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Notes: Abstract  In amphioxus embryos, the nascent and early mesoderm (including chorda-mesoderm) was visualized by expression of a Brachyury gene (AmBra-2). A band of mesoderm is first detected encircling the earliest (vegetal plate stage) gastrula sub-equatorially. Soon thereafter, the vegetal plate invaginates, resulting in a cap-shaped gastrula with the mesoderm localized at the blastoporal lip and completely encircling the blastopore. As the gastrula stage progresses, DiI (a vital dye) labeling demonstrates that the entire mesoderm is internalized by a slight involution of the epiblast into the hypoblast all around the perimeter of the blastopore. Subsequently, during the early neurula stage, the internalized mesoderm undergoes anterior extension mid-dorsally (as notochord) and dorsolaterally (in paraxial regions where segments will later form). By the late neurula stage, AmBra-2 is no longer transcribed throughout the mesoderm as a whole; instead, expression is detectable only in the posterior mesoderm and in the notochord, but not in paraxial mesoderm where definitive somites have formed.

Notes: Abstract  In vertebrates, the orphan nuclear receptors of the COUP-TF group function as negative transcriptional regulators that inhibit the hormonal induction of target genes mediated by classical members of the nuclear hormone superfamily, such as the retinoic acid receptors (RARs) or the thyroid hormone receptors (TRs). To investigate the evolutionary conservation of the roles of COUP-TF receptors as negative regulators in the retinoid and thyroid hormone pathways, we have characterized AmphiCOUP-TF, the homologue of COUP-TFI and COUP-TFII, in the chordate amphioxus (Branchiostoma floridae), the closest living invertebrate relative of the vertebrates. Electrophoretic mobility shift assays (EMSA) showed that AmphiCOUP-TF binds to a wide variety of response elements, as do its vertebrate homologues. Furthermore, AmphiCOUP-TF is a transcriptional repressor that strongly inhibits retinoic acid-mediated transactivation. In situ hybridizations revealed expression of AmphiCOUP-TF in the nerve cord of late larvae, in a region corresponding to hindbrain and probably anterior spinal cord. Although the amphioxus nerve cord appears unsegmented at the gross anatomical level, this pattern reflects segmentation at the cellular level with stripes of expressing cells occurring adjacent to the ends and the centers of each myotomal segment, which may include visceral motor neurons and somatic motor neurons respectively, among other cells. A comparison of the expression pattern of AmphiCOUP-TF with those of its vertebrate homologues, suggests that the roles of COUP-TF in patterning of the nerve cord evolved prior to the split between the amphioxus and vertebrate lineages. Furthermore, in vitro data also suggest that Amphi-COUP-TF acts as a negative regulator of signalling by other nuclear receptors such as RAR, TR or ER.

Notes: Abstract  A full-length Wnt1 gene (AmphiWnt1) was isolated from amphioxus. Expression is first detectable in the gastrula around the lip of the blastopore. By the early neurula, transcription is in the mesendoderm near the closed blastopore, but is down-regulated in the overlying ectoderm. In the late neurula, expression is limited to the posterior wall of the neurenteric canal. Later in development, AmphiWnt1 transcripts can no longer be detected. AmphiWnt1 has no counterpart of the predominant expression domains of vertebrate Wnt1 genes in the neural tube, but its expression may be more comparable to that of wingless in the invaginating hindgut primordium of insects.

Notes: Abstract Colonies ofPyrosoma atlanticum were collected by submersible in October 1988 in the Caribbean Sea, and testes were studied by electron microscopy. Spermatogonia, spermatocytes and early spermatids have two centrioles. The proximal centriole subsequently disappears, its remains apparently persisting in the spermatozoon as dense material adjacent to the distal centriole, which gives rise to the axoneme. At the tip of early spermatids are several 50 nm proacrosomal vesicles, which disappear leaving no trace in early elongating spermatids. The spermatozoon lacks an acrosome and has a head 35µm long. The head is differentiated into a bulbous posterior portion 5µm long × 1µm wide, a thinner anterior portion 25µm long tapering from a width of 0.7µm to a width of 0.4µm, and a very thin anterior extension 5µm long × 0.5µm wide. At the start of elongation, the anterior extension begins to form just lateral to the proacrosomal vesicles as a spiral projection comprising part of the nucleus, covered by a thin sheath of cytoplasm. This sheath of cytoplasm undergoes a complex differentiation. Ultimately, the nucleus in the anterior extension is overlain by two membrane-bound sheaths of cytoplasm connected by a spiral flange of cytoplasm. Between these two sheaths is a spiral space, open to the exterior through a subterminal pore near the sperm tip. In early spermatids the mitochondria fuse into a single mitochondrion, which remains lateral to the nucleus. The cristae become modified late in spermatogenesis. Throughout elongation of the spermatid there are patches of dense material between the nucleus and mitochondrion. A manchette of microtubules transiently encircles the thin anterior portion of the nucleus during the last phase of elongation. A manchette is not present during most of elongation. In the spermatozoon the mitochondrion, which has reticulate cristae, spirals a few times about the nucleus and extends from the junction between the bulbous portion and the thinner anterior portion of the nucleus to the junction between the thinner anterior portion and the nuclear extension. Spermatogenesis inP. atlanticum, compared to that in other tunicates, most closely resembles that in colonial ascidians, and supports the majority view that pyrosomes arose from aplousobranch ascidians that lost their attachment to the substratum. Pyrosome sperm are more highly derived than doliolid sperm, which have an acrosome that is probably capable of exocytosis. When salp and pyrosome sperm are compared, both are highly derived, but neither shares any apomorphies with the other that it does not share with at least one other tunicate order. Thus, sperm morphology does not support the majority view that pyrosomes gave rise to doliolids and neither confirms nor denies the idea that pyrosomes are intermediate between aplousobranch ascidians and salps. Therefore, it is likely that the class Thaliacea is polyphyletic, with doliolids arising very early from the ascidian lineage and with salps and pyrosomes arising somewhat later.

Notes: Abstract In Dolioletta gegenbauri and Doliolum nationalis, collected in 1987 in the bay of Villefranche-sur-mer (French Mediterranean Sea), spermiogenesis is essentially the same. Early spermatids have a round head, a flagellum arising from a single centriole with short microtubules at 45° to its base, several mitochondria, and an acrosome 50 nm thick and 250 nm long with its long axis parallel to the plasma membrane. The acrosomal contents are dense, with a central denser plate. The nuclear envelope next to the acrosome is thickened and concave. In elongating nuclei, strands of chromatin become oriented parallel to the length of the nucleus and then twist helically. Although the mitochondria surround the nucleus, they remain relatively short and do not fuse into a single mitochondrion as in sperm of other tunicates. In very late doliolid spermatids, the acrosome undergoes exocytosis, and exposes fibrous material that stays associated with the tip of the sperm; no acrosomal tubule forms. Exocytosis at this stage may be triggered by fixation. If so, exocytosis probably occurs naturally at some time before fusion of sperm and egg. Sperm have elongate heads (1 μm×10 μm), with the anterior two-thirds of the nucleus surrounded by mitochondria. Spermiogenesis in doliolids, compared to that in other tunicates, is most like that in solitary members of the class Ascidiacea, except that in the latter the sperm mitochondria fuse and the acrosome appears incapable of exocytosis. In contrast, previous work has shown that salps (class Thaliacea) and colonial didemnid ascidians have an acrosomeless sperm with a spiral mitochondrion, while the class Appendicularia has a sperm with a midpiece, a compact head and an acrosome capable of exocytosis and acrosomal tubule formation. By outgroup comparison with echinoderms and acraniates, appendicularian sperm are plesiomorphic within the Tunicata. Thus, gamete morphology indicates that (1) solitary ascidians and doliolids had a common ancestor, (2) the popular idea that doliolids gave rise to appendicularians is incorrect, and (3) the Thaliacea are polyphyletic, doliolids having arisen very early from the ascidian lineage and salps having arisen later.

Notes: The ovary of the salp Pegea socia (Bosc, 1802) is located at the end of an atrial diverticulum. The ovary consists of a single oocyte encased in a layer of follicle cells and is connected to the atrial epithelium by an oviduct. Transmission electron microscopy shows that the oocyte lacks a vitelline layer, cortical granules, and yolk granules and that the oviduct lacks a continuous lumen. What previous authors thought was a lumen is a line of dense intercellular junctions running down the center of the oviduct. The sperm nucleus in this species, as in other salps, is elongate. The tubular mitochondrion spirals about the sperm nucleus giving it a corkscrew-shape appearance. Sperm reach the ovary when the oocyte is still at the germinal vesicle stage. Many sperm swim up the atrial diverticulum and burrow through the cells of the atrial epithelium, oviduct, and follicular epithelium. Thus oviduct shortening, which occurs when the oocyte is in the meiotic divisions, is evidently unrelated to sperm moving up the oviduct. All previous authors, who argued either that a continuous lumen is necessary for sperm to move up the oviduct or that sperm bypass the oviduct, were incorrect. © 1994 Wiley-Liss, Inc.