Name: adult and pediatric porcine model of acute volume overload
Uploaded: 2024-01-12T13:40:00.000+00:00
Duration: 6 min 9 s
Description: Watch this Scientific Journal Video about Author Spotlight: Exploring Venous Waveforms in Porcine Models to Tackle Volume Overload in Medicine at JoVE.com

The authors would like to thank Dr. Jos&#233; A. Diaz, Jamie Adcock, and Mary Susan Fultz and the S.R. Light Laboratory at Vanderbilt University Medical Center for assistance and support. Another special thanks to John Poland and the rest of the Vanderbilt University Medical Center perfusionists and their students for their help with this study. This work was supported by a grant from the National Heart, Lung, and Blood Institute of the National Institutes of Health (BA; R01HL148244). The content is the sole responsibility of the authors and does not necessarily represent the official views of the National Institutes of Health.

The study protocol was approved by the Vanderbilt University Institutional Animal Care and Use Committee (protocol M1800176-00) and strictly adhered to the National Institute of Health Guidelines for the Care and Use of Laboratory Animals. Male and Female Yorkshire pigs and piglets weighing approximately 40-45 kg and 4-10 kg are used in this experiment. The present approach does not encompass a screening for preexisting medical conditions in the ordered swine. Acknowledging that this practice could potentially influence or obscure the desired results, it is essential to note that, as per the vendor&#39;s information, the likelihood of such interference is low. The limitation is acknowledged and accepted as an inherent aspect of the procedure.
1. Anesthesia and ventilation
<ol>
	<li>Adult porcine model
	<ol>
		<li>Anesthetize the pig by slowly injecting ketamine (2.2 mg/kg)/xylazine (2.2 mg/kg)/telazol (4.4 mg/kg) intramuscularly (IM). Immediately after induction, an 18-24G intravenous (IV) catheter is placed in the central or marginal ear vein on the posterior side of the auricle. Secure the IV with 1-inch adhesive tape.</li>
		<li>Place the pig onto the operating room table, in the supine position. Ask an independent animal laboratory technician, responsible for overseeing specific parameters, to assess anesthesia depth determined by factors including vital signs, responsiveness to stimuli, presence or absence of movement, jaw tone laxity, fluctuations in heart rate, changes in end-tidal CO2 levels, and variations in respiratory rate. These assessments guide adjustments to the inhaled anesthetic dosage.</li>
		<li>Endotracheally intubate the pig with a 6.5 mm endotracheal tube, through direct laryngoscopy, and inflate the endotracheal cuff with 3-5 mL of air. Maintain the pig on volume-controlled ventilation with a tidal volume of 8 mL/kg, respiratory rate titrated to an end-tidal CO2 of 35-40 mmHg, and positive end-expiratory pressure of 5 cm H2O. Maintain anesthesia through the inhalation of 1% isoflurane.</li>
		<li>Place a Foley catheter in the urethra to monitor the urine output volume in female pigs and surgical place it in male pigs secondary to the anatomical difficulty of Foley placement through the urethra.</li>
	</ol>
	</li>
	<li>Piglet model
	<ol>
		<li>Inject anesthesia solution of ketamine (2.2 mg/kg)/xylazine (2.2 mg/kg)/telazol (4.4 mg/kg)/dexmedetomidine (0.005 mg/kg) into an approximately 5-week-old piglet (equivalent to approximately 12-month-old human) via an intramuscular (IM) injection. Then, immediately place a 22-24G IV in the best available vein, likely on the posterior side of the auricle.</li>
		<li>Place piglet on the operating room table, in the supine position.</li>
		<li>Using direct laryngoscopy, endotracheally intubate the piglet with a 4.5-5.5 mm endotracheal tube. Inflate the endotracheal tube cuff with 3-5 mL of air using a syringe without a needle attached to it. Maintain anesthesia with 1% isoflurane with or without re-administration of dexmedetomidine (0.005 - 0.01 mg/kg IV) every 2 h as needed based on the depth of anesthesia appreciated at the time of re-dose.</li>
		<li>Maintain with volume-controlled ventilation a tidal volume of 8 mL/kg, respiratory rate titrated to an end-tidal CO2 of 35-40 mmHg, and a positive end-expiratory pressure of 5 cm H2O.</li>
		<li>Place a Foley catheter in the urethra to monitor urine output volume in female piglets. Place the Foley catheter surgically&#160;in male piglets secondary to the anatomical difficulty of Foley placement through the urethra. 
		&#8203;NOTE: Additionally, buprenorphine/dexmedetomidine analgesic administration will occur via bolus administration as deemed necessary. To maintain consistency, the respiratory rate on the mechanical ventilator will be adjusted to keep the end-tidal CO2 level within the range of 35-40 mmHg throughout the experiment.</li>
	</ol>
	</li>
</ol>
2. Cannulation and monitoring device placement
<ol>
	<li>Adult porcine model
	<ol>
		<li>Disinfect the entire anterior neck with 2% chlorohexidine scrub solution and follow with a spray of 5% povidone-iodine solution14.</li>
		<li>Surgically expose both right and left external jugular (EJ) veins and internal carotid arteries (CA) with bilateral vertical incisions immediately lateral to the trachea, and dissect down to the vasculature with monopolar cautery.</li>
		<li>Dissect the strap muscles and tract as needed using Kelly tissue scissors and Lahey retractors and/or tissue forceps14. Expose bilateral EJ and CAs.</li>
		<li>Place two 8.5 French (Fr) cannula into the right EJ using the Seldinger technique15. Once cannulated, place a 7 Fr pulmonary artery catheter (PAC) through the introducer of the right EJ. Use this right EJ catheter and PAC for hemodynamic monitoring.</li>
		<li>Cannulate the left EJ with a 10 Fr cannula and connect it to a dedicated roller pump tubing primed with PlasmaLyte solution. 
		NOTE: The external jugular veins tend to be of larger diameter and more appropriately angled for heart catheterization. It is for these reasons we chose to cannulate the EJ over the internal jugular (IJ) in the porcine experiments1.</li>
		<li>Using the Seldinger technique15 place a 4 Fr arterial line in the right CA for invasive blood pressure monitoring throughout the experiment.</li>
		<li>Attach the desired monitoring to pig.
		<ol>
			<li>Monitor the heart rate (HR) with telemetry leads and the systolic blood pressure (SBP), diastolic blood pressure (DBP), and mean arterial pressure (MAP) by connecting a pressure transducer attached to a blood pressure amplifier to the CA catheter.</li>
			<li>Monitor the mean pulmonary artery pressure (MPAP), systolic pulmonary artery pressure (SPAP), diastolic pulmonary artery pressure (DPAP), and central venous pressure (CVP) by attaching a pressure transducer attached to a blood pressure amplifier to the appropriate PAC ports.</li>
			<li>Determine pulse pressure by finding the variance between SBP and DBP. To calculate pulse pressure variability (PPV), compute the variation between peak pulse pressure levels during both inspiration and expiration throughout the respiratory cycle.</li>
			<li>Calculate PPV measurements using LabChart 8&#39;s Blood Pressure Module and running on a PowerLab system. In this setting, select 3 min of data in the Arterial Line channel where the minimum peak height is set to 10 mmHg and cycles averaged over 10 cycles. In the software&#39;s module the peaks of each pulse cycle can be automatically calculated and visually confirmed. The resulting minimum pulse pressure and maximum pulse pressure are then used to calculate PPV.</li>
			<li>Carry out thermodilution cardiac output (CO) by using device specific volume/temperature calibration. Obtain pulmonary capillary wedge pressure (PCWP) by inflating the PAC balloon with 1.5 mL of air and advancing the catheter until there is visualization of both V and A waves, representing restricted right to left blood flow. Read the PCWP at the value of the A wave at end expiration.</li>
		</ol>
		</li>
		<li>Administer PlasmaLyte16 at a rate of approximately 100 mL/min to obtain a starting PCWP of 8-10 mmHg (euvolemia) before the initiation of acute volume overload. 
		NOTE: The total volume needed to achieve euvolemia varies based on a multitude of variables covered in the Discussion. Approximately 500 mL is needed, on average, in pigs undergoing this experimental protocol at Vanderbilt University Medical Center. This model uses PlasmaLyte as the balanced buffered crystalloid solution. It is likely that any other balanced buffered crystalloid (e.g., Normosol-R, Lactated Ringer&#39;s) would offer similar results. Non-buffered, acidic normal saline is and should be avoided in this model to avoid the known loss of endothelial cell membrane integrity, endothelial dysfunction, and acidosis caused by normal saline16.</li>
	</ol>
	</li>
	<li>Piglet model
	<ol>
		<li>Similar to adult pigs, piglets, once anesthetized and mechanically ventilated, disinfect across their entire anterior neck with 2% chlorohexidine scrub solution and follow with a spray of 5% povidone-iodine solution14. Only cannulate the right EJ, carotid, and left femoral arteries in piglets.</li>
		<li>Surgically expose the right EJ vein and internal CA with a right sided vertical incision immediately lateral to the trachea and dissect down to the vasculature with monopolar cautery.</li>
		<li>Dissect the strap muscles and tract as needed using Kelly tissue scissors and Lahey retractors and/or tissue forceps14. Expose the right sided EJ and CA.</li>
		<li>Disinfect the lower abdomen and pubic area of the piglet with 2% chlorohexidine scrub solution and follow with a spray of 5% povidone-iodine solution. Surgically expose the left femoral artery (FA) with a classic longitudinal technique as described in17.</li>
		<li>Place a 6 Fr central venous catheter introducer into the right EJ, followed by placement of a 5 Fr PAC into the pulmonary artery.</li>
		<li>Position two 3 Fr arterial catheters: one in the right CA and the other in the left FA. Dedicate the left FA catheter to drawing blood for frequent arterial blood gas analysis. Use the open port associated with the PAC introducer for volume administration with a 60 mL syringe.</li>
		<li>Administer a PlasmaLyte16 bolus of 10 mL/kg using a 60 mL syringe at a steady push rate, stopping after each bolus to obtain a PCWP and stop once a value of 8-10 mmHg (euvolemia) is achieved. 
		&#8203;NOTE: The total volume needed to achieve euvolemia varies based on a multitude of variables covered in the Discussion of this manuscript. Approximately 50-100 mL is needed, on average, in the piglets undergoing this experimental protocol at Vanderbilt University Medical Center.</li>
	</ol>
	</li>
</ol>
3. Volume administration
<ol>
	<li>Adult porcine model
	<ol>
		<li>Once cannulation is complete and euvolemia achieved, infuse warm PlasmaLyte crystalloid solution in 500 mL increments at a rate of 100 mL/min (Figure 1).</li>
		<li>Confirm the recording of hemodynamic endpoints: HR, fraction of oxygen-saturated hemoglobin (SpO2), respiratory rate (RR), end-tidal carbon dioxide (ETCO2), CVP, SBP, DBP, MAP, PPV, SPAP, DPAP, and MPAP.</li>
		<li>Perform necessary procedures to obtain the static measures (CO and PCWP) after every 500 mL of volume until euthanasia, which will take place at 5 L total volume or until a 15% CO decrease from previous measurement exists, whichever occurs first. 
		NOTE: The drop in CO represents the start of the descending limb of the Starling curve18. At this point, volume overload leads to dilation of the heart past the optimum length for muscle fiber contraction, resulting in impaired contraction and decreased CO18.</li>
		<li>At euvolemia, and at the end of total volume administration, perform an arterial blood gas analysis to obtain the partial arterial oxygen pressure (PaO2), pH, lactate, and base excess of the pig.</li>
		<li>Record the urine output (mL) after each 500 mL increment of PlasmaLyte crystalloid solution. It is advisable to zero out the urine once euvolemia is achieved. Euthanize the pig either at a total volume of 5 L or when a 15% decrease in CO is observed, whichever occurs first.</li>
	</ol>
	</li>
	<li>Piglet model
	<ol>
		<li>Following successful cannulation and attainment of euvolemia, administer PlasmaLyte in increments of 20 mL/kg via syringe bolus every 10 min (Figure 1).</li>
		<li>Confirm the recording of hemodynamic parameters (HR, RR, SpO2, EtCO2, CVP, SBP, DBP, MPAP, PPV, and MPAP). Measure PCWP after each 10 mL/kg bolus. 
		NOTE: Owing to the volume required and the resistance created by the small internal diameter of the 5 Fr PAC, thermodilution CO is not performed in the piglets. Instead, the Fick method19,20 is employed to calculate the CO. This involves obtaining a fraction of oxygen-saturated hemoglobin from the pulmonary artery blood (SvO2), which is performed concurrently with arterial blood gas analysis.</li>
		<li>Perform arterial blood gas analysis after each 20 mL/kg volume bolus to obtain the PaO2, pH, lactate, and base excess. 
		NOTE: Given the limitations of many of these invasive data points in routine clinical care, transthoracic echocardiography (TTE) is performed after each 20 mL/kg bolus in the piglet model to measure the aortic blood flow Peak Systolic Velocity (PSV) and Left Ventricular Outflow Tract (LVOT) diameters-two data points used in pediatric clinical practice to estimate a patient&#39;s volume state.</li>
		<li>Perform TTE to measure PSV and LVOT diameter data points after each 20 mL/kg bolus. Record urine output after each 20 mL/kg bolus. Euthanize the piglet either at a total volume of 500 mL or when a 15% decrease in CO is observed, whichever occurs first.</li>
	</ol>
	</li>
</ol>
4. Euthanasia for both adult pigs and piglets
<ol>
	<li>Confirm maintenance of 1% isoflurane. Induce cardiac arrest by IV injection of sodium pentobarbital (125 mg/kg). Confirm lack of vital signs post injection to confirm demise.</li>
</ol>

Investigating Physiological Responses to Acute Volume Overload in Porcine Models

<ol>
	<li>Wise, E. 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=Hemodynamic+parameters+in+the+assessment+of+fluid+status+in+a+porcine+hemorrhage+and+resuscitation+model.">Hemodynamic parameters in the assessment of fluid status in a porcine hemorrhage and resuscitation model.</a> Anesthesiology. 134 (4), 607-616 (2021).</li><li>Rollins, K. E., Lobo, D. N. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Intraoperative+goal-directed+fluid+therapy+in+elective+major+abdominal+surgery:+A+meta-analysis+of+randomized+controlled+trials.">Intraoperative goal-directed fluid therapy in elective major abdominal surgery: A meta-analysis of randomized controlled trials.</a> Ann Surg. 263 (3), 465-476 (2016).</li><li>Som, A., Maitra, S., Bhattacharjee, S., Baidya, D. K. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Goal+directed+fluid+therapy+decreases+postoperative+morbidity+but+not+mortality+in+major+non-cardiac+surgery:+a+meta-analysis+and+trial+sequential+analysis+of+randomized+controlled+trials.">Goal directed fluid therapy decreases postoperative morbidity but not mortality in major non-cardiac surgery: a meta-analysis and trial sequential analysis of randomized controlled trials.</a> J Anesth. 31 (1), 66-81 (2017).</li><li>Xu, 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=Goal-directed+fluid+therapy+versus+conventional+fluid+therapy+in+colorectal+surgery:+A+meta+analysis+of+randomized+controlled+trials.">Goal-directed fluid therapy versus conventional fluid therapy in colorectal surgery: A meta analysis of randomized controlled trials.</a> Int J Surg. 56, 264-273 (2018).</li><li>Hassinger, A. B., Wald, E. L., Goodman, 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=Early+postoperative+fluid+overload+precedes+acute+kidney+injury+and+is+associated+with+higher+morbidity+in+pediatric+cardiac+surgery+patients.">Early postoperative fluid overload precedes acute kidney injury and is associated with higher morbidity in pediatric cardiac surgery patients.</a> Pediatr Crit Care Med. 15 (2), 131-138 (2014).</li><li>Chen, 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=A+global+view+of+porcine+transcriptome+in+three+tissues+from+a+full-sib+pair+with+extreme+phenotypes+in+growth+and+fat+deposition+by+paired-end+RNA+sequencing.">A global view of porcine transcriptome in three tissues from a full-sib pair with extreme phenotypes in growth and fat deposition by paired-end RNA sequencing.</a> BMC Genomics. 12, 448 (2011).</li><li>Hou, N., Du, X., Wu, S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Advances+in+pig+models+of+human+diseases.">Advances in pig models of human diseases.</a> Animal Model Exp Med. 5 (2), 141-152 (2022).</li><li>Spannbauer, 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=Large+animal+models+of+heart+failure+with+reduced+ejection+fraction+(HFrEF).">Large animal models of heart failure with reduced ejection fraction (HFrEF).</a> Front Cardiovasc Med. 6, 117 (2019).</li><li>Odle, J., Lin, X., Jacobi, S. K., Kim, S. W., Stahl, C. H. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+suckling+piglet+as+an+agrimedical+model+for+the+study+of+pediatric+nutrition+and+metabolism.">The suckling piglet as an agrimedical model for the study of pediatric nutrition and metabolism.</a> Annu Rev Anim Biosci. 2, 419-444 (2014).</li><li>Whitaker, E. E., 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+novel,+clinically+relevant+use+of+a+piglet+model+to+study+the+effects+of+anesthetics+on+the+developing+brain.">A novel, clinically relevant use of a piglet model to study the effects of anesthetics on the developing brain.</a> Clin Transl Med. 5 (1), 2 (2016).</li><li>Gasthuys, E., 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+potential+use+of+piglets+as+human+pediatric+surrogate+for+preclinical+pharmacokinetic+and+pharmacodynamic+drug+testing.">The potential use of piglets as human pediatric surrogate for preclinical pharmacokinetic and pharmacodynamic drug testing.</a> Curr Pharm Des. 22 (26), 4069-4085 (2016).</li><li>Alobaidi, R., 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=Association+between+fluid+balance+and+outcomes+in+critically+ill+children:+A+systematic+review+and+meta-analysis.">Association between fluid balance and outcomes in critically ill children: A systematic review and meta-analysis.</a> JAMA Pediatr. 172 (3), 257-268 (2018).</li><li>Soler, Y. A., Nieves-Plaza, M., Prieto, M., Garcia-De Jesus, R., Suarez-Rivera, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Pediatric+risk,+injury,+failure,+loss,+end-stage+renal+disease+score+identifies+acute+kidney+injury+and+predicts+mortality+in+critically+ill+children:+a+prospective+study.">Pediatric risk, injury, failure, loss, end-stage renal disease score identifies acute kidney injury and predicts mortality in critically ill children: a prospective study.</a> Pediatr Crit Care Med. 14 (4), e189-e195 (2013).</li><li>Alessa, M. 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=Porcine+as+a+training+module+for+head+and+neck+microvascular+reconstruction.">Porcine as a training module for head and neck microvascular reconstruction.</a> J Vis Exp. (139), e58104 (2018).</li><li>Higgs, Z. C., Macafee, D. A., Braithwaite, B. D., Maxwell-Armstrong, C. A. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+Seldinger+technique:+50+years+on.">The Seldinger technique: 50 years on.</a> Lancet. 366 (9494), 1407-1409 (2005).</li><li>Cheung-Flynn, J., 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=Normal+saline+solutions+cause+endothelial+dysfunction+through+loss+of+membrane+integrity,+ATP+release,+and+inflammatory+responses+mediated+by+P2X7R/p38+MAPK/MK2+signaling+pathways.">Normal saline solutions cause endothelial dysfunction through loss of membrane integrity, ATP release, and inflammatory responses mediated by P2X7R/p38 MAPK/MK2 signaling pathways.</a> PLoS One. 14 (8), e0220893 (2019).</li><li>Canteras, M., Baptista-Silva, J. C. C., Cacione, D. G. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Transverse+versus+longitudinal+inguinotomy+for+femoral+artery+approach.">Transverse versus longitudinal inguinotomy for femoral artery approach.</a> Cochrane Database Syst Rev. 2018 (10), CD013153 (2018).</li><li>Katz, A. M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+descending+limb+of+the+Starling+curve+and+the+failing+heart.">The descending limb of the Starling curve and the failing heart.</a> Circulation. 32 (6), 871-875 (1965).</li><li>Ragosta, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=.">.</a> Textbook of Clinical Hemodynamics. , (2017).</li><li>LaFarge, C. G., Miettinen, O. S. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=The+estimation+of+oxygen+consumption.">The estimation of oxygen consumption.</a> Cardiovasc Res. 4 (1), 23-30 (1970).</li><li>Soto, F., Kleczka, J. F. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Cardiopulmonary+hemodynamics+in+pulmonary+hypertension:+Pressure+tracings,+waveforms,+and+more.">Cardiopulmonary hemodynamics in pulmonary hypertension: Pressure tracings, waveforms, and more.</a> Adv Pulmonary Hypertension. 7 (4), 386-393 (2008).</li><li>Helen Chum, C. P. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Endotracheal+intubation+in+swine.">Endotracheal intubation in swine.</a> Lab Anim. 41 (11), 309-311 (2012).</li><li>Cavallaro, F., Sandroni, C., Antonelli, M. <a target="_blank" href="http://www.ncbi.nlm.nih.gov/entrez/query.fcgi?db=PubMed&cmd=Search&doptcmdl=Citation&defaultField=Title+Word&term=Functional+hemodynamic+monitoring+and+dynamic+indices+of+fluid+responsiveness.">Functional hemodynamic monitoring and dynamic indices of fluid responsiveness.</a> Minerva Anestesiol. 74 (4), 123-135 (2008).</li></ol>

No disclosures specific to the topics of this report. Kyle Hocking, PhD, is Founder, CEO and President of VoluMetrix and an inventor on intellectual property in the field of venous waveform analysis assigned to Vanderbilt University and licensed to VoluMetrix. Colleen Brophy, MD, is Founder and CIO of VoluMetrix and an inventor of intellectual property in the field of venous waveform analysis assigned to Vanderbilt and licensed to VoluMetrix. Bret Alvis, MD, CMO and is an inventor on intellectual property in the field of venous waveform analysis assigned to Vanderbilt and licensed to VoluMetrix and is married to the COO of VoluMetrix. The remaining authors have no disclosures to report.

There are two critical steps in this protocol. First, it is imperative that time is taken to obtain appropriate cannulation and ensure the positioning of hemodynamic/volume monitoring. In both adult and piglet models, surgical cutdown is necessary to cannulate the required vessel appropriately and introduce the required catheter. Percutaneous, ultrasound guided approaches have proven challenging and traumatic around the small caliber vessels seen in pigs and piglets. Two catheters that can present a challenge are the PAC and Foley urinary catheter. PACs can be difficult to float into the pulmonary artery in both pigs and piglets. When the balloon tip is inflated with 1.0 - 1.5 mL of air, it can migrate down the inferior vena cava (IVC) instead of going into the right atrium (RA). Minor retraction of the introducer can aid the PAC enter the RA. Fluoroscopy and ultrasound techniques have been used to aid PAC placement. Once the heart has been cannulated with the PAC, floating into the PA has not proven to be challenging. The other major point of pain is the placement of a Foley urinary catheter in a male pig/piglet bladder via the urethral tract due to the anatomical complexities of the porcine urinary system. Specifically, male pigs have a long, narrow, and highly coiled urethra, which can make the insertion and navigation of the catheter difficult and increase the risk of trauma or misplacement. Therefore, surgical placement of the Foley catheter is typically performed in male pigs/piglets secondary to this anatomical difficulty.
The second critical step in the experimental protocol involves ensuring euvolemia in both pig and piglet models prior to initiating volume overload. Establishing baseline measurements is essential for subsequent comparisons and accurate determination of volume overload. It is frequently observed that both adult pigs and piglets often present in a state of dehydration or volume down at the onset of experimentation. This could be attributed to factors such as transit, diet, and/or hydration protocols implemented during housing. Consequently, it is not uncommon for animals to require resuscitation before initiation of the volume overload model. Based on experience and current data a PCWP between 8-10 mmHg is the most representative data point for euvolemia in pig and piglet models, a criterion that aligns with existing literature1.
A major limitation is the CO variable and how it is obtained in both the models. While it is a critical parameter in cardiovascular physiology, the two techniques utilized are thermodilution and the Fick method19,20. Thermodilution involves the injection of a known quantity of calibrated temperature saline into the RA, and the subsequent measurement of temperature changes in the pulmonary artery by the thermistor at the tip of the PAC. The degree of dilution is inversely proportional to the CO. In contrast, the Fick method calculates CO based on oxygen consumption (VO2) and the difference in oxygen content between arterial and venous blood20. However, both methods have significant limitations. Thermodilution requires the injection velocity to remain constant, which is prone to human error, requires an appropriately functioning PAC thermistor, and is less accurate in conditions of significant intracardiac or intrapulmonary shunting or tricuspid regurgitation. In addition, specific to the piglet model, the CO monitor used must be calibrated to measure a much smaller volume than the adult CO monitors. In addition, the constant velocity of injection can prove difficult with small lumen diameters seen with a 5 Fr PAC. Therefore, the Fick method is more appropriate for a piglet model. While this is less invasive, its limitation is that it assumes a steady state of oxygen consumption, which may not be accurate for piglets experiencing such rapid physiological changes. For example, the SvO2 is a key variable of the Fick CO, and is influenced by the hemoglobin of the piglet. The acute dilution of the piglet&#39;s hemoglobin is likely significantly contributing to the fall in SvO2 and subsequently causing a calculated CO to drop which may not entirely be a true L/min reduction. Furthermore, accurate measurement of oxygen consumption can be technically challenging because aspiration from the tip of the PAC is prone to clot burden throughout experimentation.
Another challenge in the porcine volume overload model is the difficulty in obtaining an appropriate waveform for PCWP assessment. PCWP monitoring is a crucial component of comprehensive volume overload evaluation. However, to ensure the accuracy of PCWP tracing, certain quality parameters must be satisfied before recording wedge tracing. First, the pulmonary artery pressure (PAP) waveform should exhibit an appropriate change, reflecting restricted right-to-left blood flow. This change is the appropriate loss of systolic and diastolic pressures form the right ventricular and a PCWP tracing displaying both A and V waves, indicative of left atrial and ventricular contractions, respectively21,22. Finally, the PCWP value should be derived from the A wave at the end expiration21. A valid PCWP can only be reported when these three conditions are satisfied, thereby ensuring the reliability of the measurement in the context of volume overload assessment. Finally, PPV and systolic pressure variation (SPV; not reported in preliminary results) are the only dynamic circulatory indices evaluated in this model. Dynamic indices can be valuable datapoints in volume responsiveness and are based on the response of the circulatory system to a controlled preload variation, often present secondary to positive pressure ventilation and/or leg raising. The dynamic changes seen in the arterial waveforms were analyzed and PPV did demonstrate a statistically significant inverse correlation to volume given in the pig, the same results were not appreciated in the piglet. The SPV will be reported in future publications. Of note, plethysmograph variability index (PVI) and stroke volume variation (SVV) are not investigated in this model despite having some evidence supporting their use in specific resuscitation scenerios23. This limitation is secondary to monitoring capabilities in our current experimental set-up.
The development and utilization of both adult pig and piglet models of volume overload have significant implications in various research areas. Adult pigs, owing to their physiological similarities to humans1,9,10, particularly in cardiovascular function, serve as excellent models for studying the pathophysiology of volume overload in adult human patients. This model can provide valuable insights into the mechanisms underlying conditions such as heart failure and fluid overload in critical illness and can aid in the evaluation of therapeutic strategies and monitoring techniques in these settings.
Piglet models offer a unique opportunity to investigate the effects of volume overload in the pediatric population - a population in which clinical studies have proven challenging. Given the physiological and developmental differences between children and adults, the findings from adult models cannot always be extrapolated to pediatric patients. The representative results in this report are an example of this faulty, but traditional approach. Piglets, with their developmental similarities to human infants, can help bridge this gap9,10. This is particularly relevant given the high incidence of volume overload conditions in pediatric patients, such as those related to congenital heart diseases or intensive care interventions12. The piglet model can contribute to our understanding of the pediatric-specific pathophysiology of acute volume overload and aid in the development of age-appropriate therapeutic and monitoring strategies.

Acute volume overload, a condition characterized by an abrupt and excessive increase in body fluid volume, is a critical medical concern that warrants comprehensive study1. It is often associated with aggressive and/or inappropriate fluid resuscitation, blood transfusion, and comorbidities such as heart failure and renal failure. It can lead to severe morbidity and an increased likelihood of mortality1,2,3. Despite its clinical significance, the pathophysiology of acute volume overload remains poorly understood3,4. Furthermore, the lack of specific diagnostic criteria and effective monitoring strategies further underscores the need for rigorous scientific investigation. Studying acute volume overload is not only crucial for improving patient outcomes but also for advancing our understanding of human physiology. It provides a unique opportunity to explore the body&#39;s fluid homeostasis mechanisms and their responses to extreme stress1. Studies investigating goal-directed fluid therapy (GDFT) to prevent liberal fluid resuscitation and promote a more goal-directed resuscitation approach have demonstrated improved morbidity and mortality in perioperative settings and in sepsis1,3,4. These studies used a variety of devices to monitor the volume state, including central venous catheters with central venous pressure measurements, ScVO2, arterial line lactate measurements, stroke volume/cardiac output measurements through transesophageal Doppler, lithium dilution cardiac output, arterial pulse contour analysis, thoracic electrical bioimpedance, and transpulmonary thermodilution1,3,4,5. The multiple approaches utilized to assess volume status, each with limitations in accuracy and usability, suggest that there is room for significant improvement in GDFT by enhancing intravascular volume assessment3,4.
Porcine models have emerged as particularly valuable tools in the study of human cardiovascular physiology6. The anatomical and physiological similarities between porcine and human cardiovascular systems, such as heart size, coronary anatomy, and hemodynamic parameters, make pigs ideal models for translational research6. Furthermore, pigs exhibit a comparable response to volume overload as humans, making them excellent models for studying the pathophysiology of acute volume overload and the effectiveness of various therapeutic interventions7,8. The use of porcine models also allows for the collection of high-quality, detailed data points, such as real-time hemodynamic measurements and tissue samples, which are often unattainable in human studies7. This superiority of data points provides a more comprehensive understanding of acute volume overload, which could ultimately contribute to the development of more effective monitoring and prevention strategies.
The use of piglet models in studying acute volume overload is of paramount importance, particularly given the scarcity of pediatric research in this field. Piglets, with their physiological and developmental similarities to human infants, provide an invaluable model, like their adult counterparts, for understanding the pediatric population9,10,11. Despite the high incidence of volume overload conditions in pediatric patients, such as those related to congenital heart diseases or intensive care interventions, research in this area has been markedly limited, especially when it comes to animal models that accurately represent human infants5,12,13. Utilizing piglet models can help bridge this gap, offering insight into the pediatric-specific pathophysiology of acute volume overload and the efficacy of potential therapeutic strategies7,11.
This manuscript describes a method of using a continuous infusion of crystalloid solution directly into the external jugular vein of both adult and pediatric pigs to induce acute volume overload and to study the hemodynamic effects of such volume changes on common peripheral and central data points used in volume status monitoring. This outlined method should serve as a valuable tool to help future scientists investigate the underlying pathophysiological mechanisms of acute volume overload and evaluate potential superior monitoring modalities and innovations.

<table><tbody><tr><td>1% Isoflurane</td><td>Primal, Boston, MA, USA</td><td>26675-46-7</td><td>https://www.sigmaaldrich.com/US/en/product/aldrich/792632?gclid=Cj0KCQjw9fqnBhDSARIsAHl&lt;br/&gt; cQYS_W-q6tS2s6LQw2Qn7Roa3TGIpTLPf5&lt;br/&gt; 2351vrhgp44foEcRozPqtYaAtvfEAL&lt;br/&gt; w_wcB</td></tr><tr><td>Arterial Catheter</td><td>Merit Medical, South Jordan, UT, USA</td><td>MAK401</td><td>MAK Mini Access Kit 4F</td></tr><tr><td>Arterial Catheter</td><td>Cook Medical, Bloomington, IN, USA</td><td>C-PMS-300-RA/G01908</td><td>Radial Artery Catheter Set 3.0Fr./5cm</td></tr><tr><td>Blood Pressure Amp</td><td>AD Instruments, Colorado Springs, CO, USA</td><td>FE117</td><td>https://www.adinstruments.com/products/bp-blood-pressure-amp</td></tr><tr><td>Central Venous Catheter Introducer</td><td>Arrow International Inc, Reading, PA, USA</td><td>AK-09800</td><td>8.5 Fr. x 4&quot; (10 cm) Arrow-Flex</td></tr><tr><td>Central Venous Catheter-Introducer</td><td>Arrow International</td><td>CP-07611-P</td><td>Super Arrow-Flex Percutaneous Sheath Introducer Kit 6Fr./7.5cm</td></tr><tr><td>Disposable BP Transducers</td><td>AD Instruments, Colorado Springs, CO, USA</td><td>MLT0670</td><td>https://www.adinstruments.com/products/disposable-bp-transducers</td></tr><tr><td>Kendall 930 FoamElectrodes</td><td>Covidien, Mansfield, MA, USA</td><td>22935</td><td>https://www.cardinalhealth.com/en/product-solutions/medical/patient-monitoring/electrocardiography/monitoring-ecg-electrodes/radiolucent-electrodes/kendall-930-series-radiolucent-foam-electrodes.html</td></tr><tr><td>LabChart 8 software</td><td>AD Instruments, Colorado Springs, CO, USA</td><td>N/A</td><td>https://www.adinstruments.com/products/labchart</td></tr><tr><td>Peripheral IV Catheter Angiocath 18-24 Gauge 1.16 inch</td><td>McKesson, Irving, TX, USA</td><td>329830</td><td>https://mms.mckesson.com/product/329830/Becton-Dickinson-381144</td></tr><tr><td>PlamaLyte Crystilloid Solution</td><td>Baxter International, Deerfield, IL USA</td><td>2B2544X</td><td>https://www.ciamedical.com/baxter-2b2544x-each-solution-plasma-lyte-a-inj-ph-7-4-1000ml</td></tr><tr><td>PowerLab</td><td>ADInstruments, Colorado Springs, CO, USA</td><td>N/A</td><td>https://www.adinstruments.com/products/powerlab/c?creative=532995768429&amp;amp;keyword=&lt;br/&gt; powerlab&amp;amp;matchtype=e&amp;amp;network=&lt;br/&gt; g&amp;amp;device=c&amp;amp;gclid=CjwKCAjwysipB&lt;br/&gt; hBXEiwApJOcu-ulfO0bfCc-j6B7PpO&lt;br/&gt; kOAGur8IZ4SWNkhNZ7mORGstO&lt;br/&gt; vKON6plWLxoCigsQAvD_BwE</td></tr><tr><td>Pulmonary Artery Catheter</td><td>Edwards Life Sciences, Irvine, CA, USA</td><td>TS105F5</td><td>True Size Thermodilution Catheter 24cm Proximal Port- Swan Ganz&amp;nbsp;</td></tr><tr><td>Pulmonary Artery Catheter (7F)</td><td>Edwards Life Sciences, Irvine, CA, USA</td><td>131F7</td><td>Swan Ganz 7F x 110cm&amp;nbsp;</td></tr><tr><td>Telazol (Tiletamine HCl and Zolazepam HCl), Injectable Solution, 5 mL</td><td>Patterson Veterinary, Loveland, CO 80538</td><td>07-801-4969</td><td>https://www.pattersonvet.com/ProductItem/078014969?omni=telazol</td></tr><tr><td>Terumo Sarns 8000 Roller Pump</td><td>Terumo Cardiovascular, Ann Arbor, MI, USA</td><td>16402</td><td>https://aamedicalstore.com/products/terumo-sarns%E2%84%A2-8000-roller-pump</td></tr><tr><td>Xylazine HCl 100 mg/mL, Injectable Solution, 50 mL</td><td>Patterson Veterinary, Loveland, CO 80538</td><td>07-894-5244</td><td>https://www.pattersonvet.com/ProductItem/078945244</td></tr><tr><td>Yorkshire Adult Pigs</td><td>Oak Hill Genetics, Ewing, IL, USA</td><td>N/A</td><td>Yorkshire/Landrace 81-100lbs</td></tr><tr><td>Yorkshire Piglets</td><td>Oak Hill Genetics&amp;nbsp;</td><td>N/A</td><td>Female &quot;piglet&quot;, specify age 5 weeks with a correlating healthy weight range (approximately 10-20lbs.)</td></tr></tbody></table>

adult and pediatric porcine model of acute volume overload

This protocol describes an acute volume overload porcine model for adult Yorkshire pigs and piglets. Both swine ages undergo general anesthesia, endotracheal intubation, and mechanical ventilation. A central venous catheter and an arterial catheter are placed via surgical cutdown in the external jugular vein and carotid artery, respectively. A pulmonary artery catheter is placed through an introducer sheath of the central venous catheter. PlasmaLyte crystalloid solution is then administered at a rate of 100 mL/min in adult pigs and at 20 mL/kg boluses over 10 min in piglets. Hypervolemia is achieved either at 15% decrease in cardiac output or at 5 L in adult pigs and at 500 mL in piglets. Hemodynamic data, such as heart rate, respiratory rate, end-tidal carbon dioxide, fraction of oxygen-saturated hemoglobin, arterial blood pressure, central venous pressure, pulmonary artery pressure, pulmonary capillary wedge pressure, partial arterial oxygen pressure, lactate, pH, base excess, and pulmonary artery fraction of oxygen-saturated hemoglobin, are monitored during experimentation. Preliminary data observed with this model has demonstrated statistically significant changes and strong linear regressions between central hemodynamic parameters and acute volume overload in adult pigs. Only pulmonary capillary wedge pressure demonstrated both a linear regression and a statistical significance to acute volume overload in piglets. These models can aid scientists in the discovery of age-appropriate therapeutic and monitoring strategies to understand and prevent acute volume overload.

The preliminary representative pilot data after linear regression analysis for the adult pig model demonstrated linearity to volume administration in the first eight pigs (Figure 2). While many other data points, and volume beyond 2.5 L, were measured during this experiment, these data represent the analysis to date. The two vital signs most used for volume assessment, HR (R2=0.15) and MAP (R2=0.79), both demonstrated a linear relationship during forced hypervolemia, but did not demonstrate statistical significance (p&#62;0.05; Figure 2A,B). In comparison, the central hemodynamic variables CVP (R2=0.93), CO (R2=0.95), and PCWP (R2=0.98) demonstrated both linearity and statistical significance (p&#62;0.05; Figures 2C-E). A commonly used noninvasive measure of volume status, PPV, had a moderate inverse correlation (R2=0.41) and statistical significance (p&#60;0.5; Figure 2F); however, it was not as strong of a regression as the central hemodynamic data points to volume administered. Of note, no adult pig demonstrated a 15% decrease in CO up to the volume analyzed to date, i.e., 2.5 L. The average urine output was 1.2 L (SD=500 mL; n=6). While these data represent an analysis of 8 pigs (only 6 had accurate urine outputs), more data is being collected and future reports will include a more robust analysis of more pigs and their measured data points.
Representative pilot results after linear regression analysis from three piglets demonstrated that only the PCWP had a linear regression and statistical significance to the volume increase (R2=0.43, p=0.02; Figure 3E). Interestingly, MAP did have statistical significance; however, it was for an inverse relationship to volume given (R2=0.38, p=0.03; Figure 3B). Demonstrating that the administration of volume will decrease the MAP of a piglet. The other central variables did demonstrate a statistically significant relationship to the volume increase (CVP: R2=0.31, p=0.048; CO: R2=0.33, p=0.04; Figure 3C,D). However, based on their R2 values, they had weaker linear regressions to volume administered in these three piglets compared to PCWP. No other hemodynamic variable (HR, SvO2, PPV) demonstrated a linear regression to volume administration or any statistically significant relationship (Figures 3A,B,F; HR: R2=0.17, p=0.16; SvO2: R2=0.24, p=0.09; PPV: R2=0.01, p=0.097). While other data points were measured, specifically echocardiographic variables like PSV and LVOT diameter, these data represent the analysis to date. As with the adult pigs, the volume to data point regression was the strongest with the PCWP as volume overload was achieved. Of note, all piglets experienced a 15% decrease in CO at approximately 80 mL/kg. The average urine output was 115 mL (SD=111 mL; n=3). Piglet volume was measured as a % blood volume instead of absolute numbers as the variation in piglet weight and estimated blood volume varied greatly between piglets while the estimated blood volume for larger porcine studies did not. It should be emphasized that these data are representative pilot data from only 3 piglet experiments. Future reports will include an appropriate volume of data to draw stronger conclusions.
<img alt="Figure 1" class="xfigimg" src="/files/ftp_upload/65967/65967fig01v2.jpg" /> 
Figure 1: Timeline of porcine volume overload model in Yorkshire pigs and piglets. Adult pigs (top) and piglets (bottom) are both resuscitated to a euvolemic state (PCWP=8-10 mmHg).&#160;Once achieved, volume administration begins. Adult hemodynamic data (black arrow; heart rate, respiratory rate, blood pressure, SpO2, end-tidal carbon dioxide, pulmonary artery pressures, PCWP, central venous pressure, CO) and arterial blood gas analysis (<img alt="Equation 1" src="/files/ftp_upload/65967/65967eq01.jpg" style="margin: auto;" />; PaO2, PaCO2, Lactate, Base Excess, pH) are obtained during the experimentation after each 500 mL of fluid. In piglets, hemodynamic data (white arrow; heart rate, respiratory rate, blood pressure, SpO2, end-tidal carbon dioxide, pulmonary artery pressures, PCWP, and central venous pressure) and arterial blood gas analysis (<img alt="Equation 1" src="/files/ftp_upload/65967/65967eq01.jpg" style="margin: auto;" />; PaO2, PaCO2, Lactate, Base Excess, pH, and SvO2) are obtained until euthanasia after each 20 mL/kg bolus of fluid (volumes shown are based on average mass of 5 kg of 5-week piglets). Transthoracic echocardiography measurements are made at each volume point until euthanasia (<img alt="Equation 1" src="/files/ftp_upload/65967/65967eq01.jpg" style="margin: auto;" />; Aortic Blood Flow Peak Systolic Velocity and Left Ventricular Outflow Tract diameter). Euthanasia occurs at a 15% decrease in CO or 5L (adult)/ 500 mL (piglet). Abbreviations: PaO2= partial pressure of oxygen; PaCO2= partial pressure of carbon dioxide; mL= millimeter; kg= kilogram; CO= cardiac output; PCWP= pulmonary capillary wedge pressure. <a href="https://www-jove-com.remotexs.ntu.edu.sg/files/ftp_upload/65967/65967fig01largev2.jpg" target="_blank">Please click here to view a larger version of this figure.</a>
<img alt="Figure 2" class="xfigimg" src="/files/ftp_upload/65967/65967fig02.jpg" /> 
Figure 2: Representative results of adult pig hemodynamic variable response to acute volume administration. In the preliminary analysis of 8 pigs, all vital signs, (A) heart rate (R2=0.15) and (B) mean arterial pressure (R2=.79) did demonstrate a linear regression. (C) Central hemodynamic indices, CVP (R2=0.93), (D) CO (R2=0.95), (E) PCWP (R2=0.98), demonstrated stronger linear regressions when compared to vital signs. (F) Calculated pulse pressure variability also did, appropriately, demonstrate an inverse correlation with the volume administered (R2=0.41). The volume to data point regression was the strongest with the PCWP as volume overload was achieved. Abbreviations: HR= heart rate; MAP= mean arterial pressure; CVP= central venous pressure; CO= cardiac output; PCWP= pulmonary capillary wedge pressure; PPV= pulse pressure variation; mL= milliliters. <a href="https://www-jove-com.remotexs.ntu.edu.sg/files/ftp_upload/65967/65967fig02large.jpg" target="_blank">Please click here to view a larger version of this figure.</a>
<img alt="Figure 3" class="xfigimg" src="/files/ftp_upload/65967/65967fig03.jpg" /> 
Figure 3: Representative results of piglet hemodynamic variable response to acute volume administration. (E)&#160;In the preliminary analysis of 3 piglets, only PCWP demonstrated a linear regression and a statistically significant relationship to the % volume increase (R2=0.43, p=0.2). (C) CVP and (D) Fick based CO did demonstrate a statistically significant relationship with volume increase (R2=0.31, p=0.048; D left y-axis: R2=0.33, p=0.04), however, a weaker linear regression was witnessed compared to PCWP. (B) MAP demonstrated an inverse linear regression to volume increase with statistical significance (R2=0.38, p=0.03). (A, F) No other hemodynamic variable (D right y-axis) demonstrated a linear regression to volume administration or any statistically significant relationship (HR: R2=0.17, p=0.16; SvO2: R2=0.24, p=0.09; PPV: R2=0.01, p=0.097). The volume to data point regression was the strongest with the PCWP as volume overload was achieved. Piglet volume was measured as a % blood volume instead of absolute numbers as the variation in piglet weight and estimated blood volume varied greatly between piglets. Abbreviations: HR= heart rate; MAP= mean arterial pressure; CVP= central venous pressure; CO= cardiac output; PCWP= pulmonary capillary wedge pressure; PPV= pulse pressure variation; mL= milliliters; SvO2=oxygen-saturated hemoglobin from the pulmonary artery blood. <a href="https://www-jove-com.remotexs.ntu.edu.sg/files/ftp_upload/65967/65967fig03large.jpg" target="_blank">Please click here to view a larger version of this figure.</a>

Watch this Scientific Journal Video about Author Spotlight: Exploring Venous Waveforms in Porcine Models to Tackle Volume Overload in Medicine at JoVE.com

Author Spotlight: Exploring Venous Waveforms in Porcine Models to Tackle Volume Overload in Medicine

The protocol here shows how continuous administration of crystalloids into the central veins of a euvolemic pig/piglet allows for the appropriate investigation of the physiological effects of acute volume overload.

Adult and Pediatric Porcine Model of Acute Volume Overload

This protocol describes an acute volume overload porcine model for adult Yorkshire pigs and piglets. Both swine ages undergo general anesthesia, ...

adult-and-pediatric-porcine-model-of-acute-volume-overload

Nanyang Technological University

Research

JoVE Journal

Medicine

490 Views.  Vanderbilt University Medical Center. The protocol here shows how continuous administration of crystalloids into the central veins of a euvolemic pig/piglet allows for the appropriate investigation of the physiological effects of acute volume overload.

Video: Author Spotlight: Exploring Venous Waveforms in Porcine Models to Tackle Volume Overload in Medicine

Tachycardia-Induced Cardiomyopathy As a Chronic Heart Failure Model in Swine

A stable and reliable model of chronic heart failure is required for many experiments to understand hemodynamics or to test effects of new treatment methods. Here, we present such a model by tachycardia-induced cardiomyopathy, which can be produced by rapid cardiac pacing in swine.
A single pacing lead is introduced transvenously into fully anaesthetized healthy swine, to the apex of the right ventricle, and fixated. Its other end is then tunneled dorsally to the paravertebral region. There, it is connected to an in-house modified heart pacemaker unit that is then implanted in a subcutaneous pocket. 
After 4 - 8 weeks of rapid ventricular pacing at rates of 200 - 240 beats/min, physical examination revealed signs of severe heart failure - tachypnea, spontaneous sinus tachycardia, and fatigue. Echocardiography and X-ray showed dilation of all heart chambers, effusions, and severe systolic dysfunction. These findings correspond well to decompensated dilated cardiomyopathy and are also preserved after the cessation of pacing.
This model of tachycardia-induced cardiomyopathy can be used for studying the pathophysiology of progressive chronic heart failure, especially hemodynamic changes caused by new treatment modalities like mechanical circulatory supports. This methodology is easy to perform and the results are robust and reproducible.

Here, we present a protocol to produce tachycardia-induced cardiomyopathy in swine. This model represents a potent way to study the hemodynamics of progressive chronic heart failure and the effects of applied treatment.

A stable and reliable model of chronic heart failure is required for many experiments to understand hemodynamics or to test effects of new treatment ...

Invasive Hemodynamic Monitoring of Aortic and Pulmonary Artery Hemodynamics in a Large Animal Model of ARDS

One of the leading causes of morbidity and mortality in patients with heart failure is right ventricular (RV) dysfunction, especially if it is due to pulmonary hypertension. For a better understanding and treatment of this disease, precise hemodynamic monitoring of left and right ventricular parameters is important. For this reason, it is essential to establish experimental pig models of cardiac hemodynamics and measurements for research purpose. 
This article shows the induction of ARDS by using oleic acid (OA) and consequent right ventricular dysfunction, as well as the instrumentation of the pigs and the data acquisition process that is needed to assess hemodynamic parameters. To achieve right ventricular dysfunction, we used oleic acid (OA) to cause ARDS and accompanied this with pulmonary artery hypertension (PAH). With this model of PAH and consecutive right ventricular dysfunction, many hemodynamic parameters can be measured, and right ventricular volume load can be detected. 
All vital parameters, including respiratory rate (RR), heart rate (HR) and body temperature were recorded throughout the whole experiment. Hemodynamic parameters including femoral artery pressure (FAP), aortic pressure (AP), right ventricular pressure (peak systolic, end systolic and end diastolic right ventricular pressure), central venous pressure (CVP), pulmonary artery pressure (PAP) and left arterial pressure (LAP) were measured as well as perfusion parameters including ascending aortic flow (AAF) and pulmonary artery flow (PAF). Hemodynamic measurements were performed using transcardiopulmonary thermodilution to provide cardiac output (CO). Furthermore, the PiCCO2 system (Pulse Contour Cardiac Output System 2) was used to receive parameters such as stroke volume variance (SVV), pulse pressure variance (PPV), as well as extravascular lung water (EVLW) and global end-diastolic volume (GEDV). Our monitoring procedure is suitable for detecting right ventricular dysfunction and monitoring hemodynamic findings before and after volume administration.

We present a protocol of creating right ventricular dysfunction in a pig model by inducing ARDS. We demonstrate invasive monitoring of left and right ventricular cardiac output using flow probes around the aorta and the pulmonary artery, as well as blood pressure measurements in the aorta and pulmonary artery.

One of the leading causes of morbidity and mortality in patients with heart failure is right ventricular (RV) dysfunction, especially if it is due to ...

Induction and Phenotyping of Acute Right Heart Failure in a Large Animal Model of Chronic Thromboembolic Pulmonary Hypertension

The development of acute right heart failure (ARHF) in the context of chronic pulmonary hypertension (PH) is associated with poor short-term outcomes. The morphological and functional phenotyping of the right ventricle is of particular importance in the context of hemodynamic compromise in patients with ARHF. Here, we describe a method to induce ARHF in a previously described large animal model of chronic PH, and to phenotype, dynamically, right ventricular function using the gold standard method (i.e., pressure-volume PV loops) and with a non-invasive clinically available method (i.e., echocardiography). Chronic PH is first induced in pigs by left pulmonary artery ligation and right lower lobe embolism with biological glue once a week for 5 weeks. After 16 weeks, ARHF is induced by successive volume loading using saline followed by iterative pulmonary embolism until the ratio of the systolic pulmonary pressure over systemic pressure reaches 0.9 or until the systolic systemic pressure decreases below 90 mmHg. Hemodynamics are restored with dobutamine infusion (from 2.5 &#181;g/kg/min to 7.5 &#181;g/kg/min). PV-loops and echocardiography are performed during each condition. Each condition requires around 40 minutes for induction, hemodynamic stabilization and data acquisition. Out of 9 animals, 2 died immediately after pulmonary embolism and 7 completed the protocol, which illustrates the learning curve of the model. The model induced a 3-fold increase in mean pulmonary artery pressure. The PV-loop analysis showed that ventriculo-arterial coupling was preserved after volume loading, decreased after acute pulmonary embolism and was restored with dobutamine. Echocardiographic acquisitions allowed to quantify right ventricular parameters of morphology and function with good quality. We identified right ventricular ischemic lesions in the model. The model can be used to compare different treatments or to validate non-invasive parameters of right ventricular morphology and function in the context of ARHF.

We present a protocol to induce and phenotype an acute right heart failure in a large animal model with chronic pulmonary hypertension. This model can be used to test therapeutic interventions, to develop right heart metrics&#160;or to improve the understanding of acute right heart failure pathophysiology.

The development of acute right heart failure (ARHF) in the context of chronic pulmonary hypertension (PH) is associated with poor short-term outcomes. The ...

A Pre-Clinical Porcine Model of Orthotopic Heart Transplantation

Fifty-years following the first successful report, cardiac transplantation remains the gold-standard treatment for eligible patients with advanced heart failure. Multiple small-animal models of heart transplantation have been used to study the acute and long-term effects of novel therapies. However, few are tested and demonstrated success in clinical trials. It is of critical importance to evaluate new therapies in a clinically relevant large-animal model for efficient and reliable translation of basic studies&#39;&#160;findings. Here, we describe a pre-clinical large-animal (porcine) model of orthotopic heart transplantation that has been firmly established and previously used to investigate novel cardioprotective strategies. This procedure focuses on acute ischemia-reperfusion injury and is a reliable method to investigate novel interventions which have been tested and validated in smaller experimental models, such as the murine model. We demonstrate its usefulness in assessing cardiac performance during the early post-transplantation period and other potential possibilities enabled by the model.

Here, we describe a pre-clinical large-animal (porcine) model of orthotopic heart transplantation that has been firmly established and utilized to investigate novel cardioprotective strategies.

Fifty-years following the first successful report, cardiac transplantation remains the gold-standard treatment for eligible patients with advanced heart ...

A Novel Right Ventricular Volume and Pressure Loaded Piglet Heart Model for the Study of Tricuspid Valve Function.

Heart conditions in which the tricuspid valve (TV) faces either increased volume or pressure stressors are associated with premature valve failure. Mechanistic studies to improve our&#160;understanding of the underlying pathophysiology responsible for the development of premature TV failure are lacking. Due to the inability to conduct these studies in humans, an animal model is required. In this manuscript, we describe the protocols for a novel chronic recovery infant piglet heart model for the study of changes in the TV when placed under combined volume and pressure stress. In this model, volume loading of the right ventricle&#160;and the TV is achieved through the disruption of the pulmonary valve. Then pressure loading is accomplished through the placement of a pulmonary artery band. The success of this model is assessed at four weeks post intervention surgery through echocardiography, intracardiac pressure measurement, and pathologic examination of the heart specimens.

A novel recovery piglet heart model with combined pressure and volume overload on the right ventricle is described for the study of tricuspid valve function.

Heart conditions in which the tricuspid valve (TV) faces either increased volume or pressure stressors are associated with premature valve failure. ...

Extracorporeal Cardiopulmonary Resuscitation in Animal Model After Cardiac Arrest

Pediatric Animal Model of Extracorporeal Cardiopulmonary Resuscitation After Prolonged Circulatory Arrest

Congenital heart disease (CHD) is the most prevalent congenital malformation, with about one million births impacted worldwide per year. Comprehensive investigation of this disease requires appropriate and validated animal models. Piglets are commonly used for translational research due to their analogous anatomy and physiology. This work aimed to describe and validate a neonatal piglet model of cardiopulmonary bypass (CPB) with circulatory and cardiac arrest (CA) as a tool for studying severe brain damage and other complications of cardiac surgery. In addition to including a list of materials, this work provides a roadmap for other investigators to plan and execute this protocol. After experienced practitioners performed several trials, the representative results of the model demonstrated a 92% success rate, with failures attributed to small piglet size and variant vessel anatomy. Furthermore, the model allowed practitioners to select from a wide variety of experimental conditions, including varying times in CA, temperature alterations, and pharmacologic interventions. In summary, this method uses materials readily available in most hospital settings, is reliable and reproducible, and can be widely employed to enhance translational research in children undergoing heart surgery.

Author Spotlight: Simulating Pediatric Cardiac Surgery Using a Neonatal Piglet Model

This protocol describes a neonatal porcine model of cardiopulmonary bypass (CPB), with circulatory and cardiac arrest as a tool for studying severe brain damage and other complications secondary to CPB.

Congenital heart disease (CHD) is the most prevalent congenital malformation, with about one million births impacted worldwide per year. Comprehensive ...

Modified Arteriovenous Fistula Surgery to Establish the RV Volume Overload Mouse Model

Establishment and Confirmation of a Postnatal Right Ventricular Volume Overload Mouse Model

Right ventricular (RV) volume overload (VO) is common in children with congenital heart disease. In view of distinct developmental stages,the RV myocardium may respond differently to VO in children compared to adults. The present study aims to establish a postnatal RV VO model in mice using a modified abdominal arteriovenous fistula. To confirm the creation of VO and the following morphological and hemodynamic changes of the RV, abdominal ultrasound, echocardiography, and histochemical staining were performed for 3 months. As a result, the procedure in postnatal mice showed an acceptable survival and fistula success rate. In VO mice, the RV cavity was enlarged with a thickened free wall, and the stroke volume was increased by about 30%-40% within 2 months after surgery. Thereafter, the RV systolic pressure increased, corresponding pulmonary valve regurgitation was observed, and small pulmonary artery remodeling appeared. In conclusion, modified arteriovenous fistula (AVF) surgery is feasible to establish the RV VO model in postnatal mice. Considering the probability of fistula closure and elevated pulmonary artery resistance, abdominal ultrasound and echocardiography must be performed to confirm the model status before application.

Author Spotlight: Establishment and Confirmation of a Postnatal Right Ventricular Volume Overload Mouse Model  

This protocol presents the establishment and confirmation of a postnatal right ventricular volume overload (VO) model in mice with abdominal arteriovenous fistula (AVF), which can be applied to investigate how VO contributes to postnatal heart development.

Right ventricular (RV) volume overload (VO) is common in children with congenital heart disease. In view of distinct developmental stages,the RV ...

Description of a Swine Infant Model of Volume-Controlled Hemorrhagic Shock

Hemorrhagic shock is a leading cause of morbidity and mortality in pediatric patients. Interpretation of the clinical indicators validated in adults to guide resuscitation and comparison between different therapies is difficult in children due to the inherent heterogeneity of this population. As a result, compared to adults, appropriate management of pediatric hemorrhagic shock is still not well established. In addition, the scarcity of pediatric patients with hemorrhagic shock precludes the development of clinically relevant studies. For this reason, an experimental pediatric animal model is necessary to study the effects of hemorrhage in children as well as their response to different therapies. We present an infant animal model of volume-controlled hemorrhagic shock in anesthetized young pigs. Hemorrhage is induced by withdrawing a previously calculated blood volume, and the pig is subsequently monitored and resuscitated with different therapies. Here, we describe a precise and highly reproducible model of hemorrhagic shock in immature swine. The model yields hemodynamic data that characterizes compensatory mechanisms that are activated in response to severe hemorrhage.

This article aims to provide researchers with a detailed and accessible guide to set up an infant porcine model of hemorrhagic shock.

Hemorrhagic shock is a leading cause of morbidity and mortality in pediatric patients. Interpretation of the clinical indicators validated in adults to ...

Oleic Acid&#45;Induced Acute Respiratory Decompensation in Pigs

A Porcine Model of Acute Respiratory Failure with a Continuous Infusion of Oleic Acid

This protocol outlines an acute respiratory distress model utilizing centrally administered oleic acid infusion in Yorkshire pigs. Prior to experimentation, each pig underwent general anesthesia, endotracheal intubation, and mechanical ventilation, and was equipped with bilateral&#160;jugular vein central vascular access catheters. Oleic acid was administered through a dedicated pulmonary artery catheter at a rate of 0.2 mL/kg/h. The infusion lasted for 60-120 min, inducing respiratory distress. Throughout the experiment, various parameters including heart rate, respiratory rate, arterial blood pressure, central venous pressure, pulmonary artery pressure, pulmonary capillary wedge pressure, end-tidal carbon dioxide, peak airway pressures, and plateau pressures were monitored. Around the 60 min mark, decreases in partial arterial oxygen pressure (PaO2) and fraction of oxygen-saturated hemoglobin (SpO2) were observed. Periodic hemodynamic instability, accompanied by acute increases in pulmonary artery pressures, occurred during the infusion. Post-infusion, histological analysis of the lung parenchyma revealed changes indicative of parenchymal damage and acute disease processes, confirming the effectiveness of the model in simulating acute respiratory decompensation.

Author Spotlight: Exploring Venous Waveforms for Non&#45;Invasive Respiratory Monitoring in Pigs

Infusing oleic acid continuously into the pulmonary artery of an anesthetized adult pig induces acute respiratory failure, enabling controlled experimentation during acute respiratory decompensation.

This protocol outlines an acute respiratory distress model utilizing centrally administered oleic acid infusion in Yorkshire pigs. Prior to ...

A Porcine Model of Acute Autologous Pulmonary Embolism

Acute pulmonary embolism (PE) is a potentially life-threatening condition that causes abrupt obstruction of the pulmonary arteries, leading to acute right heart failure. Novel diagnostic methods and catheter-directed therapies are being developed rapidly, and there is an obvious need for a realistic PE animal model that can be used for pathophysiological evaluation and preclinical testing.
This protocol introduces a porcine model employing large autologous pulmonary emboli. Instrumentations are performed with minimally invasive techniques, creating a close-chest model that enables the investigation of various treatment options with high reproducibility. Three hours after drawing blood to create autologous emboli ex vivo, the induction of PE caused an immediate increase in the mean pulmonary arterial pressure (17&#160;&#177; 3 mmHg to 33&#160;&#177; 6 mmHg, p &#60; 0.0001) and heart rate (50&#160;&#177; 9 beats&#183;min-1 to 63&#160;&#177; 6 beats&#183;min-1,&#160;p &#60; 0.0003) accompanied by a decreased cardiac output (5.0&#160;&#177; 0.8 L/min to 4.5&#160;&#177; 0.9 L/min, p &#60; 0.037) compared to baseline. The CT pulmonary angiography revealed multiple emboli, and the pulmonary obstruction percentage was increased compared to baseline (0% [0-0] to 57.1% [38.8-63.3], p &#60; 0.0001). In the acute phase, the phenotype is comparable to intermediate-risk PE.
The model represents a realistic and well-characterized phenotype of intermediate-risk PE and creates an opportunity to test novel diagnostic methods, interventional and pharmaceutical treatments, and hands-on training for healthcare workers in interventional procedures.

This study presents a porcine model of pulmonary embolism (PE) using large autologous emboli that replicate acute intermediate-risk PE. The model is well-suited for the evaluation of both pathophysiology and treatment responses.

Acute pulmonary embolism (PE) is a potentially life-threatening condition that causes abrupt obstruction of the pulmonary arteries, leading to acute right ...