Friday, May 28, 2010
Thursday, May 27, 2010
A week later I checked on the twins to see how they were developing. The juvenile urchins have grown very large and the larval body mostly degenerated. Three of the remaining larval arms are visible on the upper right in the bottom picture. Because this specimen was so thick and non-transparent it was difficult to get a clear picture in the transmitted light. This picture is taken using cross-polarized light to make the juvenile skeletal spicules glow. You can see the spines from the two different juvenile rudiments - some to the left and others to the right of the plane of bilateral symmetry of the larva (which cuts across from the upper right to the bottom left of the picture). The larger of the two rudiments in the first picture was more developed and is now mobile on its tube feet. The other rudiment has spines, but I could not see any tube feet stick out.
The case of twins in this situation intrigues me very much. I am a fraternal twin myself, which means that my twin sister came from a different fertilized egg. These urchin twins came from a single fertilized egg, but they are conjoined (share a single gut). They each have an oral side, but have no aboral side. Likely they will not survive much longer after metamorphosis (S. A. Maslakova, pers. communication).
Tuesday, May 25, 2010
The Sabellaria cementarium trochophore larvae have two large bundles of setae on the trunk. These setae are barbed and are longer than the larva itself. They are held close to the body when the larva is swimming (see another post by Kristina Sawyer), but can be fanned out when the larva stops moving. In 1984 J. Timothy Pennington and Fu-Shiang Chia observed Sabellaria cementarium larvae using their setae to prevent recognition and capture by predators, such as ctenophores (comb jellies). The barbed setae also might irritate the oral tissues of the predator and act as a deterrent. At the posterior end there is another ciliated band, which assist in swimming called the telotroch.
The spines of the sand dollar are the darker “limbs” with no circles at the tips. There are large and small spines, which are homologous to the interambulacral and ambulacral spines in sea stars. The sand dollar juveniles also have two extra large spines marking the posterior end of the animal. These extra long spines can be seen more clearly in the two dark-field photos at the bottom right portion of the juvenile urchin. The middle photo shows the spines of the sand dollar clearly. In the bottom photo, one can see black network-like pigment cells on the aboral (the opposite of oral, which is facing down) side of the juvenile.
Thursday, May 20, 2010
By using cross-polarized light (one polarizing filter placed above the specimen and one below), I was able to visualize the spicules in stark contrast. With this technique, the light that passes through the first polarizer is blocked by the second, and the only structures that remain bright are those that rotate the plane of polarized light e.g. various crystalline structures - in this case skeletal spicules, composed of calcium carbonate. This highlights the spicules on a dark background. The first two spicules in urchin larvae form at the base of the archenteron, one on each side, where the primary mesenchyme cells are concentrated. The initial spicule is tri-radiate. The three branches grow and form the postoral and antero-lateral arm rods, and the body rod of the pluteus larva.
Lebour MV. 1922. The food of plankton organisms. Journal of the Marine Biological Association of the United Kingdom. 12: 644-677.
Smidt ELB. 1951. Animal production in the Danish Waddensea. Meddelelser Kommission fra Danmarks Fiskeri- og Havundersogelser. 11 (6): 151.
Wilson DP. 1982. The larval development of three species of Magelona (Polychaeta) from localities near Plymouth. Journal of the Marine Biological Association of the United Kingdom. 62: 385-401.
These photos are of 11-day old trochophore larvae. The first one shows the ciliated band, called the prototroch, which encircles the larva just anterior to the mouth. The long bristles are called setae (or chaetae) and are characteristic of both the larvae and adults of polychaete worms. The setae serve as defense against planktonic predators (Pennington & Chia, 1984). Fanning out the setae (second picture), the larva can nearly double its diameter (140μm without the setae, and 250μm with setae spread out).
In the third photo you can also see the two reddish eyespots anterior to the prototroch. This trochophore will continue adding new segments, each segment bearing more setae. Once it has more than three setigers (segments with setae) it will find a suitable place to settle and build its sand tube. In some areas species of Sabellariaform extensive reefs, because their larvae prefer to settle on the tubes of adult worms of their species.
Kozloff, E.N. 1974. Seashore Life of the northern Pacific Coast; an illustrated guide to northern California, Oregon, Washington, and British Columbia. U of Washington P: Seattle.
Pennington, J. T., & Chia, F.-S. (1984). Morphological and Behavioral Defenses of Trochophore Larvae of Sabellaria cementarium (Polychaeta) against Four Planktonic Predators. Biological Bulletin. 167 (1), 168-175.
The next photo shows a 14-day old larva which is more advanced. It has longer antero-lateral arms, which project outwards from the anterior end and frame the mouth of the pluteus. The formation of extra arm pairs extends the length of the continuous ciliated band which surrounds the larval mouth and is used to capture microscopic food particles. The longer the ciliated band, the more efficient pluteus can feed. The small bumps at the base of the post-oral arms are the newly developing postero-dorsal arms.
Although one can barely see them, one can already distinguish the small calcareous spicules, which support the new pair of arms, using cross-polarized light. The four spicule rods supporting the antero-lateral pair of arms (towards the midline) and the longer post-oral pair of arms are clearly visible. The shorter postero-dorsal spicule is visible on the left side.
The final pair of arms to form are the pre-oral arms, which, true to their name, form just anterior to the mouth of the pluteus larva. These arms are more or less parallel to the anterolateral arms, and can be seen as small bumps between the antero-lateral arms. On this bottom picture you can also see an unpaired rudiment of the juvenile sand dollar (a bean shaped mass to the left of the larval stomach - which is a large darkish oval occupying the majority of space inside the larval body.
One of the largest noticeable differences between sea stars and sea urchins in early development is the formation of polar bodies. A polar body is a tiny sister-cell of the primary oocyte, produced during meiosis. It contains discarded DNA, and very little of anything else. Polar bodies are not usually observed in sea urchin, because meiosis is completed within the ovary, and spawned eggs have already parted with their polar bodes.
However, we were able observe polar bodies in sea stars. This is because in sea stars, sperm entry occurs before the oocytes have completed meiosis (cell division, reducing the number of chromosomes). Polar bodies form after fertilization and are trapped within the fertilization envelope. The photos here show an immature unfertilized oocyte (with a large nucleus and a nucleolus inside) and a fertilized secondary oocyte with homogenenous cytoplasm, a tight fertilization envelope around it, and one polar body (at about 5 o'clock), in the ochre sea star Pisaster ochraceous. The fertilization envelope in starfish is much closer to the surface of the egg than in sea urchins.
Wednesday, May 19, 2010
Chernyshev AV. 2003. Novy vid roda Hubrechtella (Nemertea, Anopla) i obosnovanie semeistva Hubrechtellidae. [A new species of the genus Hubrechtella (Nemertea, Anopla) from the Sea of Japan, and establishement of the family Hubrechtellidae]. In Russian. Biologiya Morya. 29(5): 368-370.
Eight days after fertilization, I looked at my Calliostoma culture again. At this point, most of the embryos were dead or abnormal. Larvae can be so temperamental! There were some larvae resting at the bottom of the dish that looked normal and moved their cilia. While under a cover slip, they used cilia on their velum to move around, and they moved FAST! Those I could observe had two eyespots and two tentacles forming in the apical area. At metamorphosis, the velum will degenerate, and the miniature snail will start crawling using its foot. This picture shows the veliger on its side, with one of the eyespots facing us. The velum is out on this picture (top). You can also see the foot (left) and the operculum (trap door) attached to it. At this time, I witnessed one of the other juveniles moving on its foot, — all I could see was the shell waddling around the slide.
Sunday, May 16, 2010
Friday, May 14, 2010
This picture shows the same larva in the same orientation, only with different parts in focus. It is easy to see the intestine (narrow tube in the posterior third of the larva) with the same red pigment as in the stomach, only darker! The intestine curves up and leads to the anus, located on the ventral side (in focus here). These larvae are able to swim and feed using a ciliated band composed of many tightly opposed epithelial cells, each with a cilium that looks like a little hair. The cilia (plural of cilium) beat continuously to create a water current away from the mouth, and locally redirect the flow toward the mouth, when they encounter food particles. One cannot distinguish individual cilia on this photo, however the portion of the ciliated band that flanks the mouth above and below is distinguishable and sharply in focus. The portion of the band that is in focus above the anus is called "postoral", and the portion of the band above the mouth is called "preoral". During development of the bipinnaria larva the preoral and postrodal portions of the ciliated band separate to form two separate loops (preoral and postoral).
Thursday, May 13, 2010
Temereva EN. 2009. New data on distribution, morphology and taxonomy of phoronid larvae (Lophophorata: Phoronida). Invertebrate Zoology 6(1): 47-64.
Tuesday, May 11, 2010
Sunday, May 9, 2010
Saturday, May 8, 2010
Due to the small size of the adults (only 3-5 mm), they utilize a different reproductive strategy than larger sea stars. Instead of investing energy in producing large numbers of small eggs to free-spawn into the water column (and allow them to develop into feeding ophiopluteus larvae - as pictured in the next post), these small animals produce only a few small eggs (about 100 μm) in each gonad, and invest energy in brooding them internally until they are large enough to crawl away. Although it is a larger investment per egg, this direct development strategy ensures that the young develop to the juvenile stage. Amphipholis squamata can brood multiple cohorts simultaneously, and in our dissections we found brooded young in multiple stages of development within the same adult brittle star.
Amphipholis squamata is also interesting because it is a simultaneous hermaphrodite and appears to be capable of self-fertilization. The grey colored adult brittle stars feed on diatoms and detritus, and can be found under small rocks on sand or gravel in intertidal zones worldwide (Kozloff 1974).
Kozloff, E.N. 1974. Seashore Life of the Northern Pacific Coast; an illustrated guide to northern California, Oregon, Washington and British Columbia. U of Washington P: Seattle.
Friday, May 7, 2010
Within about a minute of sperm contacting the egg plasma membrane, a fertilization envelope forms around the egg to prevent polyspermy (penetration by additional sperm). A hyaline layer forms directly on the surface of the egg plasma membrane to help hold the blastomeres in the dividing egg together. I had to remove the fertilization envelope and the hyaline layer in order to separate the blastomeres. To accomplish this I fertilized the eggs in filtered sea water (FSW). Within thirty seconds of fertilization I replaced the FSW with calcium-magnesium-free sea water. This prevents the fertilization envelope and hyaline layer from hardening, and allows one to remove them by sheering the eggs with a pipette. Once the eggs are denuded (their coats removed) they become very sticky, so I coated the culture dishes and glass knives used for surgery with BSA (Bovine Serum Albumin) to prevent the embryos from sticking to things and being damaged. The top two pictures show two 8-cell stage embryos of S. purpuratus - one with the fertilization envelope, and one without (denuded).