Recording the EOD of Weakly Electric Fish
Equipment Needed Per Rig:
Standard. MacLab data acquisition system and Macintosh, MacLab Chart and Spike histogram extension software, 1 DAM60 Amp and connectors, Grass stimulator, oscilloscope and audio monitor, BNC cables, barrels, and Ts.(see Equipment page for model information etc. for lab report), male banana to BNC adapter, alligator clips.
Special. 3 x25 cm insulated solid EOD monitoring wires (bath end stripped .5 cm; other end female pin miniconnector), 2 lengths of wire to connect two tanks electrically, 20 l glass tank (lwh 40 x 20 x 27 cm) filled with distilled water and brought to around 300 microsiemans conductivity using fish bath solution (see Maler's Sludge formula), aquarium water heater, bubbler & air stone, fish tube, 2 pieces of fine mesh plastic screen to cover both ends of tube and elastic bands to hold these on, pH meter and conductivity meter, NaHCO3 to raise pH of bath to around 7.
Part 1 - Equipment Orientation
1) Setup analog scope, audio monitor and stimulator to view and hear specific stimuli sent to the scope and audio monitor from the stimulator. Think about polarity.
2) Attempt to display waves and notice:
a) The x and y units and scales. Do the readings make sense from the input values?
b) Try to vary the input stimulus (voltage and frequency) and notice changes that occur in
c) Try to trigger the stimulus on the scope.
d) Is it possible to lock a wave into position on the scope without triggering? Why does
Part 2 - Experimental Protocol
1) Verify that the bath is around 27 degrees C, ph 7, and conductivity is around 300 microsiemans. Setup bath electrode, connect to amp. Signal output of amp should eventually terminate on to MacLab once ready for data acquisition; use a BNC "T" to branch off to oscilloscope and audio monitor.
2) Expose one end of tube and gently encourage fish to go into it; at this point put the screen back over tube with elastic band. Orient tube so that you can see into it easily (to tell which way the fish is oriented). Roughly center the tube in the middle of the tank.
3) Tape on 3 electrodes to the side of the tank; connect to male DAM60 miniconnectors; then position electrodes so that A goes to the end of the tube which is closest to the fish's head; B goes to the tail end; and the ground electrode is near the middle of the tank.
4) Turn on amp (try 10K gain to start, open filters completely (0.1 Hz low filter cutoff, and 10KHz high cutoff, A-B recording mode) and adjust scope, you should hear the animal's discharges. Note the variation in discharge rate, attempt to roughly ascertain firing rate on the scope (it can be quite slow, which makes it difficult to measure on a scope with no storage function).
5) Set MacLab sampling rate to maximum (40,000 samples/s). Given that in fast mode, MacLabs can only acquire 16,000 samples at a time, how many pulses would you expect to acquire at the approximate EOD discharge rate measured above? Acquire the 16,000 samples. Use the zoom tool to examine the waveform up-close. Characterize about 5 discharges (you may need to acquire more than one set of 16,000 samples to get this many). Save your individual discharge recordings to a file on the desktop with the format "eod_lab_[lastname]". Here's a minimal set of parameters you can characterize (if you think of additional ones as you look at the pulses include those):
*What is the minimum and maximum amplitudes for both phases of the waveform
(go through each pulse to determine this).
*What is the minimum and maximum widths for each phase?
* Are the pulses "head positive" (first phase positive) or "head negative"
6) In terms of the efferents to the electrocytes within the electric organ, how does Gnathonemus coordinate all of its electrocytes to obtain a pulse that is so short?
7) Predict (in the form of sketches of the expected waveforms) what will happen to the measured EOD when you do the following manipulations of the electrodes (each manipulation from the initial set-up, not cumulative):
a) Move the A electrode over to a corner at the same end of the tank.
b) Move the B electrode near to the ground electrode
c) Switch input select to A only
d) Switch input select to -B only
Predictions in hand, do these manipulations and observe if you were correct. If so, why; if not, explain the discrepancy.
8) Close your individual EOD recordings file and open a new one. Now that we've characterized individual discharges, characterize a few trains of pulses. To do this, reduce your sampling rate to 10K (how many pulses should you get now?). Acquire 16K of samples--this will be your "resting activity" data. Now let the fish out of the tube and again record another 16K of samples to obtain your "active" data. Characterize each train using the Spike Histogram extension. Here are some parameters to note down:
* What's the mean, standard deviation, and coefficient of variation of the interval distribution (remember that the histogram will be drawn for the selected portion of data, and the statistics will be done on the selected portion of the interval distribution)? Select an appropriate bin width.
* Use the ratemeter function to look at the binned firing rate. What's the minimum and maximum firing rates? How does the firing rate histogram relate to the interspike interval histogram?
* Compare the statistics you collected for the "resting activity" and "active" data sets. Comment on the differences you see; suggest neural/behavioral reasons for these differences. Are their analogous phenomena in other animals? (clue: variations in whisker movement depending on the behavioral state of a rat).
9) Jamming avoidance response in pulse fish. Pulse fish will try to avoid firing a pulse that overlaps with that of a nearby pulse fish; this is seen as a "following response" with one fish following the pulse of another fish with a pulse of their own. This is the pulse-fish analog of jamming avoidance in fish that emit a continuous wave-type EOD. to the jamming avoidance response We will try to observe this by shorting one tank to another so that each fish can sense the other. Place one length of insulated wire near the fish's head, the other near the ground electrode, and do the same with the other ends in a nearby tank. Put fish back in tubes in both tanks. Close down the filters of your amp (300 Hz for the low filter, 0.1 KHz for the high filter), and increase the gain x10. Now the discharges will be distorted; but since we are here just interested in their onset times this doesn't matter. With the filters closed we can stretch the .5 ms pulse to something that can be recorded by the MacLab's in continuous mode (1K samples/s). Set two channels to have this acquisition rate, and plug the leads from each tank's amp into channels 1 & 2. Now, acquire some data without the leads between the two tanks connected; then connect the leads together while continuing to acquiring data and see if you can observe the following response. What is the typical latency of the following pulse? How long does it take for the animals to start "cooperating" in this way? What happens to the pulse rate of a fish when you excite the other one by feeding it or gently moving it?
10) Extra credit: How would an animal have to modify electroreception/generation if it went from being aquatic to being terrestrial?