​Sea Urchin Treatments

Danica Heck
4/25/17
Dr. Burleigh
Introduction

Lithium Chloride is known to affect the Wnt pathway, it does so by blocking the GSK-3.

The LiCl stabilizes beta catenin so it does not degrade and is found throughout cells, not just

localized in the micromeres. This leads to animalization which is the animal cells becoming

specified as endo and mesoderm. Injecting LiCl will also lead to abnormal ingression and

development. To begin, sea urchin eggs develop based on temperature, study shows spawning

has a direct correlation with the temperature of the water. The rate at which the temperature

changes can either stimulate gonad development or spawning. LiCl is thought to affect the cell

fate in the sea urchins. This procedure was run from February 7​th​ until February 14​th​ collecting

multiple samples each day to record as much development as possible.

Materials and Methods

Spawning of sea urchins can be induced by injecting them with 1mL of 0.5 M KCl

solutions in the soft membrane around the mouth. This causes the sea urchins to release their

gametes, males are white and the female’s eggs are orange. To collect these gametes, the

females are turned upside down and the males are right side up letting the sperm and eggs drip

into a beaker. The female’s eggs were falling into a beaker with ASW and once collected the

ASW was poured out. The gametes were then collected after several minutes. LiCl was then

used to treat the fertilized sea urchins in four different doses, 3.8mM, 7.5mM, 15mM, 30mM.

Each dose however had multiple samples incase of any errors that could occur. So the beaker

full of fertilized eggs were divided equally between each sample, there were two 15mM, three

3.8mM, three 30mM, and two 7.5mM doses. When the samples were collected the tubes were
labeled then placed in a refrigerator at 4 degrees Celsius so the development progresses but at

a slower pace, making it easier for each student to document the progress. If kept at room

temperature the eggs would develop too fast and once a student would to check on it multiple

life stages would have gone by without documentation ruining the experiment.

Results/Conclusions

Based on the hypothesis will treating with LiCl affect the gastrulation of sea urchin

embryos due to disruption of the Wnt pathway was proved correct. The more LiCl injected the

more deformities occurred. As shown by the pictures below each dose developed in accordance

to each other, all reaching the gastrula or hatched and swimming but the physical development

was abnormal. The gastrula would be deformed and would often be mistaken for a prism or

early pluteus. Each cell was pronounced dead at the end of the experiment due to a power

outage stunting the experiment. Without these extra hours/days the embryos were unable to

develop into a full pluteus. This is a very big issue because the Pluteus is where all abnormalities

and deformities are observed. In other experiments the Pluteus is malformed looking like a

snowman or having a greatly enlarged gut. Those experiments performed by Tufts University

show the extent of the deformities. With the experiment conducted in this experiment it is

seen that there are some abnormal ingressions, however it is not as prominent as It would be if

the eggs developed fully. We were able to see some gastrulas that were deformed, taking a

shape of a lightbulb or other hatched and swimming embryos having difficulty swimming, also

showing some deformities. Because of the early death due to power outage a lot of information

was taken away from this experiment. Students were unable to see the development during

that day and could have missed a series of developmental progress. Overall, each student got
as much information as possible, although the experiment was stunted it was still proven that

LiCl does in fact affect the gastrulation.