Epitoky in Nereis (Neanthes) virens (Polychaeta: Nereididae): a story about sex and death

The nereidid Nereis (Neanthes) virens undergoes drastic behavioural, morphological and physiological changes during its sexual maturation (epitoky). This metamorphosis prepares benthic worms for a brief pelagic existence devoted to mating although in N. virens only mature males leave their burrows to swarm. After spawning, individuals of both sexes die. Specific adjustments of energy metabolism pathway allowing higher muscular activity and swimming capacity remain to be eluded. This study compared atokous worms (immature) and epitokous (mature) swimming males and benthic females of N. virens to detect metabolic changes that could occur during epitoky. Epitokous males showed significantly higher electron transport system, citrate synthase and aspartate aminotransferase activities (p<0.01) and significantly lower lactate dehydrogenase activity (p<0.01) compared to atokous worms and epitokous females. There was no difference in antioxidant enzyme capacities between epitokes and atokes. Lipase and trypsin activities were significantly lower (p<0.01) in epitokous males. The enzymatic changes observed are likely related to the metabolic adjustments required to support higher swimming abilities. Maintenance of antioxidant capacities could be related to protection of germinal tissues more than long term survival, since N. virens die after spawning.     

Gametogenesis in the cirratulid polychaetes Dodecaceria concharum and D. caulleryi

In Northumberland Dodecaceria concharum coexists in the same habitat as D. caulleryi . Morphologically both species are very similar but reproductively they are not. D. concharum is a parthenogenetic atoke while D. caulleryi has an asexual cycle that ends in an epitokous sexual phase. Histologically the gonad of both species had six cell types and the number of cells in each type were found during gametogenesis. In D. concharum the period over which gametogenesis occurs was followed from the increase in size of free coelomic oocytes and numbers of body segments. In D. caulleryi the period was followed by means of changes in the asexual cycle and metamorphosis into epitokes. In spite of the differences in the methods of reproduction of the two species gametogenesis of each was found to be almost identical. Only oogenesis could be compared since D. concharum does not have males. Prior to spawning in both species the production of oocytes by the ovary appeared to be inhibited. Possible hormonal mechanisms used to control atokous and epitokous spawning are discussed. From the histology of the gonads and oocytes D. concharum appeared to have half the number of chromosomes of D. caulleryi.     

The presence of the epitokous type of Perinereis cultrifera (Polychaeta) in the Mediterranean

The reproduction of Nereid Perinereis cultrifera in the English Channel and the Atlantic is of an epitokous type. The same Nereid in the Mediterranean (Bay of Alger) has been described as atokous (Marcel, 1962) and since that time epitoky in mediterranean specimens has been considered to be an accidental phenomenon. However, in the present study polychaetes collected at Salammb? (near Tunis) show that those P. cultrifera reproduce exclusively by epitoky.     

Zur Determination der zeitlichen Verteilung von Fortpflanzungsprozessen in Laborkulturen des PolychaetenTyposyllis prolifera

Factors controlling the timing of reproduction in laboratory cultures of the polychaete Trposyllis prolifera.Typosyllis prolifera (Krohn) from Pore (Yugoslavia) has been cultured for 12 successive generations. The life cycle of the species in the laboratory is described briefly. During their life individuals reproduce several times (up to 15) by stolonization which, under constant laboratory conditions (LD 16:8, 20 °C), is cyclic and takes place about every 30 days. Based on the investigations of Durchon (1959) and Wissocq (1966), experiments on extirpation and transplantation of the proventriculus have been carried out. The results suggest that an endocrine system anatomically connected to the proventriculus is important in the control of reproduction. Most likely, the endogenous reproductive cycle of an individual is controlled by periodical changes of the activity of this system. During the period following stolonization, the endocrine system of the proventriculus, which at this time shows its maximal activity, inhibits sexual development and enables regeneration of the segments lost as stolon. A subsequent decrease of the hormonal activity induces sexual maturation and epitokous metamorphosis, thus leading to further stolonization. Exogenous factors influencing the timing of reproduction probably affect the endocrine function of the proventriculus. Short-day photoperiods (LD 10:14) and low temperatures (12 °C) given simultaneously (i.e. winter conditions) totally suppress reproduction. Under normally favourable conditions (LD 16:8, 20 °C), reproductive processes can be prevented by starving or amputation of caudal segments. In all these cases, however, stolonization can be induced by removing the proventriculus. Exogenous factors also play a decisive role in synchronizing reproductive events within the species population. Under field conditions reproduction shows a lunar periodicity. The endogenous reproductive cycles of cultured specimens can be synchronized by a periodical light regime simulating the change of the moon.     

A review of Asian Hediste species complex (Nereididae, Polychaeta) with descriptions of two new species and a redescription of Hediste japonica (Izuka, 1908)

Three Asian species of the genus Hediste (Nereididae, Polychaeta, Annelida), including two new species, are reviewed. Hediste japonica (Izuka, 1908) was redescribed based on recently discovered type material and on additional specimens newly collected from the Ariake Sea in Japan and the west coast of Korea. Two new species, H .  diadroma and H. atoka , were described based on specimens collected from Japan. These three species are morphologically similar, but distinguishable by some differences in parapodial and chaetal morphology, at least at the sexually mature stage. The presence of homogomph falcigers and the absence of heterogomph spinigers are unique characteristics of H. japonica among this genus. Inconspicuous epitokous metamorphosis occurred as a slight enlargement of the eyes and parapodial ligules in mature adults of H. diadroma sp. nov. and H. japonica in combination with their reproductive swarming and pelagic larval development. The epitoke-specific sesquigomph spinigers were added in mature adults of H. diadroma sp. nov., while the number of ordinary homogomph spinigers increased in some adults of H. japonica. Epitokous metamorphosis in the Hediste species was compared with that in other nereidid genera. No epitokous metamorphosis was observed in H. atoka sp. nov., corresponding to its reproduction without swarming and to its development without a planktonic larval stage. Significant differences in paragnath numbers in the maxillary ring of the proboscis were found among the three species, suggesting different feeding habits. In Japan, the present distribution of H. japonica is restricted to muddy tidal flats in the inner part of the Ariake Sea, probably due to recent human impact, while the other two species are widespread.  © 2003 The Linnean Society of London. Zoological Journal of the Linnean Society, 2003, 137, 403−445     

COMPARATIVE STUDIES ON THE CELL WALLS OF SEXUAL AND ASEXUAL BANGIA ATROPURPUREA (RHODOPHYTA). II ELECTROPHORETIC PATTERNS OF POLYSACCHARIDES1

Cellulose acetate electrophoresis of the hot water soluble polysaccharide extracts from whole filaments, as well as base, mid and tip segments, of marine asexual and sexual Bangia atropurpurea (Roth) C. Ag. Yielded distinctive patterns which demonstrated that changes occur in the outer cell walls during sexual reproduction. Heterogeneity of the sulfated polysaccharide components isolated from outer cell walls was shown to be specifically related to sexual reproduction. Two components (Band I and II) were detected in extracts from tips of sexual filaments, whole only one (Band I) was present in the vegetative segments of all filaments and in asexual reproductive regions. The faster running component (Band II) was detected during the later stages of sexual development, prior to maturation.     

Genetic studies on the role of octopine T-DNA border regions in crown gall tumor formation

Summary Crown gall tumors result from transfer and integration of the T-DNA from the Ti plasmid of Agrobacterium tumefaciens into plant nuclear DNA. In the present study, recombinant plasmids containing deletion and rearrangement deriviatives of the T-DNA region of the octopine Ti plasmid pTiA6 were tested in a binary tumorigenesis system (Hoekema et al. 1983) to determine the requirements for T-DNA border regions in tumor formation. Since two defined segments of the T-DNA region of octopine Ti plasmids can be detected in tumor DNA (the left (T_L-) and right (T_R-) DNA), four border regions exist in this Ti plasmid. Agrobacteria harboring plasmid constructs which contain a T-DNA gene capable of inciting tumors (gene 4, the tmr gene, which is involved in cytokinin biosynthesis) and various T-DNA border regions were tested for ability to cause tumors on Nicotiana glauca and other host plants. Such tmr constructs containing as their only border region the right border of either the T_L-DNA or the T_R-DNA are fully tumorigenic. Analogous tmr constructs containing only the T_L-DNa left border region are not tumorigenic. These results do not depend on the orientation or position of the single border with respect to the tmr gene; furthermore, the T_R-DNA right border can confer tumor-forming ability despite the presence of an intervening copy of the T_L-DNA left border.These results for relatively small plasmids are contrasted with previously determined requirements for border regions in tumorigenesis by intact Ti plasmids. A model previously proposed by Wang et al. (1984) for the role of border regions in DNA transfer to plant cells is extended in order to explain the tumor-forming ability of plasmid constructs containing a single border region. The results of this study interpreted according to the model suggest that the octopine T_L-DNA left border is defective in this DNA-transfer process.     

Comparative sequence analysis of a single-gene conserved segment in mouse and human

Comparative mapping and sequencing of the mouse and human genomes have defined large, conserved chromosomal segments in which gene content and order are highly conserved. These regions span megabase-sized intervals and together comprise the vast majority of both genomes. However, the evolutionary relationships among the small remaining portions of these genomes are not as well characterized. Here we describe the sequencing and annotation of a 341-kb region of mouse Chr 2 containing nine genes, including biliverdin reductase A (Blvra), and its comparison with the orthologous regions of the human and rat genomes. These analyses reveal that the known conserved synteny between mouse Chromosome (Chr) 2 and human Chr 7 reflects an interval containing one gene (Blvra/BLVRA) that is, at most, just 34 kb in the mouse genome. In the mouse, this segment is flanked proximally by genes orthologous to human chromosome 15q21 and distally by genes orthologous to human Chr 2q11. The observed differences between the human and mouse genomes likely resulted from one or more rearrangements in the rodent lineage. In addition to the resulting changes in gene order and location, these rearrangements also appear to have included genomic deletions that led to the loss of at least one gene in the rodent lineage. Finally, we also have identified a recent mouse-specific segmental duplication. These finding illustrate that small genomic regions outside the large mouse–human conserved segments can contain a single gene as well as sequences that are apparently unique to one genome. The nucleotide sequence data reported in this paper have been submitted to GenBank and assigned the accession numbers AC074224 and AC074041.     

Genetic compatibility between sexual and clonal genomes in local populations of the hybridogeneticRana esculenta complex

Summary Hybridogenetic species possess a hybrid genome: half is clonally inherited (hemiclonal reproduction) while the other half is obtained each generation by sexual reproduction with a parental species. We addressed the question of whether different hemiclones of the hybridogenetic water frogRana esculenta are locally adapted for genetic compatibility with their sexual parental hostRana lessonae. We artificially crossedR. esculenta females of three hemiclones (GUT1, GUT2 and GUT3) from a pond near Gütighausen, Switzerland and one hemiclone (HEL1) from near Hellberg, Switzerland each toR. lessonae males from both populations. We also created primary hybrids by crossing the sameR. lessonae males from both populations toR. ridibunda females from Pozna, Poland (POZ). Tadpoles were then reared in the laboratory at two food levels to assess their performance related to early larval growth rate, body size at metamorphosis and length of the larval period. Tadpoles from hemiclones GUT1, GUT3 and POZ had higher growth rates than those from hemiclones GUT2 and HEL1 at the low food level, but at the high food level all growth rates were higher and diverged significantly between hemiclones GUT2 and HEL1. Tadpoles from the intrapopulational crosses GUT2 × GUT and HEL1 × HEL were larger at metamorphosis than those from the interpopulational crosses GUT2 × HEL and HEL1 × GUT. A high food level increased the size at metamorphosis in all tadpoles. A high food level also decreased the days to metamorphosis and tadpoles from GUT1, GUT3 and POZ had the shortest larval period whereas those from GUT2 and HEL1 had the longest. These results indicate that the differential compatibility of clonal genomes may play an important role in hybridogenetic species successfully using locally adapted sexual genomes of parental species and that interclonal selection is likely important in determining the distribution of hemiclones among local populations.     

Ultrastructure of the Genital Organs in the Interstitial Syllid Petitia amphophthalma (Annelida,Polychaeta)

Abstract The gonochoristic syllid Petitia amphophthalma is one of the truly interstitial polychaetes. P. amphophthalma does not show any epitokous modifications at maturity such as those that usually occur in syllids. The reproductive structures are unique: the male genital organs consist of a seminal vesicle in chaetigers 6–10, subdivided into a dorsal part tightly filled with spermatozoa and a ventral part with contents in different stages of spermatogenesis, one pair of sperm ducts and conspicuous gland cells situated in chaetigers 10 and 11. Their glandular secretions are discharged into the sperm duct together with those of other types of gland cells that form the duct. The oocytes develop freely within the body cavity of the females. Each of the fertile segments possesses a paired oviduct ending in a large ciliated funnel. Sperm ducts and oviducts are probably modifications of excretory organs; nephridia are absent in segments where gonoducts occur. A direct sperm transfer by lytic opening of the integument of the female and internal fertilization are inferred. Copyright © 1996 Published by Elsevier Science Ltd on behalf of the Royal Swedish Academy of Sciences     

Abnormal-abdomen beiDrosophila hydei

Several strains ofDrosophila hydei exhibiting an abnormal-abdomen (aa) phenotype were investigated. They proved to be mutant at one and the same major gene, located on chromosome 2 (homologous to 3R ofD. melanogaster).This mutant gene,aa, causes skeletal abnormalities of the adult abdomen. Sternites and, less frequently, tergites are imperfectly sclerotized. They may be dissolved into separate areas, or may be missing altogether. Often they are normal in shape, while the number of hairs is reduced. Compound sclerites originating from parts of adjoining segments were never observed. Small hypoderm spherules with internal hairs occur frequently under the ventral skin. In aa larvae, segmentation is normal, in contrast to the situation in abnormal-abdomen mutants ofD. melanogaster. Even where theaa mutant expression is extremely strong, puparia are perfectly normal. Theaa gene thus appears to act exclusively during metamorphosis. This was confirmed by temperature experiments. The expression of theaa gene is much reduced at low temperature, but low temperature is effective during metamorphosis only.The seven aa strains compared, showed considerable differences in penetrance and expressivity as well as in the amount of dominance in crosses. Reciprocal crosses yield in the F₁ values either intermediate, or lower or higher, compared to those of the parent strains. The F₁ values show a nonlinear correlation between penetrance and expressivity.On the basis of chromosomal location and phenotypic effect, it is unlikely thataa ofD. hydei is homologous to any of the abnormal-abdomen mutants ofD. melanogaster described so far. However, a new dominant autosomal mutant was found inD. simulans, which seems to be a homologue.     

Untersuchungen am pazifischen PalolowurmEunice viridis (Polychaeta) in Samoa

This paper deals with the problem of lunar-periodic reproduction in the palolowormEunice viridis Gray; in Samoa this famous event takes place regularly once a year in October or November while the moon is in its last quarter. In 1966 most of the epitokous posterior ends, the so-called “palolos”, spawned on the 5th of November. Between the 14th of June and the 20th of November 1966 we isolated continuously complete paloloworms from the coral reef at Faga'itua (Tutuila/American Samoa) for an examination of their sexual status. During this whole period we found not only various stages of ripening (respectively mature) specimens but also other worms of the same size the posterior ends of which contained either immature or no germ-cells at all. Apparently, the sexual development of the population as a whole is not synchronized all year round. Obviously, external factors interfere acting only on certain specimens as promoter for sexual maturation and as “Zeitgeber” for simultaneous breeding. Those paloloworms, which lose their epitokous part at this time, subsequently start to regenerate a new posterior end. We kept many of the collected paloloworms alive for several weeks or even months in our Samoan laboratory. Females, which had been isolated from the reef on the 22nd of October or later, were the only ones which continued their sexual development in the laboratory, whereas among the males also some specimens which had been collected 1 to 2 months earlier became sexually mature. In all paloloworms taken from the reef before the 15th of August and kept thereafter in the laboratory, the sexual development was totally inhibited. The same block appeared in all headless posterior fragments of male or female paloloworms even if they had been isolated on the 22nd of October in a nearly mature status; only 3 of such fragments performed their sexual maturation after regeneration of a new head. This result and other facts suggest the existence of a hormone in the head which induces sexual maturation in the posterior end of the worm. Histological investigations agree with this assumption so far as they have shown neurosecretion in the brain of the paloloworm; those maturing animals which had been fixed one month before spawning included about 10 times more neurosecretory material in their brain than specimens collected earlier.     

Postmetamorphic growth,survival, and age at first reproduction of the salamander,hynobius nebulosus tokyoensis Tago in relation to a consideration on the optimal timing of first reproduction

The postmetamorphic growth and survival of the salamander Hynobius nebulosus tokyoentisTago were surveyed in the study site located in Habu village of Hinodemachi, a suburb of Tokyo City, during 1975–1981. A laboratory experiment on the growth rate of juveniles was conducted in parallel with the field survey. The result indicated that this salamander grew at the rate of 8,mm in s.v.l. per year during the juvenile stage, but its growth rate decreased markedly as low as 1.8 mm for males and 1.1 mm for females, once it had attained sexual maturity. According to the “capture-recapture” procedure the annual survival rate after metamorphosis was found to be quite high; that is, approximately 0.7. By using the growth rate of juveniles and the difference between the sizes at metamorphosis and sexual maturity, the age at first reproduction was estimated to be 4 year for males and 5 year for females. From the data obtained in this study, the intrinsic rates of increase (r) were calculated for various values of age at first reproduction under different survival schedules, and the relationship between the age at first reproduction and fitness as measured by r was examined. The result indicated that an optimal age maximizing fitness always existed under respective survival schedules, and the observed age at first reproduction of this salamandei was found to coincide well with the predicted optimal age.     

Cladoendesis and a new look at the evolution of insect metamorphosis

Cladoendesis is a method of phylogeny analysis opposed to various matrix methods. In contrast to matrix methods, in cladoendesis phylogenetic trees are not built each time as new ones, but are reconstructed based on the previous results. Each character of each taxon is compared with its ancestral condition in the ground-plan of the higher taxon. The revealed part of the phylogeny is represented in a form of hierarchical classification. In addition to the principles of natural classification elaborated by C. Linnaeus, P.-A. Latreille, and others, evolutionary theory elaborated by Ch. Darwin and others, and principles of phylogenetic systematics elaborated by W. Hennig and others, cladoendesis includes rank-free dual nomenclature system (DualNom) and advanced method of text layout elaborated by the author. Important components of cladoendesis are the use of the term “plesiomorphon” and taking into account such important evolutionary events as conservation and deconservation of characters. Cladoendesis allowed the author to make comprehensive revision of the phylogeny of Ephemeroptera, find homology in the structure of the maxillae of adult Aphaniptera, larval maxillae of Neuropteroidea-Birostrata, larval legs of Nannomecoptera, and genital parts of some insect taxa. It also allows understanding the nature and evolution of insect metamorphosis. It is generally believed that initially insect ontogenesis proceeds the shortest way, while such phenomena as substitutions of organs by means of their atrophy and subsequent restoring are secondary. In contrast to this, the flagellum of the antenna of Amyocerata initially grows by addition of proximal segments and simultaneous loss of distal ones; in many taxa, including Metabola (insects with complete metamorphosis), distal segments are not aborted. Some authors tried to explain the origin of complete metamorphosis by various reasons: ecological, morphogenetic, or others. This approach is wrong because all insects with complete metamorphosis constitute the holophyletic taxon Metabola Burmeister 1832, i.e., originate from a single ancestral species which acquired this kind of metamorphosis. If complete metamorphosis could appear in response to some factor, it would appear many times in different species, in which case the taxon Metabola, characterized by complete metamorphosis, would be polyphyletic. The holophyly of Metabola is well proven by cladoendesis but cannot be revealed by any matrix method. Based on understanding of these facts, the author was able to discover the specific features which appeared in the common ancestor of Metabola and then became conserved and were inherited by all its descendants. These features include loss of the scape in the larval antenna (leading to a peculiar transformation of antennae in metamorphosis) and a peculiar mode of leg transformation during molt from larva to pupa. During the larval/pupal molt, the leg loses musculature and gets an immobile knee bend, so that the pupa is unable to use its legs. This conserved feature determines the inactive mode of life of the pupae of most insects. It is usually believed that male coccids (Gallinsecta De Geer 1776) have metamorphosis similar to the complete metamorphosis of Metabola. But the phylogenetic position of Gallinsecta and Metabola, as revealed by cladoendesis, does not allow one to assume common modifications in their metamorphoses. Now, when concrete autapomorphies of Metabola have been found, it becomes possible to compare metamorphoses of Metabola and Gallinsecta. Examination of Orthezia urticae and some other coccids shows that metamorphosis of their males shares no modifications with the true complete metamorphosis. The inactive mode of life of their nymphs is not connected with any anatomical reason, but purely with the fact that in the course of transformation from the feeding wingless larva to the non-feeding winged adult, the nymph has already lost its mouth apparatus and has not yet got functional wings, so it does not need to move. At the same time, actively feeding stages of Gallinsecta, both males and females, have an unusual mode of molt transformation of the legs and antennae; this is a unique autapomorphy of Gallinsecta not found in any other insect taxon.     

Developmental fates of larval tissues after metamorphosis in the ascidian, Halocynthia roretzi

Cell lineages during ascidian embryogenesis are invariant. Developmental fates of larval mesodermal cells after metamorphosis are also invariant with regard to cell type of descendants. The present study traced developmental fates of larval endodermal cells after metamorphosis in Halocynthia roretzi by labeling each endodermal precursor blastomere of larval endoderm. Larval endodermal cells gave rise to various endodermal organs of juveniles: endostyle, branchial sac, peribranchial epithelium, digestive organs, peripharyngeal band, and dorsal tubercle. The boundaries between clones descended from early blastomeres did not correspond to the boundaries between adult endodermal organs. Although there is a regular projection from cleavage stage and larval stage to juvenile stage, this varies to some extent between individuals. This indicates that ascidian development is not entirely deterministic. We composed a fate map of adult endodermal organs in larval endoderm based on a statistical analysis of many individual cases. Interestingly, the topographic position of each prospective region in the fate map was similar to that of the adult organ, indicating that marked rearrangement of the positions of endodermal cells does not occur during metamorphosis. These findings suggest that fate specification in endoderm cells during metamorphosis is likely to be a position-dependent rather than a deterministic and lineage-based process. Received: 16 June 1999 / Accepted: 16 August 1999     

The function ofbithorax genes in the abdominal central nervous system ofDrosophila

Summary We have analysed the influence of the bithorax gene complex (BX-C) on two segment-specific features of the central nervous system ofDrosophila larvae: the “presumptive leg neuromeres” (PLN), which are present only in the thoracic ganglia of the larva and develop into the leg neuromeres of the adult fly during metamorphosis; and the “lateral dots” (LD) which are found in the first abdominal as well as thoracic ganglia. We show in both cases that consecutive BX-C genes can suppress the development of these structures. We also show that each gene is expressed in several consecutive segments, leading to an apparent redundancy of the suppression in posterior segments.     

Isotopic fractionation with morphological change and sexual specificity in the lappet moth Euthrix potatoria

The δ¹⁵N values of adult holometabolous insects exceed those of larvae, but otherwise little information on terrestrial invertebrates has been obtained in food‐web analyses using stable isotope ratios (δ¹⁵N, δ¹³C). Changes in δ¹³C during metamorphosis and differences between males and females have not been examined. We collected the larvae and cocoons of Euthrix potatoria (L.) (Lepidoptera: Lasiocampidae) in the field and used them to assess the species’ isotopic fractionation. Each emerged moth was divided into five body parts. We conducted stable N and C isotope analyses for each body part, as well as for cocoons and exuviae, and also compared stable isotope ratios between sexes. We confirmed δ¹⁵N enrichment through metamorphosis and estimated that δ¹⁵N enrichment is accomplished by the relative concentration of ¹⁵N due to the excretion of copious meconium, which contains abundant ¹⁴N. We also observed changes in δ¹³C values through metamorphosis. Both isotope values tended to change more in males than in females. The proportion of the whole‐adult weight represented by meconium was higher in males than in females, suggesting that high meconium secretion in males contributes to the sexual difference in δ¹⁵N. These phenomena may be common in Holometabola, which require a pupal stage. For more accurate food‐web assessments, it is important to consider stable isotope changes during different life cycles, as well as sexual differences.     

The ultrastructure of odontogenesis in larval and adult urodeles; differentiation of the dental epithelial cells

Summary Sections of undemineralized tooth germs ofAmbystoma andTriturus were examined. The ultrastructure of early germs, both larval and adult, and of dentinogenesis, resembled that of mammals. In adult bicuspid teeth, once the dentine of the cusps was mineralized, mineral crystals of a similar size to early mammalian enamel crystals, appeared between the dentine and the inner dental epithelium (i.d.e). Concomitantly, the i.d.e showed features of mammalian secreting ameloblasts. This new layer, regarded as true enamel, lacked collagen, possessed an ordered arrangement of crystals and reached a maximum thickness of 6 m.In larval monocuspid teeth, once dentine mineralization had reached the plasma membranes of the i.d.e at the tip of the cusp, the i.d.e developed a ruffled border. At this stage the dentine of the tip, regarded as enameloid, was very hard and difficult to section. The ruffled border, characteristic of other cells which transport materials, was regarded as indicating that the i.d.e was removing organic matter from the enameloid. The differences in development between larval and adult teeth support the concept that there is a change in cellular activity of the i.d.e which occurs during metamorphosis from the larval to the adult urodele.