Can Embryonic Zebra Finch Survive Without Their Shell?

Hopefully, this peculiar title is what brought you here. I certainly am intrigued by the potential of creating a shell-like environment ex ovo that allows a zebra finch embryo to develop normally for a few days. You may be asking, “What’s the point of  a shell-less bird and why should I care?”. That is the question I am attempting to answer in one of my research projects this summer.

As I previously stated in my past blogs, I am interested in axon degeneration in zebra finch and how SARM affects that process. I also began to talk about a project I was working on where I was crafting “hammocks” for the embryos. The point of these hammocks is to create an environment where the finch, ex ovo, can grow normally and stay alive for a day or two. The purpose of this is so that I can better understand how axon degeneration proceeds in a living organism rather than looking at a DRG or retinal explant on a microscope. Instead, I would be injecting an indicator of some kind into the embryo’s vasculature (Mito-Tracker perhaps?) and let the body process and stain axons so that I can visualize degeneration during growth. This all sounds wonderful and grand, but the question is, “How can I keep them alive?”.

After researching the topic of ex ovo experiments, I came across a helpful piece of literature. In the paper, the researchers were able to grow a chicken embryo outside of their shell for multiple days in an incubator. When I first read this, I was expecting them to have used some type of expensive, complex technology. Boy was I wrong. Much to my surprise, all I needed was a plastic cup, Saran wrap, and water. Miraculously, they were able to use these simple household items to create what is called a “hammock”. The reason for this name is that you fill up the cup roughly 3/4 with water, incubate the water to 37 degrees, then take the Saran wrap and place it over the cup and droop it so that it is touching the water. This creates a hammock-type look to the Saran wrap, hence the name. They cracked open the egg and set it in the hammock so that it is touching the water through the wrap and can be exposed to heat without direct contact with the water. You then cover the cup with more wrap and leave it in the incubator. Sounds easy enough to make and succeed, right? Wrong.

My first attempt at making a hammock with a surviving embryo did NOT go so well. After I had opened the egg into the hammock, I struggled to determine if the embryo was alive. To do so, I either needed to see movement or blood flow, none of which I could find. My only option was to manipulate the embryo around to try and view it’s major veins and arteries to detect blood flow. I finally found the right area but in the process, I had manipulated the subject too much and killed it due to blood pooling. I decided I needed to manipulate the embryos with forceps as little as possible so I ran an experiment where I simply cracked open the egg and placed it in the hammock. I immediately took it to the incubator and checked it for blood flow every 30 minutes. After 2 hours, all three subjects had died. This led me to believe that it is not my manipulation that is killing them, but rather the environment they are exposed to.

Currently, my goal is making these embryos survive. So, I have to alter the environment and make it more controlled and similar to their life inside of a shell. The next step in my experimentation is using a more advanced incubator where I can monitor carbon dioxide levels, humidity, and temperature more closely. I will begin this new experiment in the coming week and hope to see better results. I look forward to my next blog and what kind of advances I have made in my research!