Name: a recovery cardiopulmonary bypass model without transfusion or inotropic agents in rats
Uploaded: 2018-03-23T13:00:00
Description: Watch this Scientific Journal Video about A Recovery Cardiopulmonary Bypass Model Without Transfusion or Inotropic Agents in Rats at JoVE.com

Appreciation is extended to Dr. T. Taki and Dr. M. Funamoto for their technical support.

Prior to experiment, all rats should be given one week to acclimate. All surgical procedures on animals should be carried out in accordance with the Guide for the Care and Use of Laboratory Animals (www.nap.edu/catalog/5140.html) or other appropriate ethical guidelines. Protocols should be approved by the animal welfare committee at the appropriate institution before proceeding. All subsequent procedures must be performed under aseptic conditions.&#160;
1. Preparing CPB Circuit
<p class="jov...

<ol>
	<li>Gibbon, J. H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Application+of+a+mechanical+heart+and+lung+apparatus+to+cardiac+surgery.">Application of a mechanical heart and lung apparatus to cardiac surgery.</a> Minn Med. 37 (3), 171-185 (1954).</li><li>Apostolakis, E., Filos, K. S., Koletsis, E., Dougenis, D. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Lung+dysfunction+following+cardiopulmonary+bypass.">Lung dysfunction following cardiopulmonary bypass.</a> J Cardiac Surg. 25 (1), 47-55 (2010).</li><li>Popovic, P., Horecky, J., Popovic, V. P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Instrumental+responses+in+rats+after+hypothermic+cardiopulmonary+by-pass.">Instrumental responses in rats after hypothermic cardiopulmonary by-pass.</a> P Soc Exp Biol Med. 126 (1), 225-228 (1967).</li><li>Fabre, O., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+recovery+model+of+partial+cardiopulmonary+bypass+in+the+rat.">A recovery model of partial cardiopulmonary bypass in the rat.</a> Perfusion. 16 (3), 215-220 (2001).</li><li>Hirao, S., Masumoto, H., Minatoya, K. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Rat+cardiopulmonary+bypass+models+to+Investigate+multi-organ+injury.">Rat cardiopulmonary bypass models to Investigate multi-organ injury.</a> Clin Surg. 2, 1-6 (2017).</li><li>Jungwirth, B., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Neurologic+outcome+after+cardiopulmonary+bypass+with+deep+hypothermic+circulatory+arrest+in+rats:+description+of+a+new+model.">Neurologic outcome after cardiopulmonary bypass with deep hypothermic circulatory arrest in rats: description of a new model.</a> J Thorac Cardiov Sur. 131 (4), 805-812 (2006).</li><li>de Lange, F., Yoshitani, K., Podgoreanu, M. V., Grocott, H. P., Mackensen, G. B. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+novel+survival+model+of+cardioplegic+arrest+and+cardiopulmonary+bypass+in+rats:+a+methodology+paper.">A novel survival model of cardioplegic arrest and cardiopulmonary bypass in rats: a methodology paper.</a> J Cardiothorac Surg. 3, 51 (2008).</li><li>Peters, S., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=An+experimental+model+of+myocardial+infarction+and+controlled+reperfusion+using+a+miniaturized+cardiopulmonary+bypass+in+rats.">An experimental model of myocardial infarction and controlled reperfusion using a miniaturized cardiopulmonary bypass in rats.</a> Interact Cardiovasc Th. 19 (4), 561-564 (2014).</li><li>Engels, M., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=A+cardiopulmonary+bypass+with+deep+hypothermic+circulatory+arrest+rat+model+for+the+investigation+of+the+systemic+inflammation+response+and+induced+organ+damage.">A cardiopulmonary bypass with deep hypothermic circulatory arrest rat model for the investigation of the systemic inflammation response and induced organ damage.</a> J Inflamm. 11 (26), (2014).</li><li>Pinto, A., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+extracellular+isoform+of+superoxide+dismutase+has+a+significant+impact+on+cardiovascular+ischaemia+and+reperfusion+injury+during+cardiopulmonary+bypass.">The extracellular isoform of superoxide dismutase has a significant impact on cardiovascular ischaemia and reperfusion injury during cardiopulmonary bypass.</a> Eur J Cardio-Thorac. 50 (6), 1035-1044 (2016).</li><li>Zhang, Z., Wu, Y., Zhao, Y., Xiao, X., Liu, J., Zhou, X. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Dynamic+changes+in+HMGB1+levels+correlate+with+inflammatory+responses+during+cardiopulmonary+bypass.">Dynamic changes in HMGB1 levels correlate with inflammatory responses during cardiopulmonary bypass.</a> Exp Ther Med. 5 (5), 1523-1527 (2013).</li><li>Kohno, T., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Impact+of+serum+high-mobility+group+box+1+protein+elevation+on+oxygenation+impairment+after+thoracic+aortic+aneurysm+repair.">Impact of serum high-mobility group box 1 protein elevation on oxygenation impairment after thoracic aortic aneurysm repair.</a> Heart Vessels. 26 (3), 306-312 (2011).</li><li>Tseng, C. C., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Impact+of+serum+biomarkers+and+clinical+factors+on+intensive+care+unit+mortality+and+6-month+outcome+in+relatively+healthy+patients+with+severe+pneumonia+and+acute+respiratory+distress+syndrome.">Impact of serum biomarkers and clinical factors on intensive care unit mortality and 6-month outcome in relatively healthy patients with severe pneumonia and acute respiratory distress syndrome.</a> Dis Markers. 2014, (2014).</li><li>Paparella, D., Yau, T. M., Young, E. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Cardiopulmonary+bypass+induced+inflammation:+pathophysiology+and+treatment.+An+update.">Cardiopulmonary bypass induced inflammation: pathophysiology and treatment. An update.</a> Eur J Cardio-Thorac. 21 (2), 232-244 (2002).</li><li>Hirao, S., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Recombinant+human+soluble+thrombomodulin+prevents+acute+lung+injury+in+a+rat+cardiopulmonary+bypass+model.">Recombinant human soluble thrombomodulin prevents acute lung injury in a rat cardiopulmonary bypass model.</a> J Thorac Cardiov Sur. , (2017).</li><li>Yamazaki, S., Inamori, S., Nakatani, T., Suga, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Activated+protein+C+attenuates+cardiopulmonary+bypass-induced+acute+lung+injury+through+the+regulation+of+neutrophil+activation.">Activated protein C attenuates cardiopulmonary bypass-induced acute lung injury through the regulation of neutrophil activation.</a> J Thorac Cardiov Sur. 141 (5), 1246-1252 (2011).</li><li>Wang, C. T., Zhang, L., Wu, H. W., Wei, L., Xu, B., Li, D. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Doxycycline+attenuates+acute+lung+injury+following+cardiopulmonary+bypass:+involvement+of+matrix+metalloproteinases.">Doxycycline attenuates acute lung injury following cardiopulmonary bypass: involvement of matrix metalloproteinases.</a> Int J Clin Exp Patho. 7 (11), 7460-7468 (2014).</li><li>Liu, K., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Curcumin+attenuates+cardiopulmonary+bypass-induced+lung+oxidative+damage+in+rats.">Curcumin attenuates cardiopulmonary bypass-induced lung oxidative damage in rats.</a> J Cardiovasc Pharm T. 17 (4), 395-402 (2012).</li><li>Taki, T., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Fetal+mesenchymal+stem+cells+ameliorate+acute+lung+injury+in+a+rat+cardiopulmonary+bypass+model.">Fetal mesenchymal stem cells ameliorate acute lung injury in a rat cardiopulmonary bypass model.</a> J Thorac Cardiov S. 153 (3), 726-734 (2017).</li><li>Zhu, X., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Establishment+of+a+novel+rat+model+without+blood+priming+during+normothermic+cardiopulmonary+bypass.">Establishment of a novel rat model without blood priming during normothermic cardiopulmonary bypass.</a> Perfusion. 29, 63-69 (2014).</li><li>Inoue, K., et al. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Deep+anesthesia+worsens+outcome+of+rats+with+inflammatory+responses.">Deep anesthesia worsens outcome of rats with inflammatory responses.</a> Inflamm Res. 65 (7), 563-571 (2016).</li><li>Bradfield, J. F., Schachtman, T. R., McLaughlin, R. M., Steffen, E. K. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Behavioral+and+physiologic+effects+of+inapparent+wound+infection+in+rats.">Behavioral and physiologic effects of inapparent wound infection in rats.</a> Lab Anim Sci. 42 (6), 572-578 (1992).</li></ol>

In this rat CPB model, the serum and lung expression levels of inflammatory cytokines and HMGB-1, a key transcription factor regulating the inflammatory responses, dramatically increased after CPB. Previous clinical studies showed that the serum secretion of HMGB-1 level is elevated in patients undergoing cardiovascular surgery11, and the peak serum HMGB-1 level during CPB was associated with more severe systemic inflammatory response syndrome and lung oxygenation impairment after CPB<sup class="x...

In 1953, Dr. John H. Gibbon Jr. successfully performed the first cardiac surgery using CPB1, and it subsequently became an essential modality in cardiovascular surgery. While the techniques and devices have been dramatically refined, multi-organ complications related to CPB still compromise the outcome of cardiovascular surgeries, and may affect postoperative morbidity and mortality2. CPB-related organ damage is caused by immune system activation and synthesis of proinflammatory cytokines, compliment activation, and production of oxygen free radicals2. Its pathophysiology, however, has not been fully elucidated.
Animal models recapitulating the clinical usage of CPB enable the clarification of the pathophysiological processes during and after CPB; this can facilitate pre-clinical studies in developing strategies to avoid these complications. Since Popovic et al. first reported a rat CPB model in 19673, rat CPB models have been refined, and have gradually taken the place of large-animal models due to greater cost-effectiveness, convenient experimental processes, and a plethora of testing methods in genetic and protein levels. Additionally, inbred rats can be genetically identical, reducing possible biological biases.
Fabre et al. first established a recovery model that allowed the study of the long-term multiple organ sequelae of CPB4. The advantages of this simple survival model are the flexibility (CPB flow and duration), stable vital condition, and reproducibility in systemic inflammation. Rat CPB models have become crucial for the investigation of therapeutic strategies that aim to prevent multi-organ injury during CPB5, and various models for simulating the clinical situations during CPB have recently been developed. De Lange et al. developed a cardiac arrest model, which can be used to characterize the enzymatic, genetic, and histological responses related to myocardial injury7. Peters et al. arranged myocardial infarction and controlled reperfusion using a miniaturized CPB model to analyze heart disfunction through the focal ischemia and reperfusion injury8. Jungwirth et al. first established a deep hypothermic circulatory arrest (DHCA) model, which can elucidate the global ischemia and reperfusion injury by DHCA and supports potential neuroprotective strategies6. Studies using DHCA investigate the influence of hypothermia, reperfusion, and/or hemolysis-triggered signaling events9. Deep hypothermia may affect the activation and inactivation of various enzymes and pathways and the mechanisms remain unknown10. On the other hand, cardiac arrest models or heart ischemia models must be used to investigate ischemia and reperfusion heart injury. These various rat CPB models that highly recapitulate human CPB may reveal pathological processes related to CPB and help mitigate CPB-related complications.
This protocol demonstrates a simple CPB model without transfusion or inotropic agents in a rat. This model allows for the study of long-term multiple organ sequelae of CPB.

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		<col />
		<col />
		<col />
	</colgroup>
	<tbody>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">Rodent Ventilator 7025</td>
			<td style="width:140px;">Ugo Basile</td>
			<td align="right" style="width:145px;">7025</td>
			<td style="width:167px;">Ventilator</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">OxiQuant B</td>
			<td style="width:140px;">ENVITEC</td>
			<td style="width:145px;">46-00-0023</td>
			<td>Oxygen Sensor</td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:216px;">CMA 450 Temperature Controller</td>
			<td style="width:140px;">CMA</td>
			<td align="right" style="width:145px;">8003759</td>
			<td>Temperature Controller</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">CMA 450 Heating Pad</td>
			<td style="width:140px;">CMA</td>
			<td align="right" style="width:145px;">8003763</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">CMA 450 Rectal Probe</td>
			<td style="width:140px;">CMA</td>
			<td align="right" style="width:145px;">8003761</td>
			<td></td>
		</tr>
		<tr height="42">
			<td height="42" style="height:42px;width:216px;">DIN(8) to Disposable BP Transducer</td>
			<td style="width:140px;">ADInstruments</td>
			<td style="width:145px;">MLAC06</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">Disposable BP Transducer</td>
			<td style="width:140px;">ADInstruments</td>
			<td style="width:145px;">MLT0670</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">IX-214 Data Recorder</td>
			<td style="width:140px;">iWorx Systems</td>
			<td style="width:145px;">IWX-214</td>
			<td>amplifier</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">LabScribe software</td>
			<td style="width:140px;">iWorx Systems</td>
			<td style="width:145px;"></td>
			<td>software</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">Roller pump</td>
			<td style="width:140px;">Furue Science</td>
			<td style="width:145px;">Model RP-VT</td>
			<td>pump</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">Happy Cath</td>
			<td style="width:140px;">Medikit</td>
			<td style="width:145px;">EB 19G 4HCLs PP</td>
			<td>17-gauge multiorifice angiocatheter</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">SURFLO ETFE I.V. Catheter</td>
			<td style="width:140px;">Terumo</td>
			<td style="width:145px;">SR-OX2419CA</td>
			<td>24-gauge angiocatheter</td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">Oxygenator</td>
			<td style="width:140px;">Mera</td>
			<td style="width:145px;">HPO-002</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">CPB circuit</td>
			<td style="width:140px;">Mera</td>
			<td style="width:145px;">custom-made</td>
			<td></td>
		</tr>
		<tr height="21">
			<td height="21" style="height:21px;width:216px;">Hespander fluid solution</td>
			<td style="width:140px;">Fresenius Kabi</td>
			<td style="width:145px;">3319547A4035</td>
			<td>Hydroxyethyl starch</td>
		</tr>
	</tbody>
</table>

a recovery cardiopulmonary bypass model without transfusion or inotropic agents in rats

Cardiopulmonary bypass (CPB) is indispensable in cardiovascular surgery. Despite the dramatic refinement of CPB technique and devices, multi-organ complications related to prolonged CPB still compromise the outcome of cardiovascular surgeries, and may worsen postoperative morbidity and mortality. Animal models recapitulating the clinical usage of CPB enable the clarification of the pathophysiological processes that occur during CPB, and facilitate pre-clinical studies to develop strategies protecting against these complications. Rat CPB models are advantageous because of their greater cost-effectiveness, convenient experimental processes, abundant testing methods at the genetic or protein levels, and genetic consistency. They can be used for investigating the immune system activation and synthesis of proinflammatory cytokines, compliment activation, and production of oxygen free radicals. The rat models have been refined and have gradually taken the place of large-animal models. Here, we describe a simple CPB model without transfusion and/or inotropic agents in a rat. This recovery model allows the study of the long-term multiple organ sequelae of CPB.

Figure 1&#160;shows the entire CPB circuit. The physiological variables in this model are shown in Figure 2, and include rectal temperature, mean arterial blood pressure, and heart rate. Figure 3 shows the arterial blood gas analyses during CPB, including partial pressure of arterial oxygen, partial pressure of arterial carbon dioxide, hematocrit, base excess, serum expression of potassium, and poten...

Watch this Scientific Journal Video about A Recovery Cardiopulmonary Bypass Model Without Transfusion or Inotropic Agents in Rats at JoVE.com

A Recovery Cardiopulmonary Bypass Model Without Transfusion or Inotropic Agents in Rats

Here, we present a protocol to describe a simple recovery cardiopulmonary bypass model without transfusion or inotropic agents in a rat. This model allows the study of the long-term multiple organ sequelae of cardiopulmonary bypass.

Cardiopulmonary bypass (CPB) is indispensable in cardiovascular surgery. Despite the dramatic refinement of CPB technique and devices, multi-organ ...

The overall goal of this procedure is to investigate multiple organ complications related to cardiopulmonary bypass in a rodent model without the use of transfusion or inotropic agents. This method for CPB in a small animal can help improve key operations in the cardiovascular cellular feed. Ultimately, use of this technique will help develop strategies to protect against complications encountered in cardiopulmonary bypass surgeries.First, set up the cardiopulmonary bypass circuit, connect the polyvinyl chloride tubes with the venous reservoir pre-designed for the CPB circuit and connect the tubes to a modified neonatal membrane oxygenator. Then, ensure that all of the connections are tight and do not leak. Next, on a height adjustable table, secure the circuit to a roller pump.Then, set the table's height to 10 centimeters below the plane of the experiment. Now, prime the circuit with 11 milliliters of the priming solution and start the pump roller gently rotating. Then, put an 18 gauge venting needle into the reservoir and heat the circuit using a heat lamp positioned over the reservoir.Next, tilt the membrane oxygenator and trigger it several times to de-air the line. This step is essential to preventing air embolisms and insufficient oxygenation. After anesthetizing and intubating the rat, transfer it to an operating table equipped with an electric heating pad.A respiratory rate of 70 cycles per minute and with 30%of inspired oxygen. Monitor the oxygen level using the oxygen sensor. To maintain the anesthesia, provide 1.5 to 2%isoflurane and administer pentobarbital at CPB initiation.Next, set up a rectal temperature probe and maintain a normal thermic body by adjusting the temperature of the heat pad and by heating the circuit with a lamp. Then, monitor the heart rate using an ECG attached at the bilateral shoulder and the left abdomen. Now, place moisture gauze on the rat's face to cover the eyes.Next, shave the hair over the bilateral inguinal region and right cervical region. Then, disinfect the skin using three alternating scrubs of 70%ethanol and betadine or chlorhexidine. The rat is now ready to be cannulated.Begin the surgery by making a five millimeter incision at the bilateral inguinal region and right cervical region using scissors. Then, bluntly dissect the tissues to expose the right main femoral artery. Next, carefully separate the artery from the vein and nerve.Then, ligate the end of the common femoral artery using 4-0 silk suture. Then, make a one millimeter perpendicular incision into the right femoral artery and carefully cannulate it using a 24 gauge intravenous catheter to a depth of one centimeter. Use this catheter to monitor systemic arterial pressure and analyze gas partial pressure in arterial blood.Next, administer heparin sodium using the catheter. Repeat this process to cannulate the left common femoral artery, and use this cannula as an arterial infusion line for the CPB circuit. Next, insert a 17 gauge multi-orifice angiocatheter into the right internal jugular vein, and advance it into the right atrium and inferior vena cava.Be gentle, as the vessel can be easily ruptured. Then, connect the angiocatheter to the CPB circuit for venous drainage. Finally, cover each cannulated region with moist gauze to avoid contamination.The animal is now ready for CPB. Before starting the CPB, begin delivering 100%oxygen gas to the oxygenator at 0.8 liters per minute and decrease the respiratory rate to 30 cycles per minute. Maintain the arterial oxygen partial pressure between 200 and 400 millimeters of mercury.Next, raise the heat pad temperature to 42 degrees celsius to mitigate the immediate drop in the body temperature after CPB initiation. Now, carefully start the CPB flow and keep an eye on the volume of blood in the reservoir. An empty reservoir may cause an air embolism, so adjust the pump flow by adjusting the table height.Do not reposition the venous drainage catheter to change the flow rate. Movement of this catheter might perforate the atrium or create an arrhythmia. Next, increase the pump flow up to 100 milliliters per kilogram per minute.Now, maintain the mean blood pressure at 70 millimeters of mercury. When the blood pressure is stable, a small one milliliter volume of blood in the reservoir is acceptable. Any less risks an air embolism.If the blood pressure does not stabilize, add two to three milliliters of priming solution to the circuit. When the body temperature returns to normal, reduce the heat pad temperature to 37 degrees celsius. After the CPB is completed, clamp the venous drainage tube and remove it from the circuit.Then, gradually infuse the remaining blood in the circuit to the artery to maintain the blood pressure. Next, increase the respiratory rate to 70 cycles per minute. Then, remove the venous drainage catheter and the left arterial catheter.Next, ligate the vessel in the proximal and distal site. Next, remove the arterial line from the right femoral artery. Then, clean each wound with saline and close them with sutures.As soon as spontaneous breathing begins, end the anesthesia and extubate the intratracheal tube. Then, administer warmed sterile isotonic fluids and use a heat mat and electrical heat lamp to keep the animal warm until it recovers from the anesthesia. Watch the animal carefully and provide respiratory support as needed.Arterial blood gas analyses during CPB include partial pressure of arterial oxygen, partial pressure of arterial carbon dioxide, hematocrit, base excess, serum expression of potassium, and potential of hydrogen. Throughout the procedure, the heart rate and mean arterial pressure were stable. The rectal temperature and hematocrit decreased at the beginning of CPB because of hemodilution caused by the priming volume.The partial pressure of arterial oxygen significantly increased during CPB due to membrane oxygenation. Hematoxylin and eosin staining of the lungs after CPB show interstitial edema, inflammatory cell infiltration, and hemorrhage in the CPB group as compared to the control group. In addition, serum concentration of tumor necrosis factor alpha, interleukin six and high mobility group box one were each augmented after CPB.After watching this video, you should have a good understanding of how to conduct a rough CPB procedure without transfusion or entropic agents. Once mastered, this technique can be done in two hours if it is performed properly. While attempting this procedure, it's important to remember to frequently check the hemodynamics of the rat.After its development, this technique paved the way for researchers in the field of cardiovascular surgery to explore inflammatory and other major organ complications in CPB.

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