14 Lab 4: Leaky Neuron (Spatial and Temporal Summation)

Leaky Neuron Activity

Ensure you have all the supplies you will need at your table:

• large tray to contain spillage of water
• a 1000 ml beaker or measuring cup (or a large container to catch leaking water and a way to measure that amount of water)
• a 300 ml+ clear cup with holes
• a 300 ml+ clear cup without holes
• a large bowl with water
• six ~50 ml (very small) cups; red cups
• four-five ~100 ml (small) cups; wax paper cups
• one-three ~200 ml (medium sized) cups; paper cups
• a large white tub (for water)
• a data sheet

Background knowledge:

Recall what you learned in class about the chemical synapse and the action potential. Specifically, recall that a presynaptic neuron releases neurotransmitters which diffuse across the synapse. When the neurotransmitter binds with a ligand-gated channel, ions can move through this opened channel and into the cytoplasm of the postsynaptic neuron. Some postsynaptic potentials are excitatory while others are inhibitory. The change in the postsynaptic neuron may be received at a dendrite; however, ions can continue to diffuse through the soma if their concentration gradient allows. Recall that both spatial and temporal summation of PSPs at the axon hillock may allow the activation threshold to be achieved.

For simplicity, only synapses that are excitatory such that the PSPs (EPSPs) bring the membrane at the axon hillock to threshold or beyond are considered in this lesson.

General instructions:

Members of the group will have different roles:

• One person will hold the leaky neuron (cup with holes and threshold marked) over the 1000 ml measuring cup; they will stop the trial by quickly nesting the two cups (quickly putting the cup without holes under the one with the water) when threshold is reached (see Fig. 2). Threshold is reached when the water gets to the green tape line near the top of the 300 ml+ clear cup with holes.
• One person will be the timer and record the time elapsed to reach threshold.
• One person will count cups, watch to see when threshold is reached, and immediately tell the timer and the person who stops the trial. This person needs to be able to see the green line from where they sit.
• One to three students will add water to the leaky neuron, dipping cups of various volumes (based on specific activity) into the reservoir bowl to refill them as needed.

Each team will record the:

• number of cups (EPSPs) needed to reach threshold at the axon hillock
• time to reach threshold
• total amount of water (positive charge) needed to reach threshold
• amount of water (charge) that leaked out
• percentage of water (charge) that leaked out

There is a tray and a large bowl with water at your station/table. The tray is to contain spilled water; the reservoir bowl may need to be refilled after a few trials; if so, add water to the reservoir container with your measuring cup. Keep the reservoir bowl inside the tray at one corner, and the measuring cup in the opposite corner at the other end of the tray.

Before you begin, feel free to confirm that three to four ~100 ml cups of water should fill your clear cup without holes.

Activity 1: Spatial summation of EPSPs from multiple inputs

Fill four ~100 ml paper cups with water from the reservoir bowl.

Hold the leaky neuron (300 ml+ cup with holes and threshold marked) over the 1000 ml measuring cup.

Start the timer and then have two people each dump two cups of water into the leaky neuron at the same time; stop the experiment (by quickly nesting the clear cup without holes under the leaky neuron cup) when the water reaches the threshold line. Record your results (trial 1) on the data sheet. Notice you need the amount of water added and the amount of water lost to leakage. A calculation must also be performed.

If the threshold is not reached with the use of four cups, try again. Fill five of the ~100 ml cups with water, and use three people in your group, pouring 1 or 2 cups of water at the same time, to try to fill the leaky cup to the marked line. Five wax paper cups should be enough to fill your leaky cup to the marked threshold.

Repeat the experiment a few times to practice calling out threshold, stopping the experiment, and recording the time. Record your two fastest times (trials 2 & 3) on the data sheet.

Activity 2: Temporal summation of EPSPs from a single input

Practice:

• Hold the leaky neuron cup over the large beaker.
• Fill the small ~50 ml cup with water.
• Have one person repeatedly empty the water from the small cup into the leaky neuron (large clear plastic cup with lines and holes), refill the cup, and continue to empty it until threshold is reached (see Figure 3).
• Stop the trial when threshold is reached. Repeatedly emptying water from a small beaker represents the responses in the target neuron to a burst of action potentials that produces temporal summation.

Repeat the experiment three times and record the data after each trial on the data sheet. The same group member can perform the repetitions, or another member may take a turn.

Activity 3: Graded local potentials; larger EPSPs

Repeat Activity 2 using a ~200 ml paper or plastic cup; if using paper, a new cup may be needed for subsequent trials if it becomes soggy. Record the data after each trial on the data sheet.

Activity 4: The effect of distance of the synapse from the axon hillock

Position the white container at one of the shorter sides of the tray. Place the water reservoir bowl at the center of this container. Add water to the reservoir container with your measuring cup if needed.

Hold the leaky neuron cup over the large measuring cup at the opposite short side of the tray, just in one corner – as it was for the other three activities. Repeat Activity 2, using the ~50 ml cup, under these conditions three times.. Record your data on the data sheet.

Activity 5: Homework questions

Complete the quiz on Moodle.