STUDENT HANDOUT Modeling a microvascular network with tubing

Transcription

1 STUDENT HANDOUT Modeling a microvascular network with tubing Introduction The microvasculature system (the portion of the circulatory system composed of the smallest vessels, such as the capillaries, arterioles, and venules) is comprised a series of vessels and networks with extensive redundant connections. Figure 1: Branching of a typical microvasculature showing change of blood pressure, total area, and velocity. These extensive connections are necessary to provide a high surface area for blood flow to the various organs in the body. In addition if there is an occlusion in any of these vessels e.g. a blood clot, the redundant nature of the network allows blood flow to be rerouted with minimal damage to the organ. As a result of a blockage in the microvasculature one would expect a redistribution of flow in the neighboring vessels, a change in velocity and a change in pressure across the network. The objective of this experiment is to characterize a simple model of the microvascular system using tubes and connectors. You will measure flow velocities within the model network before and after a blockage and track the redistribution of flow when a channel is occluded. Figure 2 shows a schematic a network of pipes and connectors analogous to the vasculature in the brain. 1

2 inlet/source outlet Figure 2. Schematic of microvascular network that can be made with tubing and connectors Materials: Flexible tubing 1/16 ID Tygon lab tubing Cole Parmer Catalog# ZW Connectors Y-connectors Cole Parmer Catalog# ZW Beads Micrometer size beads - Duke Scientific Cat # 2010A (10 micron) or 2070A (70 micron). Place few drops in a solution of water and detergent (to keep them suspended). Binder Clip to block tubing Equipment: Pump Light microscope with camera Methods: 1. Use the tubing and connectors and create your own network. The inner diameter of the tube is 1/16. Take note of the length of each tube segment or channels as you are building your device (you ll need channel dimensions to calculate velocity). We ll assume that the Y-connectors also have an inner diameter of 1/ Draw a schematic of your network and hypothesize the network flow before and after a block in one of the channels. 3. Fill your syringe with bead solution, invert the syringe, tap on the sides, then gently squirt out any air bubbles that rose to the top. 4. Manually pump the bead solution through your network so that the solution covers the entire network. Place your syringe in the syringe pump. Start pumping at a flow rate of µl/min, until liquid is dripping out of the tubing. 2

3 5. Look at the channels underneath the microscope and make sure you can visualize beads. If the beads are moving too fast, decrease the flow rate. 6. You are expected to measure the velocity of the particles in each channel, some channels may have similar velocities. 7. Measure velocity by counting the number of beads that pass through a channel over a given amount of time. Or the time it takes for 1 bead to pass through the channel. You ll need to know the field of view of the microscope to calculate the velocity. You will be expected to track more than 10 particles per channel type in order to get statistically relevant data (i.e., to calculate the standard deviation). 8. When you have obtained the velocities for each channel, you will use the blocking tool to block 1 channel (binder clip). The purpose of blocking is to see a redirection of flow e.g. flow backwards and a change in velocity. Block the channel wait a minute until the network has reached a steady state. 9. Measure the velocity in each channel as well as note if there is redirection of flow. Measuring bead velocity using ImageJ Image J using Manual Tracking plugin Image J can be downloaded for free windows, download first choice (6MB) put in Program Files or Google Image J and follow download instructions. You also need to download an Manual Tracking plugin from Open Image J software Under File Import Image Sequence- the files of 1 series of data (e.g one set of channel data) then click OK. It helps if all the data is in a separate folder. Set the Scale. This converts pixels to microns. With the line tool measure across the channel (this is a known distance use the width of the channel) Analyzeset scale type in known distance type in umcheck global Note the conversion factor of pixels to microns Click on the image sequence you just opened Go to Plugins Manual tracking In the Parameters panel, set the time interval (the time interval between each picture frame), the xy scale (put in the conversion factor of pixel to microns this is 1/(value in set scale), and ignore the z scale To start a new track, click on Add track. Then click on the bead you are tracking in each frame as the program goes through the image sequence. After tracking one bead, click on Add track again to start tracking another bead. A track can be removed by selecting the appropriate track number in the central listing and clicking the Delete track n button. You want to keep a note of the order you track the beads in since Manual tracking puts all tracks (labeled with track number) in one excel file. Also note that the parameters are now hidden. To show parameters, tick the Show parameters option. 3

4 The resulting excel file contains the coordinates of each track and its velocity. Results and Data Analysis Expected. Average velocity for each channel before and after blockage Prediction of flow pattern after channel blockage. Include the schematic. If you expect a reversal of flow support with an image of flow reversal. Report: Please write your report in the following format. Organize your report into: 1. Abstract 2. Introduction 3. Results 4. Discussion 5. Questions Abstract: A synopsis of the purpose of the experiment, what method was used, and the results and conclusions obtained. (1 short paragraph) Introduction: State the objective of the experiment and discuss the importance of the subject and what other people are doing in this area (from supplemental information given). Results: Present the data/results in a logical manner with appropriate images and tables. Detailed calculations can go in an appendix. Discussion: Discuss your experiments and results, your limitations and assumptions you made. Discuss what your results mean in context of the field and what would it implications be in a more complex network Questions: 1. How would different pressures for each channel affect flow through the network? 2. What are some physiological examples of pressure changes our body undergoes? 3. Would a change in vessel diameter or vessel flow rate contribute to a greater change in pressure? What do you think our body does to compensate for changes in pressure? 4. What does a block in one channel do to network flow and direction? Is a blockage necessarily bad? Explain why it is or is not. 5. When a vessel branches, what will happen to the resistance? pressure drop? Relate this to the picture shown in the introduction graphing pressure from arteries to the capillary beds Useful References 4

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