzi. Shuichi Wada and Hidetoshi
Saiga 206
Analysis of a cis-Regulatory Element of Hroth, the Ascidian
Homologue of the otx Genes, That Drives Its Transcription in the Anterior
Larval Central Nervous System of the Ascidian, Halocynthia roretzi.
Izumi
Oda and Hidetoshi Saiga 211
Comparison of the Structure and Expression of otx Genes
between Ciona intestinalis and Halocynthia roretzi. Nanami
Utsumi and Hidetoshi Saiga 215
Phylogeny of the Urochordates: Implications for Chordate
Evolution. Billie J. Swalla 219
Evolution of Anural Developmemt in Ascidians: Roles of
Muscle-Specific Differentiation Genes. Takehiro Kusakabe 225
Maximum Direct Development and the Ascidiotypic Stage.
William
R. Bates 230
The Origin of the Neural Crest and Insights into Evolution
of the Vertebrate Face. Hiroshi Wada 235
Participation of Neurotransmitters and Adrenergic Receptor
in the Metamorphosis of Ascidian Larvae. Yukiko Kimura, Manabu Yoshida,
and Masaaki Morisawa 241
4. Taxonomy and Ecology
A Global Overview of Ascidian Introductions and Their
Possible Impact on the Endemic Fauna. Gretchen Lambert 249
Settlement and Metamorphosis of the Tropical Ascidian
Herdmania
curvata. [now synonymized under H. momus--GL]
Bernard
M. Degnan 258
Ascidians in Brazil: The State of the art of Research
in Taxonomy, Ecology and Natural Products. Rosana M. Rocha and Roberto
G. S. Berlinck 264
The Biological Substratum Eudistoma carolinense Van Name,
1945 in the Beach Itapema do Norte, Santa Catarina, Brazil. Tatiane
R. Moreno and Rosana M. Rocha 271
Ascidians of South Africa: A Historical Perspective.
Shirley
Parker-Nance 278
Mitochondrial DNA Analysis of Boltenia echinata iburi
(Oka, 1934). Tsuneo Kakuda 283
5. Colonial Ascidians
Molecular and Cellular Advantage of Transdifferentiation
System for Asexual Reproduction of the Tunicate, Polyandrocarpa misakiensis.
Kazuo
Kawamura 293
Molecular Bases of Bud Development in Ascidians. Shigeki
Fujiwara, Mika Kamimura, Mitsuko Ohashi, and Kazuo Kawamura 300
Laboratory Studies of Mating in the Aplousobranch Diplosoma
listerianum. John D. D. Bishop, Andrew J. Pemberton, A. Dorothea
Sommerfeldt, and Christine A. Wood 305
Environmental Effect on the Reproductive Effort of Botryllus
schlosseri. J. Stewart-Savage, Anne Stires, and Philip O. Yund
311
Phylogeny of Botryllid Ascidians. Yasunori Saito,
Maki Shirae, Makiko Okuyama, and Sarah Cohen 315
Epithelial Differentiation in the Dorsal Strand of a
Budding Ascidian, Polyandrocarpa misakiensis (Protochordata, Ascidiacea).
Hiromichi
Koyama 321
6. Biologically Active Substances
Lumichrome Is a Putative Intrinsic Substance Inducing
Larval Metamorphosis in the Ascidian Halocynthia roretzi. Sachiko
Tsukamoto, Haruko Kato, Hiroshi Hirota, and Nobuhiro Fusetani 335
Biological Activity and Chemistry of the Compound Ascidian
Eusynstyela
tincta. S. K. Chithra Lekha Devi, K. N. Rajasekharan, K. Padmakumar,
Jun'ichi Tanaka, and Tatsuo Higa 341
Aquaculture of Ecteinascidia turbinata Herdman,
1880 as Source of Marine Anticancer Agents. S. A. Naranjo, H. B. Kukurtçu,
C. Barbero, S. Martin, and J. L. Carballo 355
7. Heavy Metals
The Mechanism of Accumulation and Reduction of Vanadium
by Ascidians. Hitoshi Michibata, Taro Uyama, Tatsuya Ueki, and Kan Kanamori
363
Immunotoxicity in Ascidians: the Case of Organotin Compounds.
Loriano
Ballarin and Francesca Cima 374
8. Host Defense Mechanisms
Immunodefense in Tunicates: Cells and Molecules. Edwin
L. Cooper and Nicolò Parrinello 383
Immunological Activity of Ascidian Hemocytes. Nicolò
Parrinello, Matteo Cammarata, Mirella Vazzana, Vincenzo Arizza, Aiti Vizzini,
and Edwin L. Cooper 395
Identification of Type I and IX Collagens in the Ascidian
Ciona
intestinalis. Aiti Vizzini, Vincenzo Arizza, Melchiorre Cervello,
Cinzia Chinnici, Matteo Cammarata, Roberto Gambino, Eleonora Patricolo,
and Nicolò Parrinello 402
Primitive Complement System of the Solitary Ascidian,
Halocynthia
roretzi. Seita Miyazawa, Kaoru Azumi, and Masaru Nonaka 408
Aggregation, Tyrosine Phosphorylation, and Gene Expression
in Hemocytes of the Ascidian Halocynthia roretzi. Kaoru Azumi
and Hideyoshi Yokosawa 414
Common Cell Surface Ligands Functioning in Allogeneic
Cytotoxic Reaction and Fertilization in Halocynthia roretzi. Makoto
Arai, Shin-Ichi Ohtake, Hiroyoshi Ohba, Kunio Tanaka, and Joe Chiba
419
Allorecognition and Microsatellite Allele Polymorphism
of Botryllus schlosseri from the Adriatic Sea. Baruch Rinkevich,
Guy Paz, Jacob Douek, and Rachel Ben-Shlomo 426
Isolation of Marine Birnavirus from Sea Squirts Halocynthia
roretzi. Sung-Ju Jung, Myung-Joo Oh, Tatsuya Date, and Satoru Suzuki
436
Colony Specificity in Botrylloides leachi (Savigny):
Preliminary Reports. Giovanna Zaniolo and Loriano Ballarin 442
The Viriform Cell of Halocynthia roretzi: Fine
Structure, Distribution, and Appearance. Shin-Ichi Ohtake, Teruhisa
Ishii, Makoto Arai, Takeyuki Abe, Fumio Shishikura, Joe Chiba, and Kunio
Tanaka 445
Hemopoiesis in Solitary Ascidians. Tomoo Sawada, Teruhisa
Ishii, and Shin-Ichi Ohtake 450
9. Food Science
Antioxidant Activity of Quinone-derivatives from Freeze-dried
Powder of the Ascidians. Osamu Inanami, Tohru Yamamori, Haruhisa Shionoya,
and Mikinori Kuwabara 457
Gastroprotective Effect of Ascidian, Halocynthia aurantium
(Akaboya), Extract on Acute Gastric Hemorrhagic Lesions in Rats. Hideyuki
Chiji, Chizuko Hayashi, and Megumi Matsumoto 463
NEW PUBLICATIONS (also see the Table of Contents above)
Adachi, Y., Nagao, T., Saiga, H. and Furukubo-Tokunaga, K. 2001. Cross-phylum regulatory potential of the ascidian Otx gene in brain development in Drosophila melanogaster. Dev. Genes Evol. 211: 269-280.
Armsworthy, S. L., MacDonald, B. A. and Ward, J. E. 2001. Feeding activity, absorption efficiency and suspension feeding processes in the ascidian, Halocynthia pyriformis (Stolidobranchia: Ascidiacea): responses to variations in diet quantity and quality. J. Exp. Mar. Biol. Ecol. 260: 41-69.
Ballarin, L. 2001. Morula cells as the major immunomodulatory hemocytes in ascidians: evidences from the colonial species Botryllus schlosseri. Biol. Bull. 201: 59-64.
Baylies, M. K. and Michelson, A. M. 2001. Invertebrate myogenesis: looking back to the future of muscle development. Curr. Opin. Genet. Dev. 11: 431-439.
Beaulieu, S. E. 2001. Life on glass houses: sponge stalk communities in the deep sea. Mar. Biol. 138: 803-817.
Bellas, J., Vazquez, E. and Beiras, R. 2001. Toxicity of Hg, Cu, Cd, and Cr on early developmental stages of Ciona intestinalis (Chordata, Ascidiacea) with potential application in marine water quality assessment. Water Res. 35: 2905-2912.
Bishop, C. D., Bates, W. R. and Brandhorst, B. P. 2001. Regulation of metamorphosis in ascidians involves NO/cGMP signaling and HSP90. J. Exp. Zool. 289: 374-84.
Bonetta, L. 2001. Anticancer squirt. Nat. Med. 7: 891.
Burighel, P., Brena, C., Martinucci, G. B. and Cima, F. 2001. Gut ultrastructure of the appendicularian Oikopleura dioica (Tunicata). Invert. Biol. 120: 278-293.
Burighel, P., Sorrentino, M., Zaniolo, G., Thorndyke, M. C. and Manni, L. 2001. The peripheral nervous system of an ascidian, Botryllus schlosseri, as revealed by cholinesterase activity. Invert. Biol. 120: 185-198.
Carballo, J. L., Naranjo, S., Kukurtzu, B., de la Calle, F. and Hernandez-Zanuy, A. 2000. Production of Ecteinascidia turbinata (Ascidiacea: Perophoridae) for obtaining anticancer compounds. J. World Aquacult. Soc. 31: 481-490.
Cima, F., Perin, A., Burighel, P. and Ballarin, L. 2001. Morpho-functional characterization of haemocytes of the compound ascidian Botrylloides leachi (Tunicata, Ascidiacea). Acta Zool. 82: 261-274.
Coma, R., Ribes, M., Gili, J.-M. and Hughes, R. N. 2001. The ultimate opportunists: consumers of seston. Mar. Ecol. Prog. Ser. 219: 305-308.
Connell, S. D. 2001. Predatory fish do not always affect the early development of epibiotic assemblages. J. Exp. Mar. Biol. Ecol. 260: 1-12.
Connell, S. D. 2001. Urban structures as marine habitats: an experimental comparison of the composition and abundance of subtidal epibiota among pilings, pontoons and rocky reefs. Mar. Env. Res. 52: 115-125.
Corbo, J. C., Di Gregorio, A. and Levine, M. 2001. The ascidian as a model organism in developmental and evolutionary biology. Cell 106: 535-538.
Darras, S. and Nishida, H. 2001. The BMP signaling pathway is required together with the FGF pathway for notochord induction in the ascidian embryo. Development 128: 2629-2638.
Darras, S. and Nishida, H. 2001. The BMP/CHORDIN antagonism controls sensory pigment cell specification and differentiation in the ascidian embryo. Dev. Biol. 236: 271-288.
Dumollard, D. and Sardet, C. 2001. Three different calcium wave pacemakers in ascidian eggs. J. Cell Science 114: 2471-2481.
Exposito, A., Fernandez-Suarez, M., Iglesias, T., Munoz, L. and Riguera, R. 2001. Total synthesis and absolute configuration of minalemine A, a guanidine peptide from the marine tunicate Didemnum rodriguesi. J. Org. Chem. 66: 4206-4213.
Farina, J. M. and Castilla, J. C. 2001. Temporal variation in the diversity and cover of sessile species in rocky intertidal communities affected by copper mine tailings in northern Chile. Mar. Pollution Bull. 42: 554-568.
Ferrier, D. E., Minguillon, C., Cebrian, C. and Garcia-Fernandez, J. 2001. Amphioxus Evx genes: implications for the evolution of the midbrain-hindbrain boundary and the chordate tailbud. Dev. Biol. 237: 270-281.
Fontana, A., Cimino, G., Gavagnin, M., Gonzalez, M. C. and Estornell, E. 2001. Novel inhibitors of mitochondrial respiratory chain: endoperoxides from the marine tunicate Stolonica socialis. J. Med. Chem. 44: 2362-2365.
Frank, P., Robinson, W. E., Kustin, K. and Hodgson, K. O. 2001. Unprecedented forms of vanadium observed within the blood cells of Phallusia nigra using K-edge X-ray absorption spectroscopy. J. Inorg. Biochem. 86: 635-648.
Garrido, L., Zubia, E., Ortega, M. J., Naranjo, S. and Salva, J. 2001. Obscuraminols, new unsaturated amino alcohols from the tunicate Pseudodistoma obscurum: structure and absolute configuration. Tetrahedron 57: 4579-4588.
Glasby, T. M. 2001. Development of sessile marine assemblages on fixed versus moving substrata. Mar. Ecol. Prog. Ser. 215: 37-47.
Graber, N. A. and Ellington, W. R. 2001. Gene duplication events producing muscle (M) and brain (B) isoforms of cytoplasmic creatine kinase: cDNA and deduced amino acid sequences from two lower chordates. Mol. Biol. Evol. 18: 1305-1314.
Greenwood, A., O'-Riordan, R. M. and Barnes, D. K. A. 2001. Seasonality and vertical zonation of zooplankton in a semi-enclosed sea lough. J. Mar. Biol. Ass. U.K. 81: 213-220.
Hernandez-Zanuy, A. C. and Carballo, J. L. 2001. Distribution and abundance of ascidian assemblages in Caribbean reef zones of the Golfo de Batabano (Cuba). Coral Reefs 20: 159-162.
Hirose, E. 2001. Acid containers and cellular networks in the ascidian tunic with special remarks on ascidian phylogeny. Zool. Sci. 18: 723-731.
Hirose, E., Yamashiro, H. and Mori, Y. 2001. Properties of tunic acid in the ascidian Phallusia nigra (Ascidiidae, Phlebobranchia). Zool. Sci. 18: 309-314.
Holland, L. Z. and Holland, N. D. 2001. Evolution of neural crest and placodes: amphioxus as a model for the ancestral vertebrate? J. Anat. 199: 85-98.
Hyslop, L. A., Carroll, M., Nixon, V. L., McDougall, A. and Jones, K. T. 2001. Simultaneous measurement of intracellular nitric oxide and free calcium levels in chordate eggs demonstrates that nitric oxide has no role at fertilization. Dev. Biol. 234: 216-230.
Jantzen, T. M., de Nys, R. and Havenhand, J. N. 2001. Fertilization success and the effects of sperm chemoattractants on effective egg size in marine invertebrates [Ciona intestinalis]. Mar. Biol. 138: 1153-1161.
Juge, M., Grimaud, N., Biard, J. F., Sauviat, M. P., Nabil, M., Verbist, J. F. and Petit, J. Y. 2001. Cardiovascular effects of lepadiformine, an alkaloid isolated from the ascidians Clavelina lepadiformis (Müller) and C. moluccensis (Sluiter). Toxicon 39: 1231-1237.
Kathiresan, K. and Bingham, B. L. 2001. Biology of mangroves and mangrove ecosystems. Adv. Mar. Biol. 40: 81-251.
Kim, G. J. and Nishida, H. 2001. Role of the FGF and MEK signaling pathway in the ascidian embryo. Dev. Growth Differ. 43: 521-533.
Kimura, Y., Yoshida, M. and Morisawa, M. 2001. Participation of neurotransmitters and adrenergic receptor in the metamorphosis of ascidian larvae. In: Sawada, H., Yokosawa, H. and Lambert, C. C. (ed.), The Biology of Ascidian. Tokyo, Springer-Verlag, pp. 241-245.
Kobayashi, K. and Nishida, H. 2001. Nuclear plasticity and timing mechanisms of the initiation of alkaline phosphatase expression in cytoplasm-transferred blastomeres of ascidians. Dev. Biol. 234: 510-520.
Kodama, E., Baba, T., Yokosawa, H. and Sawada, H. 2001. cDNA cloning and functional analysis of ascidian sperm proacrosin. J. Biol. Chem. 276: 24594-24600.
Kott, P. 2001. The Australian Ascidiacea part 4, Aplousobranchia (3) Didemnidae. Mem. Queensland Mus. 47: 1-407.
Kusakabe, T., Kusakabe, R., Kawakami, I., Satou, Y., Satoh, N. and Tsuda, M. 2001. Ci-opsin1, a vertebrate-type opsin gene, expressed in the larval ocellus of the ascidian Ciona intestinalis. FEBS Lett. 506: 69-72.
Lacalli, T. C. 2001. New perspectives on the evolution of protochordate sensory and locomotory systems, and the origin of brains and heads. Phil. Trans. Roy. Soc. Lond. B 356: 1565-1572.
Lambert , C. C. 2001. Fertilization-induced glycosidase release and interspecific sperm competition in ascidians. In: Sawada, H., Yokosawa, H. and Lambert, C. C. (ed.), The Biology of Ascidians. Tokyo, Springer-Verlag, pp. 24-29.
Lambert, G. 2001. A global overview of ascidian introductions and their possible impact on the endemic fauna. In: Sawada, H., Yokosawa, H. and Lambert , C. C. (ed.), The Biology of Ascidians. Tokyo, Springer-Verlag, pp. 249-257.
Lambert, G. and Sanamyan, K. 2001. Distaplia alaskensis sp. nov. (Ascidiacea, Aplousobranchia) and other new ascidian records from south-central Alaska, with a redescription of Ascidia columbiana (Huntsman, 1912). Can. J. Zool. 79: 1766-1781.
Lane, N. J., Manni, L., Burighel, P. and Zaniolo, G. 2001. Ascidian brain originates from the neural gland primordium. In: Goos, H. J. T., Rastogi, R. K., Vaudry, H. and Pierantoni, R. (ed.), Perspective in Comparative Endocrinology: Unity and Diversity--proceedings of the meeting May 26-30, 2001 Sorrento (Napoli), Italy. Monduzzi Editore, pp. 239-244.
Lee, I. H., Lee, Y. S., Kim, C. H., Kim, C. R., Hong, T., Menzel, L., Boo, L. M., Pohl, J., Sherman, M. A., Waring, A. and Lehrer, R. I. 2001. Dicynthaurin: an antimicrobial peptide from hemocytes of the solitary tunicate, Halocynthia aurantium. Biochim. Biophys. Acta 1527: 141-148.
Lopez-Gonzalez, P. J., Bresciani, J. and Conradi, M. 1998. Tarificola bulbosus, new genus, new species, a highly transformed parasitic copepod, with information on its parasitism and larval development. J. Crust. Biol. 18: 581-589.
Makabe, K. W., Kawashima, T., Kawashima, S., Minokawa, T., Adachi, A. and al., e. 2001. Large-scale cDNA analysis of the maternal genetic information in the egg of Halocynthia roretzi for a gene expression catalog of ascidian development. Development 128: 2555-2567.
Matsumoto, J., Nakamoto, C., Fujiwara, S., Yubisui, T. and Kawamura, K. 2001. A novel C-type lectin regulating cell growth, cell adhesion and cell differentiation of the multipotent epithelium in budding tunicates. Development 128: 3339-3347.
Mazouni, N., Gaertner, J.-C. and Deslous-Paoli, J.-M. 2001. Composition of biofouling communities on suspended oyster cultures: an in situ study of their interactions with the water column. Mar. Ecol. Prog. Ser. 214: 93-102.
McCoy, M. C. and Faulkner, D. J. 2001. Uoamines A and B, piperidine alkaloids from the ascidian Aplidium uouo. J. Nat. Prod. 64: 1087-1089.
McHenry, M. J. 2001. Mechanisms of helical swimming: asymmetries in the morphology, movement and mechanics of larvae of the ascidian Distaplia occidentalis. J. Exp. Biol. 204: 2959-2973.
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