Author: Allison J. Gong

After much teasing and titillation, my urchin larvae have finally gotten down to the serious business of metamorphosis. It seems that I had jumped the gun on proclaiming them competent about a week ago, or maybe they were indeed competent and just needed to wait for some exogenous cue to commit to leaving the plankton for good. In any case, I’ve spent much of the last five days or so watching and photographing the larvae to document the progress of metamorphosis as it occurs. While I was unable to follow any individual larva through the entire process of metamorphosis, I did manage to put together a series of photographs that document the sequence of events.

To recap: A competent larva is anatomically and physiologically prepared to undergo metamorphosis. This batch of larvae reached competence at the age of about 45 days. The larva below is very dense and opaque in the main body. It can still swim, but has become “sticky” and tends to sit on the bottom of the dish.

Sometimes the first tube feet emerge from the larva while it is still planktonic. Other times the larva falls to the benthos and lands on its (usually) left side, where the rudiment is located.

This larva is lying on its right side, so the tube feet are sticking straight up out of the plane of view. You can clearly see two of them, though.

Just for kicks, here’s the same larva, photographed with dark-field lighting. This kind of light illuminates the surface of the object being viewed, which is very helpful when the subject is opaque, making it possible to see four tube feet in this picture.

As the tube feet are emerging from the juvenile rudiment, the larval body contracts and gets denser. The arms shrink and the internal skeletal rods that supported them are discarded. At this stage the larval juvenile (larvenile? juvenal?) begins to crawl around on the bottom. The ciliated band that used to propel it through the water and create the feeding current may still be beating, but eventually will stop, as the larvenile will no longer need it. This is usually the time that I see the first spines waving around; it’s interesting to note that tube feet, which originate from the inside of the animal, come first, then are followed by spines. Then again, the spines are part of the animal’s endoskeleton, so maybe it’s not so noteworthy after all.

So they’re getting close to becoming real urchins! Next up: Learning to walk.

Did you notice that I invented a new word? I’m going to start using it regularly.

In the parlance of invertebrate zoologists, competence is the state of development when a larva has all of the structures and energy reserves it needs to undergo metamorphosis into the juvenile form. In the case of my sea urchins, this means that they have four complete pairs of arms, each with its own skeletal rod, and a fully formed juvenile rudiment, which contains the first five tube feet of the water vascular system. A continuous ciliated band runs up and down all eight arms and provides the water current used both for swimming and feeding. The larva will have been eating well and its gut will be full of food. It will have lost the transparency it had when it was younger and will appear to be more solid-looking in the central area.

The first batch of larvae that I began culturing this season are now 42 days old. Some of these are competent, or very nearly so. Last week I isolated about a dozen of these big guys into a small dish, making it easier for me to observe them closely every day. Today they looked decidedly opaque and dumpy, and although some of them were still swimming others were heavy and tended to rest on the bottom of the dish.

Here’s a photo that I took yesterday:

General orientation: This is a ventral view. The animal swims with its arms forward, which defines the anterior portion. Thus the bottom of the cup-shaped body is the posterior. This larva measures ~900 microns along the anterior-posterior axis. Plutei have bilateral symmetry that goes all to hell during metamorphosis, from which the urchin crawls away with typical echinoderm pentaradial symmetry. This wholescale change in body organization is one of the truly amazing things about metamorphosis in these animals. It boggles my mind every time I think about it.

You can see that this pluteus has eight arms. The oblong reddish structure in the middle is the stomach, which has taken on the color of the food the animal has been eating. The strange mixed-up looking structure adjacent to the stomach on the animal’s left side is the juvenile rudiment. Focusing up and down through the rudiment shows that it contains five tube feet. After metamorphosis, the juvenile urchin will use those first five tube feet to walk around as a benthic creature, having spent all of its life up to this point as a member of the plankton.

Today I captured about 20 seconds of a larva feeding. This individual is a day older than the one in the photo above and has more of that opacity that I associate with competence.

This is a dorsal view; if you imagine that you’re looking at the animal’s back, you see that the rudiment is indeed on its left side. The larva’s ciliated band is moving a lot of water, and the little specks that you can see flying around are food cells. There wasn’t enough water in this drop for the pluteus to do any actual swimming, but at this point it’s pretty heavy and would tend to sink to the bottom.

Some time in the next several days these guys are going to start metamorphosing. I will be examining them every day; keep your fingers crossed that I catch them in the act!

On another glorious afternoon low tide the other day, with the help of a former student I collected six purple urchins, Strongylocentrotus purpuratus. Given that we’re in about the middle of this species’ spawning season, I reasoned that collecting six gave me a decent chance of ending up with at least one male and one female that hadn’t spawned yet.

Yesterday, after the urchins had been in the lab for somewhat less than a whole day, I shot them up and waited. Three females began spawning almost immediately (yes!) and one male started a few minutes later. When all was said and done I ended up with four females and two males. It turns out that the largest individual, with a test diameter of almost 10 cm, was a male but didn’t spawn very much at all. I infer from this that he had already spawned in the field before I collected him.

At the current ambient sea water temperature of 14°C, hatching begins around 24 hours post-fertilization. Early this afternoon I checked on the beakers and they had indeed begun hatching. Sea urchins hatch at the blastula stage of development, when they are essentially a ciliated hollow ball of cells. The cilia allow the larvae to swim, but at this size they are at the mercy of even the weakest current. Thus, for the most part they act as particles, getting carried wherever the current takes them.

As the embryos hatch, they swim up to the top of the beaker, then move down towards the bottom. I call this “streaming.” At this point in our artificial culturing system the embryos are living in still water without any current, so this behavior is due primarily to their ability to swim. There is probably some interesting physics involved, but I’m not enough of a physicist to figure out what’s going on at that level. But whatever it is, it’s a really cool behavior to watch:

Rather mesmerizing, isn’t it? Each of those tiny orange dots is an individual embryo. Once the embryos hit the water column I pour them off into larger jars and begin stirring them. Right now they’re small enough to swim on their own, but once they start feeding and growing they get heavier and would sink to the bottom without some current to keep them suspended. The contraption we use to stir jars of larvae is a manifold of paddles connected to a motor that moves the paddles back and forth, creating the right amount of current to keep the larvae from settling on the bottom without getting beat up by the turbulence.

Here’s the paddle table in action. It’s a noisy SOB.

For now the embryos just hang out in the jars and get stirred. Their first gut, the archenteron, will be visible tomorrow and the larvae will be able to eat on Friday. Stay tuned!