Hannah Liechty A09522659 May 22, 2014 Phys 173 Spring 2014 Medicinal Leech Electrophysiology: Retzius Readings Introduction Initially when this lab began, the idea was to be able to set up a system that would allow two electrodes to be placed into two different cells from a fresh leech ganglion and measure the effect of stimulus into one cell on another cell to understand the complex interactions that occur within the ganglion. Yet many difficulties arose such as lighting, focus, issues with the amplifier and the probes, and lack of time to practice probing cells other than the Retzius cell. Finally a reading of the Retzius cell was obtained as seen in figure 4 with periodic pulses for 10 seconds averaging about 1 pulse per second. Experimental Procedure Solutions Preparation In order to properly conduct basic electrophysiology of the leech ganglion two solutions were prepared, saline and potassium chloride/potassium acetate. The saline solution (aka instant ocean) was made with the following salts per liter of solution: mM Composition g/L g/6L g/2L 115 NaCl 6.71 40.26 13.42 4 KCl 0.30 1.80 0.60 1.8 CaCl2 * 2H2O 0.27 1.62 0.54 1.5 MgCl2 * 6H2O 0.30 1.80 0.60 10 Glucose 1.80 10.80 3.60 10 Tris 1.58 9.46 3.16 This was used as a frozen mixture along with the liquid form to transport the leech and conduct the “wet” dissection. Then the potassium chloride/potassium acetate solution was made with 20mM potassium chloride in 1M potassium acetate which was used in the micropipette as an electrolyte pathway for the silver chloride-coated electrode. Leech Preparation Leeches were first obtained from the Kristan Lab then placed in a cold dissection tray as shown in Figure 1A with saline solution. The smaller end of the leech is the “head” where it was pinned to one end of the wax and the large end with the sucker was pinned and stretched to the opposite end. Then using a small scalpel (obtained from Kristan Lab) the leech was fillet open in a zipper-like fashion while being careful to not prod the scalpel too deep as to not damage the ganglia. Then using the smaller pins, the epithelia of the leech was pinned open at intervals of ~1in to reveal the internal structure of the leech as shown in Figure 1B. The lining of the
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Medicinal Leech Electrophysiology: Retzius Readings...2014/05/22 · Medicinal Leech Electrophysiology: Retzius Readings Introduction Initially when this lab began, the idea was to
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