Effects of Shear Rate, Temperature, and Blood Composition on Platelet Adhesion and Aggregation in Whole Blood of Thirteen-lined Ground Squirrels

Significant alterations in shear rate, temperature, and blood composition have been recorded in hibernating animals. These factors directly impact platelet clot formation, but it is currently unknown which factor has the greatest influence. To discover how each factor directly affects coagulation, capillary blood flow was replicated by use of a channeled microfluidic device. Flow rate was controlled with a syringe pump while temperature was controlled using stage heating and cooling techniques. Nonhibernating conditions were simulated using high flow rate at 35°C, and hibernating conditions were simulated using low flow rate at 15°C. Whole blood was run from both hibernating and non-hibernating thirteen-lined ground squirrels under both conditions. To allow for observation of blood flow and clot formation, microfluidic devices were placed on slides lined with a strip of collagen. As platelets flowed through the channels within the device, they adhered to the collagen strip. Slides were then fixed for further study of these clots. Flow of blood through the chambers was observed using an inverted phase contrast microscope and recorded on video. Clot formation over time was evaluated by use of these recordings. Platelets were also labeled with a fluorescent dye for analysis of the clots adhering to the collagen strip after flow. Clots were assessed using fluorescent microscopy to determine differences in morphology and total surface area under the different conditions. Preliminary results suggested that hibernating conditions (low shear rate and temperature) have a more prominent effect on the resistance of platelets to adhesion and aggregation than blood composition does. Further studies will investigate the effects of similar conditions on the blood of non-hibernating mammals to determine whether or not the observed response to cold temperature with low shear rate is a unique quality of hibernator blood