First, you'll want to set up your brain model. Follow the instructions on whatever agar or gelatin you've purchased and put the contents of the whole package into your 1-2 quart container. We use a brain mold, but any container will work.
If it's available to you or you can order it, agar is a bit better than gelatin (such as Jello), since it is more resistant to temperature fluctuations and is more comparable to real brain tissue.
Once in the refrigerator your brain should take 2-3 hours to set. Our final brain product is shown below.
Step 2While your brain model is setting up, you can start building neurons. Open the NEURON graphical user interface (GUI) (it may be named "nrngui" in your program files). Click "Build" and select "CellBuilder." It should look something like what you see below.
If you have never used NEURON before or would like a more detailed guide through building your own neuron model, please see our "Links and Other Useful Resources" page for a link to a step-by-step tutorial through the program.
Step 3Within the CellBuild window, you can go through and build your cell, section by section. In the "Topology" section, you will draw each cell segment individually to create a neuron that looks exactly as you would like it to.
Our basic neuron with three dendrites and a single axon is shown below. Explore making more complicated shapes and neurons with different branching patterns! Don't forget to name your segments.
Step 4The "Subsets" section allows you to group segments by type. This will be helpful later if, for example, you have created a cell with a large number of dendrites and you do not want to have to change the geometry or biophysics of each dendrite individually.
Step 5The "Subsets" section will also give you the option to allow sections to be parameterized in the biophysics section. Do this in the "Parameterized Domain Page" by selecting each subtree of segments and clicking "Create a SubsetDomainIterator."
You will want to create parameterized domains for each of your sections that you hope to stimulate/record from. Don't forget to save your neuron regularly!
Step 6In the "Geometry" section, you can alter the length and diameter of each section. For now, set the length of each segment as 1000 and the diameter as 10. Later, you can play around with different settings to see what it does to your signal!
Step 7The "Biophysics" section can feel a bit complicated, but it's not as scary as it looks! Here, you can set each segment to have passive, resistive, capacitive, and "hh" properties, the last of which we will explain in a moment. The passive, resistive, and capacitive components of neurons work in much the same way as with electrical circuits. For your first neuron, try just turning on the passive and hh components, but make sure to come back and explore was resistivity and capacity do to your signal propagation!
|Hodgkin-Huxley properties, or "hh" properties as they are shown in NEURON are named after Alan Hodgkin and Andrew Huxley who studied the neurons in giant squid and were the first to mathematically describe action potentials. These properties are necessary for a neuron to send an electrical signal and involve the molecular components of how currents are generated within a neural cell. Basically, there are high concentrations of sodium (Na) outside your cells and high concentrations of potassium (K) inside your cells. When enough current is applied to a neuron either from another neuron or by an electrode, channels within the membrane of your neurons open, allowing transfer of these ions into and out of the cell. Because of the large differences in ion concentration between the inside of the cell and the outside of the cell, Na and K flow very quickly through the membrane, creating an electrical current, or action potential. This current then travels down the length of the axon and is transferred to the next neuron.
This electrical current within your body is referred to as bioelectricity and it is what we are recording when we look at neural signals. Your body is generating electricity all the time! It is the same kind of electricity you use in your house when you plug things into the wall, though at a much smaller scale. Just like you are sending an electrical message to your lights when you flip your light switch on, your brain sends electrical messages to your body when you want to move!
Note: Your neuron will only fire if you go into the parameterized domains for each of the subsections and change the distribution of potential and conductance over the length of each segment. This means that you are changing how the current is applied and carried through the segment. There needs to be a gradient to allow the signal to be propagated properly.
Step 8In the "Biophysics" section of NEURON, some of your cell sections will be marked with an "x" before the name Go into the parameterized domain page for each of these sections. This is where you can change the potential and conductance distribution over the length of your cell. You can choose how you want your gradient to be applied by choosing one of the preset mathematical functions or by inputting your own.
Step 9Once you have your cell built, you can stick an electrode in it and give it a shock! To do this, go to "Tools" then "PointProcesses," "Managers," and finally "Electrode." This will allow you to choose either a current controlled or voltage controlled electrode.
Current clamp is most realistic for this first basic experiment. In this window you can change the delay before stimulation turns on, where the stimulation is applied, for how long it is applied, and the amplitude of the current applied.
Step 10To apply the current and look at the subsequent voltage signal, go to "Tools" and click "RunControl." This will give you a window that allows you to run the simulation.
Step 11Finally, under "Graph," click "VoltageAxis." This will automatically plot the electrical potential at the center of your neuron in millivolts. Run a few simulations and see if you can elicit an action potential. It should look something like the plot below. Once you have a functional neuron, you can listen to the signals and apply them to your agar brain! To do this, we will need to take the transmembrane currents—the electrical currents created by the flow of ions into and out of the cell—and convert those into audio files. We will then play those files through our speakers if we want to listen to them or through some severed headphone wires into our brain so that you can record your neural signal.
Step 12To get the transmembrane currents that we need, right click your voltage plot and select "Plot What?" to select which segment to record from (you will likely want to choose the axon) and turn on tracking of Na, K, passive, and capacitive currents (ina, ik, ipas, and icap).
You may also want to change the color of each current type, so that they're easier to keep track of. Below is an example of all four currents on the same plot with ina shown in blue, ik shown in orange, ipas shown in green, and icap shown in brown. Positive current indicates ion flow out of the cell and negative current indicates ion flow into the cell. Notice the quick and drastic the Na flow.
Step 13Now, save each of these currents to individual files. Unfortunately, there is no option for overall membrane current in the NEURON GUI, so we have to save each of them one at a time. To do this, right click the plot and click "Select Vector," then click on your chosen current once. It should flash red while your mouse is clicked and then go back to normal color. Then, on the main menu, go to "Vector" and "Save to File" to save it to a directory. Save all of the transmembrane currents for your simulation into a single folder that does not contain any other files. Give it a name that you will remember and that makes sense!
Step 14Once you have saved all 4 files, input your directory name into the "VNW_simtosound1" Matlab code provided in our "Sample Code" page.
Step 15Run the Matlab script with your directory name and you should be able to hear the signal from the cell you created!
If you want to change how many times your signal repeats, change the sig_L number.