Intracellular recording in Snails Final Update – Juan Ferrada

Hi Everyone. Juan here! My two month tour with Backyard Brains has reached its end, and I’m really grateful to have had the opportunity to work on this project.

I had three activites during the “practica” here at Backyard Brains:

• Recording from the Ganglia of Snails
• Helping on the Anemone Project
• Assisting in Outreach.

The snail recording was my main project at 70% of my time, the anemona project occupied about 10% of my time, and the high school outreach was 20%.

Recording from Snails.

The original aim was to record the intracellular action potential of pacemaker-like cells from the parietal ganglia of Helix aspersa, using Backyard Brains hardware for the optics, acquisition and amplification. From the last blog post you can see that we had no problem with dissecting the ganglia or visualizing the neurons with hand pulled pipettes in the microscope, but we couldn’t get any recordings. Recording electrical activity from the leg of the cockroach is different from the ganglia of the snail. Because of the dry environment of the cockroach leg and the strong signal from the leg nerves, the cockroach leg nerve activity is very easy to record. Buuuuut with the snail, we have a really weak signal in a conductive aqueous salt solution, so we must take a different approach to the experiment, as repeated attempts at the BYB lab did not yield results.

Sooo we went to familiar ground (for me) and replicated the experiment with lab equipment from the Laboratorio de Neurociencias at the University of Santiago (Usach). We had divine intervention from Darwin Contreras, a PhD student who just that day had successfully defended his Ph.D. and happened to only be coming back to the lab to get his motorcycle helmet to go motor on home and relax with his growing family. Using a large Faraday cage, a dissection scope, and a high end manipulator (but yes, a Backyard Brains Neuron SpikerBox Pro), we carefully inserted an insulated  blunt tip silver wire into the ganglia.

And we listened to the low background noise coming out from the speaker of the SpikerBox. But….Every now and then there was a rattle, sparse and random enough to not be an artifact, so we recorded it and to the surprise of everyone there it was, spontaneous, asynchronous action potentials. Success!

We witnessed high amplitude spiking (download the raw data if you like!).

And we also witnessed the rhythmic “neuron dying” response.

But this was only a partial success, as it was 1) our only successful recording, and 2) made extracellularly instead of intracellularly. We found a shorter electrode, to prevent the antenna effect, and a faraday cage minimized the noise profile, but we are still far away from the original goal, recording intracellularly from the large neurons in the parietal ganglion.

At the moment, we may seek another preparation for intracellular single unit recording, as the snail preparation is a bit tricky. We may go to the intracellular recordings of the muscles of the tail of the crayfish, or perhaps try another mollusk, say a “macha,” a type of clam very common along the Chilean and Peruvian coast, that we looooovvveeee to eat.

Recording from the Nervous System of Anemones

We continue in the long term project to record from sea anemones. We built a harpoon style electrode…

but the silver wire wasn’t strong enough to pierce the membrane of the oral disc of the anemone. We had heard that there are more neurons around the oral disc (which, in an anemone, serves as its mouth, anus, and reproductive orifice). We will try tungsten next, which is the classic, strong material for small metal electrodes.

But….we had the idea that maybe we could remove the tentacles, like we remove the leg of the cockroach, and attempt a recording in a more controlled environment under a microscope. To our surprise, the tentacles kept moving for an hour  after we had cut them! We may be on to a new preparation, it is very fascinating to watch. Very primordial success, yet waiting, yet to come. See our video below.

We then inserted an insulated blunt tip silver wire into the open end of the tentacle and tried to see if we could measure spontaneous activity or evoked activity (when we touched the tentacle with a probe).

But….we did not get any successful neuron recordings. We are sure there is something here though, there have got to be neurons inside the tentacle. The tentacle is moving, and neurons must be talking to the muscles. The neurons in the anemone tentacle are arranged like sheets between rings of muscle, so it’s a matter of optimizing the preparation. We are always getting closer to the elusive anemone neurophysiology, stay tuned.

Outreach

During the last part of May we taught the students how to build a two stage amplifier circuit from a breadboard, and the students can now recognize the logic of how to manage components like resistors, capacitors, and transistors.

During the last two classes I helped teach about the difference between reactions and reflexes using the knee and elbow.

For the last class, we did experiments measuring the difference between audio and visual reaction times. Data collected in a classroom can be noisy. Supposedly auditory reactions are faster than visual reaction times but we did not observe that difference in the students who had well tabulated data. But I always continue in my experiments. I always continue trying to have compelling data that tells an interesting story.

Bye Guys, Now I have to write my thesis! I’ll miss the late night pizza party experiment sessions with Florencia and Tim and the workshops in the Fablab at the high school Colegio Alberto Blest Gana. I will not miss the cardboard-tasting garlic bread  (pizza delivery company to remain anonymous).