lesson a15: earthworm smooth muscle - data … · lesson a15: earthworm smooth muscle . ... cardiac...

Updated 02.05.14 BSL PRO Lesson A15: Earthworm Smooth Muscle Developed in conjunction with Department of Biology, University of Northern Iowa, Cedar Falls

Objectives:

1. Record smooth muscle contraction from an isolated segment of earthworm gut.

2. Observe effect of temperature on rate of contraction.

3. Measure effect of drugs on peristaltic contractions:

a. Serotonin (5HT) or Epinephrine (EPI) (Sympathomimetic) b. Acetylcholine (ACH) (Parasympathomimetic) c. Ca++-free annelid Ringer’s solution d. Atropine (ATRO) (Acetylcholine blocker)

4. Plot log-dose response curves for 5HT or EPI, ACH and ACH in ATRO

Equipment:

Biopac Student Lab System: o MP36 or MP35 hardware o BSL 4.0.1 or greater software

BSL PRO template file: “a15.gtl” Force Transducer (SS12LA includes S-hooks) 10 gram calibration weight Tension adjuster (HDW100A or equivalent) Tissue Bath 2 Ring stands Earthworm (well fed) Separation funnel support Test tube rack 16 – 10 ml test tubes Plastic 50 ml graduate cylinder (for tissue bath) Funnel with tubing 2 Petri dishes O2 cylinder with regulator 250 ml Erlenmeyer flask Surgical thread (non-stretch nylon or equivalent) Examination/surgical gloves Light source Dissection Pan Dissection Microscope

Fine dissection tools Goggles Sharpie Glass probe or small wooden swab sticks Automatic pipettes Pipette tips (1 ml) 10 T-pins for dissection Parafilm Scissors Complete Earthworm Ringer’s1 (ER): dissolve 6 gm

NaCl, 0.12 gm KCl, 0.20 gm CaCl2 (anhydrous), and 0.10 gm NaHCO3 (pH 8.0) in 1 liter of dH2O. 50 ml should be refrigerated to approx. 5° C / 41° F

Ca++-free Earthworm Ringer’s: dissolve 6.0 gm NaCl, 0.12 gm KCl, 0.10 gm, NaHCO3 (pH 8.0), in 1 liter of dH2O

Drug preparations: 5% Ethanol (dilute absolute, no methyl alcohol) Potassium Chloride (KCl) 1.0 M (3 ml) 5HT or EPI 10-2 M (0.2 ml) ACH 10-2 M (0.2 ml) ATRO 10-2 M (0.5 ml)

Ice cooler (for drugs)

1Earthworm Ringer’s should be used to maintain a healthy earthworm preparation; do not use plain water, which may act as an insulator inhibiting the signal and causing cells to hemolyze.

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BSL PRO Lesson A15 BIOPAC Systems, Inc.


Background: Although skeletal muscle is by far the most abundant muscle tissue in the vertebrate, smooth muscles is widely distributed and diverse in its behavior and physiological properties. It is important in homeostatic mediation by the autonomic nervous system. It forms layers in all hollow organs including the digestive tract, blood vessels, gall and urinary bladder, sweet and mammary glands as well as respiratory pathways. It also has specialized functions in tissues such as the kidney and the eye. It has several unique characteristics distinguishing it from other types of muscle: it lacks striations, contracts spontaneously, maintains contractility when stretched, and responds to both neurotransmitters and hormones. Smooth myocytes in many tissues are connected by gap junctions uniting them into a syncytium in which all cells response to a single stimulus. Smooth muscular tissue differs from skeletal by lacking extensive internal stores of Ca++ to regulate contraction. In smooth myocytes most Ca++ comes from external sources by passing across the myocyte membrane. Consequently, understanding the physiology of smooth muscle will provide insights into many aspects of autonomic regulation.

Spindle-shaped smooth myocytes are distinct for their apparently unorganized system of contractile proteins. Cardiac and skeletal cells have thick and thin proteins organized into parallel and series arrays of sarcomeres giving them a “striated” appearance. The filaments do not have this appearance in smooth myocytes. Rather, the organizational center for smooth muscle proteins is the dense body. These are bound to intermediate filaments or the cell membrane. They bind and support thin filaments. To contract, activated myosin chains (thick filaments) slip between the actin filaments attached to the dense bodies and initiate the motion of contraction. The “random” arrays of filaments cause the smooth myocytes to become globular distributing the force of contraction uniformly over the sarcolemmma.

To initiate contraction, Ca++ is used to promote action-myosin formation. However, smooth myocytes do not have troponin. They possess another, related Ca++-binding protein, calmodulin. Quiescent smooth myocytes are induced to contraction three ways: Membrane depolarization by (1) action potential, (2) receptor activation, or (3) stretching. Hyperpolarization of the membrane potential relaxes or inhibits contraction in the myocyte. In general, stretching and membrane depolarization will introduce Ca++ into the cytoplasm for activate contraction. On the other hand, some receptors transduce cellular enzymes causing them to produce a cytoplasmic second messenger known as inositol trisphosphate (IP3). This will signal internals stores to release Ca++ activating a specific mechanism allowing Ca++ to move in to the cell across the cell membrane. Consequently, Ca++ introduced into the myocyte by any of these processes will activate calmodulin. When activated, the Ca++-binding protein will then attach to an enzyme known as myosin light-chain kinase (MLCK) that phosphorylates myosin. This event initiates the formation of a rigor complex between actin and myosin resulting in cellular contraction. Unlike skeletal or cardiac myocytes, these are not fast or intermediate speed, all-or-none twitches. They are much slower and persistent contractions. They can vary in strength and are called graded. The degree of contraction is based on the amount of Ca++ allowed to enter the cell. Regardless of amount, as long as Ca++ is free in the cytoplasm, the contraction will continue in a fashion something like tetanus in skeletal muscle. When Ca++ is removed, a second enzyme is activated, myosin light-chain phosphatase (MLCP). This enzyme removes PO4 from the actinomyosin complex and causes the myocyte to relax. Overall cytoplasmic levels of Ca++ are kept low by either Ca++-pumps or Na+/Ca++-exchange antiports that extrude the ion from the cell.

As a result of various configurations on Ca++ entry, MLCK, and MLCP activity, four different patterns of contraction are seen in smooth muscle tissue. First, in a fast, phasic contraction, Ca++ enters and is extruded quickly following stimulation. Second is a slow, tonic contraction where Ca++ enters slowly and is extruded slowly after stimulation. Third, if Ca++ persists in the cytoplasm but is occasionally decreased rapidly, the tissue is usually contracted and occasionally relaxes but returns to the contracted state. This is known as sphincter behavior. The fourth, and last pattern of contraction, is rhythmic and known as an oscillator which is seen in peristalsis. In these tissues, Ca++ maintains a hyperpolarized membrane potential via Ca++-dependent K+ channels. As Ca++ is removed, the K+ channels close depolarizing the cell membrane potential. If the depolarization reaches a threshold, voltage-gated Ca++ channels will re-introduce Ca++ into the cytoplasm and the K+ channels will hyperpolarize the membrane potential.



Low levels of Ca++ and high K+ are achieved by both the Ca++-pump and the Na+/K+ exchange pump. The rate and amplitude of contractions in peristalsis can be regulated via hyperpolarizing or depolarizing the resting membrane potential of the cellular oscillator driving gastric motility.

The contraction state of a smooth myocyte or syncytium is regulated by the autonomic nervous system. The contractions are graded with their strength/frequency being up or down regulated by sympathetic or parasympathetic agents depending upon physiological demands at any moment. In general, the autonomic agents have antagonistic effects on a tissue; one up-regulates while the other down-regulates. The direction of regulation for a sympathetic or parasympathetic agent depends upon receptor coupling. For example, in vascular smooth muscle, norepinephrine (NE) from sympathetic sources will promote contractions. On the other hand, in the digestive system, contractions are stimulated by acetylcholine (ACH) from parasympathetic sources and relaxed by NE. In a sphincter where smooth muscles remain contracted, ACH may momentarily relax the smooth muscle allowing material to pass from one compartment to another. Without parasympathetic stimulation, the sphincter remains shut. And lastly, the rate of an oscillator regulating peristalsis may up or down regulated by the relative abundance of sympathetic or parasympathetic agents.

Here we will use the smooth muscle system in the annelid digestive tract as a model to study the basic physiology of smooth muscle. Gastric motility results from peristalsis produced by contractions of longitudinal and circular smooth muscle in the earthworm gut. There are advantages in using tissues from this invertebrate. The worms are inexpensive and easily located at most bait shops year around. An animal care facility is not required and generally no special authorization is required for their use. In addition, the tissue works well at room temperature (21 – 26° C). Consequently precisely heated or CO2 - aerated solutions are not required. The smooth muscles of the earthworm digestive tract are activated by acetylcholine (ACH) and inhibited by serotonin (5HT) or Epinephrine (EPI). Thus, we can simulate autonomic regulation of the smooth muscle in vitro.

Atropine (ATRO) belongs to a family of compounds that will bind to some ACH receptors. It occurs naturally in plants known as “nightshades.” The substance binds to the ACH receptor but produces no effect. It is known as a competitive antagonist or inhibitor of the ACH receptor. ATRO blocks the effect by covering the receptor and preventing ACH from having its usual effect in cells and tissues. In this study you will use ATRO to alter the effectiveness of ACH is stimulating the amplitude and frequency of rhythmic peristalsis in the earth worm gut. ATRO will be introduced and kept at a more or less constant concentration and the concentration of the agonist (ACH) is increased.

About the lesson data

You will observe the recordings before and after the various treatments and look for and measure changes in the rhythmicity and in the tonus of the smooth muscle contractions. The Biopac Student Lab PRO will be used to record the contractions.

Rhythmicity refers to the pattern of the muscle contractions. In particular, you will be looking at three aspects of rhythmicity: (1) the rate (frequency) of the contractions, (2) the amplitude (size) of the contractions and the (3) regularity of the contractions. You will observe and make note of changes in any of these aspects of rhythmicity. If the interval between contractions is irregular or the amplitude of the contractions varies considerably, then the contractions are arrhythmic.

The smooth muscle in the walls of the digestive tract (and many other locations) maintains a constant low level of contraction, known as tone or tonus. Tonus refers to the amount of tension continuously generated by the muscle. Usually there are a small percentage of muscle fibers that are in a constant state of contraction while the majority of the fibers undergo rhythmic contraction and relaxation. An increase in the percentage of fibers in a state of continuous contraction results in increased muscle tonus; a decrease results in a reduction of tonus. Shifts in the baseline position of the recording on the y-axis indicate changes in tonus.

Instructor Note:

One lab group will only be able to perform a few of the listed recordings/experiments in a lab period. You can assign a different series for each lab group and then have each group share the data files.


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Setup and Calibration:

It is assumed that the MP36/35 is connected to the host computer and that the Biopac Student Lab software has been installed and is known to work with the MP unit.

1. Turn OFF the MP36/35 unit.

2. Plug the SS12LA Force Transducer into CH 1.

Figure 1 Figure 2

3. Place the tension adjuster (BIOPAC HDW100A or equivalent) on the ring stand, and attach the BIOPAC SS12LA Force transducer such that the hook holes are pointing down (Figure 2). Roughly set so that transducer is level both horizontally and vertically.

4. Set the tension adjuster height such that it is approximately ¼ the distance from the lowest setting. This will allow the majority of the range to be used for adding tension (raising the adjuster).

Note: Do not firmly tighten any of the thumb-screws at this stage.

5. Place the small “S-hook” in the hole labeled “50 g.”

6. Turn ON the MP36/MP35 hardware. Turning ON the hardware after the connections are made minimizes the chance of instrumentation errors caused by Electrostatic Discharge (ESD) during plug-in.

7. Launch the BSL PRO software and open the template file “a15.gtl” in one of two ways:

a) From Startup dialog, click the PRO Lessons tab and double-click on the lesson title in the list. If you do not see the lesson title, try the next opening method.

b) From Startup dialog, choose the Create/Record a new experiment option, click Open Graph template from disk and then click OK. Navigate to the file, select it and click Open.

8. Click on the Calibration button (Wrench icon) in the vertical scale region and follow the instructions in the pop-up dialogs. The first part of calibration is performed with only the S-hook attached to the force transducer (Figure 3). The second calibration is performed with a 10 gram weight attached, but after it has stopped swinging (Figure 4).

Note: Do not touch the bronze weight with exposed fingers, hands or frog Ringer’s solution.

Figure 3 Figure 4

Calibration part 1 – only “S-hook” Calibration part 2 – 10 gram weight

Tissue Bath Setup:

© BIOPAC Systems, Inc of 14


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Figure 5 shows a low cost tissue bath system. The funnel and tubing connect to a T-valve at the bottom of the 50 ml graduated cylinder that sits within the water bath. The other end of the T-valve connects to drain hose that is controlled by a hose clamp. An O2 gas cylinder with regulator is used to create a bubbler to aerate the ER bath. The earthworm gut will be connected between the tissue bearer (bottom of tissue bath) and the BIOPAC force transducer using thread. The tissue bath is shown immersed in a water jacket, which is not required, but acts as a temperature stabilizer.

1. Add exactly 50 ml of ER solution to the internal bath.

2. Using a thermometer, measure the temperature of the ER bath (not water bath) and record it in your notebook.

Figure 5

Earthworm Preparation:

Notes:

The earthworm gut is very fragile and must be handled with care.

The earthworm should be well fed to insure a full crop/gizzard which helps with the strength of contractions (muscles are stretched a bit).

1. Rinse the earthworm under a tap to clean off any dirt.

2. Place the earthworm in a Petri dish with 5% ethanol in ER, to anesthetize it.

The worm will stop moving when it is anesthetized and should not respond to a pinch with forceps (could take 5 – 10 minutes).

Take it out soon after it stops moving as prolonged exposure to the ethanol will reduce the contraction amplitude.

3. Rinse the earthworm to remove any excess anesthesia.

4. Pin the earthworm dorsal side up on a flat dissecting dish.

5. Place the dissecting dish on the stage of the microscope.

6. Expose the crop, gizzard and about 3 cm of intestine:

a. Make a small slit with the scalpel near the anterior then cut toward the posterior taking care to cut through only the body wall.

Figure 6

b. Use T-pins to hold the body wall open (exposing the gut).

c. Flush with Earthworm Ringer’s (ER).

d. Carefully break the septa connections between the gut and the body using a non-conductive blunt probe (wood or glass).

7. Attach thread onto each end of the portion of the gut that will be used for the experiment. Make sure the two lengths of thread are enough to attach the gut to the transducer and tissue bearer.

a. Tie a ligature just above (anterior end) the crop.

b. Using the other thread, tie a ligature onto the intestine at a point 1 – 2 cm below the gizzard.



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8. Using the scalpel, cut the gut above and below the ligatures so that the portion of interest can be removed.

9. Carefully remove the gut section from the worm using the thread, and place it in the second Petri dish

10. Rinse thoroughly with ER.

11. Tie the posterior end of the worm to the tissue bearer hook and the other end to the transducer S-hook.

Note : Make sure that the thread is loose and not pulling on the gut.

12. Raise the tissue bath so that it covers the gut with solution.

13. Adjust the tension adjuster so that the slack is taken out of the thread but DO NOT apply any line tension yet.

14. Carefully turn on the O2 gas regulator for the bubbler. Only a small flow of gas is required to aerate and mix solutions.

Note: The O2 bubbler may cause artifact (noise) on the force recording. If you see excessive artifact try turning off the bubbler prior to the recording and turn it back on between recordings.

Drug Preparation:

1. Use the Erlenmeyer flask to warm about 50 ml of ER to approx. 37° C / 99° F.

2. Make up two test tubes each containing 1.5 ml of 1.0 M KCl.

3. Make a series of 6 dilutions of 10-2 M ACH from 10-3 M to 10-8 M. To make the dilutions, fill each test tube with 1.8 ml ER. Place 0.2 ml of 10-2 M ACH (stock) into test tube 1 to make a solution of 10-3 M ACH. Pour 0.2 ml of 10-3 M ACH (from test tube 1) into test tube 2 and add 1.8 ml ER to make 10-4 M ACH. Continue this process of taking 0.2 ml from the last test tube and pouring it into the next test tube to make the 6 different concentrations. Always dilute with ER.

4. Make a series of 6 dilutions of 10-2 M Serotonin (5HT) or 10-2 M Epinephrine (EPI) from 10-3M to 10-8 M the same way as for ACH.

5. Fill a test tube with 0.5 ml of ATRO (10-2 M) and dilute with 1.8 ml ER.

Note: Label the test tubes with the drug and concentration using the Sharpie. Cover the top of each tube with Parafilm.

Recordings

One data file containing several recordings will be saved for this lesson. Each time the Start button is clicked; new data will be appended onto any previous data. At the start of each recording, an Append Event Marker will be automatically inserted. To keep track of the data, manually change the append marker labels to match the recording name (i.e. “Baseline,” “Temperature Change,” etc.).

Figure 7

Notes:

Check with your instructor to determine which recording modes will be used.

If after beginning a recording, you notice that the baseline thread tension has fallen to 0 grams, carefully adjust the tension then redo the recording; click Stop, click on the Rewind toolbar button, then click Start.

If there is excessive noise on the Force signal, try turning off the bubbler prior to the recording and turn it back on between recordings.

It is assumed that the user has a basic knowledge of the software which includes data selection, zooming in and out, auto-scaling, taking measurements and placing event markers. Review the tutorial if necessary (see options in the Help menu).

Always review the recording steps prior to clicking Record.

The time interval between contractions can be very long (more than a minute,) so be patient when looking for contractions.




1: Baseline Recording

1. Press Start to begin recording.

2. Carefully rotate the tension adjuster knob to apply slight line tension to the gut while monitoring the data in the graph. A slight baseline offset should be seen in the graph along with contractions. Adjust the tension to produce maximal contractions.

3. Wait for the signal to settle into a stable baseline. Adjust the tension if needed.

4. Click Stop to end the setup recording.

5. Redo the recording: click on the Rewind toolbar button, and then click Start.

6. Click in the marker label region and then type in “Baseline”

7. Record for 3 minutes, and then click Stop.

2: Effects of Temperature Changes

1. Record the temperature of the ER bath (obtained during setup) into the append marker label field (i.e. “Baseline 21 deg. C”).

2. Drain the ER bath and refill it with 50 ml of warm (37° C / 99° F) ER.

3. Measure the temperature of the ER bath and record it in your notebook.

4. Click Start to begin recording.

5. Manually enter “Temperature Change” and the bath temperature (i.e. “Temperature Change – 37 deg. C”) into the marker label field.


7. Drain the ER bath and replace it with 50 ml of cold (5°C / 41° F) ER.

8. Measure the temperature of the ER bath and record it in your notebook.

9. Click Start to begin recording.

10. Manually enter the measured temperature into the append marker label field.


12. Drain the ER bath and replace it with 50 ml of room temperature ER.

3: Effects of Potassium (K+) Depolarization and a Ca++ - free medium

1. Click Start and enter “KCl” into the marker label field.

2. Add 1.5 ml of 1.0 M KCl into the 50 ml ER bath. This will produce a concentration of 30 mM K+ in the ER.

3. Record for 5 minutes then click Stop.

4. Drain the bath and replace with regular, room temperature ER.

5. Click Start and enter “ER after KCl” into the marker label field.

6. Record for at least 5 minutes to allow the preparation to recover, and then click Stop.

7. Drain the bath and replace it with 50 ml of Ca++-free ER.

8. Click Start and enter “Ca++ free ER” into the marker label field.

9. Wait for the contractions to diminish so that the preparation appears to be dormant.

10. Add 1.5 ml of 1.0 M KCl into the bath. Press the Esc key to enter an event marker and label it “KCl.”


12. Replace the bath with 50 ml of regular, room temperature ER and allow the rhythmicity and contraction amplitude to return to normal.



4: Effects of Acetylcholine

1. Click Start and enter “ACH” into the marker label field.

2. Starting with lowest concentration of ACH (10-8 M,) add 0.5 ml to the 50 ml bath. When the drug is placed in the bath, press the Esc key to enter an event marker and type the concentration into the label (i.e. “10-10 M”).

3. Note: Since 0.5 ml of ACH is being added to 50 ml of ER, the actual concentration of the drug is 1/100 the original dilution. (i.e. 10-8 M = 10-10 M final).

4. Record for 5 minutes.

5. Repeat Steps 3 and 4 with the next ACH concentration (10-7M,) etc. Continue until you have added 0.5 ml 10-3M ACH to the bath.

6. Click Stop.

7. Drain and refill the bath with 50 ml fresh Earthworm Ringer’s.

8. Allow the preparation to stabilize for at least 5 minutes before proceeding.

5: Effects of Serotonin or Epinephrine

1. Click Start and enter “5HT” or “EPI” into the marker label field.

2. Starting with lowest concentration of 5HT or EPI (10-8 M,) add 0.5 ml to the 50 ml bath. When the drug is placed in the bath, press the Esc key to enter an event marker and type the concentration into the label.

Note: The actual concentration of the drug is 1/100 the original dilution. (i.e. 10-8 M = 10-10 M final).


4. Repeat Steps 3 and 4 with the next concentration (10-7M,) etc. Continue until you have added 0.5 ml 10-3 M 5HT or EPI to the bath.

5. Click Stop.

6. Drain and refill the bath with 50 ml fresh Earthworm Ringer’s.

7. Allow the preparation to stabilize for at least 5 minutes before proceeding.

6: Effects of Atropine

1. Click Start and enter “ATRO” into the marker label field.

2. Add 0.5 ml of ATRO (10-2 M) to the tissue bath (final concentration is 10-4 M).

3. Starting with lowest concentration of ACH (10-8 M,) add 0.5 ml to the 50 ml bath containing ATRO. When the drug is placed in the bath, press the Esc key to enter an event marker and type in “10-10M ACH/ATRO” into the label.

Note: The actual concentration of the drug is 1/100 the original dilution. (i.e. 10-8 M = 10-10 M final).


5. Repeat Steps 3 and 4 with the next highest concentration (10-7 M) of ACH. Continue until you have added 0.5 ml (10-3 M) ACH to the bath.

6. Click Stop.

Note: In this experiment the ATRO concentration remains constant as you increase ACH concentration.


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Done

To save recorded data, choose File menu > Save As… > file type: BSL Pro files (*.ACQ) File name: (Enter Name) > Save button

Data Analysis:

Fill in the values for the tables with the aid of the software measurement tools. Ignore any table entries that do not pertain to data you recorded.

Measurement examples:

To find the amplitude a contraction, select and area that encompasses the peak and baseline and use the P-P measurement.

Figure 7

To find the level of Tonus, select an area in between the contractions and use the Mean measurement.

Figure 8

To find the contraction frequency, select the area from the peak of one contraction to the peak of the next contraction, and use the BPM (beats/pulses per minute) measurement. Because the frequency can vary greatly, you should take a few different measurements within the data segment and then compute the average frequency.

Figure 9

Temperature Notes:

Condition Measured Temperature

Normal, room

Warm

Cold

Table 1




Condition Amplitude

(grams)

Tonus

(grams)

Frequency

(Pulses per minute)

Baseline

Warm ER

Cold ER

KCl

ER after ACl

Ca++ free ER

KCl after Ca++ free ER

ACH - 10-10 M

ACH - 10-9 M

ACH - 10-8 M

ACH - 10-7 M

ACH - 10-6 M

ACH - 10-5 M

5HT or EPI - 10-10 M

5HT or EPI - 10-9 M

5HT or EPI - 10-8 M

5HT or EPI - 10-7 M

5HT or EPI - 10-6 M

5HT or EPI - 10-5 M

ATRO with 10-10 M ACH






Table 2



Calculate the percentage (%) change in frequency and amplitude vs. the Baseline (control)

ACH:

Concentration (M) % Change in Amplitude % Change in Frequency

10-10

10-9

10-8

10-7

10-6

10-5

Table 3

5HT or EPI:

Concentration % Change in Amplitude % Change in Frequency

10-10

10-9

10-8

10-7

10-6

10-5

Table 4

ATRO (constant 10-4 M) with ACH added:

ACH Concentration % Change in Amplitude % Change in Frequency

10-10

10-9

10-8

10-7

10-6

10-5

Table 5


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Plot the following log-dose response curves using data from Tables 3, 4, and 5. If both ACH and ATRO with ACH experiments were performed, plot both curves on Graphs 1 and 2 making sure to clearly label each curve.

Graph 1 Graph 2

Graph 3 Graph 4




Questions

Only answer the questions that pertain to the recordings you performed.

1. How did temperature affect the amplitude and frequency of the contractions? Explain why temperature changes had the effect.

2. What was the affect of chemical depolarization? 3. What was the affect of each drug on the amplitude and frequency of the contractions? Explain why these

drugs had the effect. 4. How does ATRO alter or interfere with the ACH response? Can you suggest a molecular mechanism for

this alteration?


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Appendix 1:

Autoscaling data:

Graph data can be autoscaled vertically and horizontally for enhanced viewing.

Autoscale vertically by using the toolbar button or “Display > Autoscale Waveforms” to optimize the vertical display and allow closer examination of the waveform.

Autoscale horizontally by using the toolbar button or “Display > Autoscale Horizontal” to display the entire horizontal time scale in a single graph window.

Zooming in and out of data:

Use the Zoom tool to magnify portions of the waveform for a closer look.

Zoom in by selecting the toolbar icon and click/drag over the area of interest.

To zoom back, use Ctrl – (minus) or “Display > Zoom Back.”

Using the Event Palette to quickly locate Event Markers in the graph:

An easy way to locate event markers in the graph is by using the Event Palette, accessible by clicking the toolbar icon or via “Display > Show > Event Palette.”

Zoom in on any section of data, simply highlight an event in the Event list, and the portion of data corresponding to that event will be displayed in the graph. Other features in the Event Palette include summarizing, renaming, removing or editing existing events.

To navigate quickly through consecutive event markers, use the arrows to the left of the Event Palette toolbar icon.

Measurements used in BSL PRO Lesson A15:

Graph data measurements are taken by using the I-beam tool to select an area of interest. The following basic measurements are used in this experiment:

P-P (Peak-to-Peak) Shows the difference between the maximum amplitude value and the minimum amplitude value in the selected area.

Mean Shows the mean of all the data within the selected area.

Delta T Calculates the time interval between the start and end of the selected area.

BPM Calculates the frequency (in Beats/Pulses per minutes) from the selected time interval.

For a full explanation of the features described above, see the BSL PRO Tutorial or BSL PRO Software Guide.


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