The successful development and testing of new thrombolytics in animal models requires monitoring of hemodynamic changes in cerebral circulation before and after stroke. The purpose of the present study was to document that percutaneous transcranial Doppler (TCD) monitoring is able to differentiate two hemodynamic situations induced with two anesthetic protocols. Twelve adult rats divided into two groups underwent general anesthesia (60 min) using combination: 1) ketamine-xylazine-diazepam (KXD); and 2) ketamine-xylazine-urethane-alpha-chloralose (URACH). The TCD was performed with the skin and skull intact. The heart rate, peak systolic velocity, pulsatility index, and resistance index were recorded in a branch of the posterior cerebral artery. Flow detection and measurement was possible in all rat brains bilaterally. The mean heart rate was lower in the KXD 243+/-4 (range: 238 to 249) than in the URACH group 265+/-12 (range: 250 to 279), the difference between means: 22; 95 % CI [8 to 34], p=0.005) only for the first 20 min of monitoring. Peak systolic velocity was lower in the KXD 73.4+/-3.3 mm/s (range 70.3 to 76.5) vs. URACH group 93.7+/-4.0 mm/s (range: 90.0 to 97.4) during the entire observation period (difference between means: 20; 95 % CI [16 to 25], p<0.001). Same difference was observed for pulsatility and resistance indexes. TCD was able to differentiate hemodynamic changes in the rat brains, making the TCD suitable for monitoring of hemodynamic changes and explores, e.g. how such changes contribute to hemorrhagic transformation after thrombolysis. Also, TCD holds promise as a tool for monitoring of recanalization induced by thrombolytics. Key words Non-invasive monitoring " Brain flow velocity " Anesthesia " Animal model.

Stroke Group St Anne's University Hospital Brno Czech Republic

See more in PubMed

Powers WJ, Rabinstein AA, Ackerson T, Adeoye OM, Bambakidis NC, Becker K, Biller J, et al. 2018 Guidelines for the Early Management of Patients With Acute Ischemic Stroke: A Guideline for Healthcare professionals from the American Heart association/American Stroke Association. Stroke. 2018;49:e46–e110. doi: 10.1161/STR.0000000000000158. PubMed DOI

Martins SCO, Pontes-Neto OM, Pille A, Secchi TL, Miranda Alves MA, Rebello LC, Oliveira-Filho J, et al. Reperfusion therapy for acute ischemic stroke: where are we in 2023? Arq Neuropsiquiatr. 2023;81:1030–1039. doi: 10.1055/s-0043-1777721. PubMed DOI PMC

Spronk E, Sykes G, Falcione S, Munsterman D, Joy T, Kamtchum-Tatuene J, Jickling GC. Hemorrhagic Transformation in Ischemic Stroke and the Role of Inflammation. Front Neurol. 2021;12:661955. doi: 10.3389/fneur.2021.661955. PubMed DOI PMC

Kovács KB, Bencs V, Hudák L, Oláh L, Csiba L. Hemorrhagic Transformation of Ischemic Strokes. Int J Mol Sci. 2023;24:14067. doi: 10.3390/ijms241814067. PubMed DOI PMC

Bang OY, Saver JL, Buck BH, Alger JA, Starkman S, Ovbiagele B, Kim D, et al. Impact of collateral flow on tissue fate in acute ischaemic stroke. J Neurol Neurosurg Psychiatry. 2008;79:625–629. doi: 10.1136/jnnp.2007.132100. PubMed DOI PMC

Freitas-Andrade M, Raman-Nair J, Lacoste B. Structural and Functional Remodeling of the Brain Vasculature Following Stroke. Front Physiol. 2020;11:948. doi: 10.3389/fphys.2020.00948. PubMed DOI PMC

Sutherland BA, Rabie T, Buchan AM. Laser Doppler flowmetry to measure changes in cerebral blood flow. Methods Mol Biol. 2014;1135:237–248. doi: 10.1007/978-1-4939-0320-7_20. PubMed DOI

Liao LD, Tsytsarev V, Delgado-Martínez I, Li ML, Erzurumlu R, Vipin A, Orellana J, et al. Neurovascular coupling: in vivo optical techniques for functional brain imaging. Biomed Eng Online. 2013;12:38. doi: 10.1186/1475-925X-12-38. PubMed DOI PMC

Zhao Y, Zhang J, Yu H, Hou X, Zhang J. Noninvasive microvascular imaging in newborn rats using high-frequency ultrafast ultrasound. Neuroimage. 2024;297:120738. doi: 10.1016/j.neuroimage.2024.120738. PubMed DOI

Premilovac D, Blackwood SJ, Ramsay CJ, Keske MA, Howells DW, Sutherland BA. Transcranial contrast-enhanced ultrasound in the rat brain reveals substantial hyperperfusion acutely post-stroke. J Cereb Blood Flow Metab. 2020;40:939–953. doi: 10.1177/0271678X20905493. PubMed DOI PMC

Wang W, Huang Z, Chen S, EY, Qi J, Xie Y, Su M, Zhang Y, Jiang T, Zhang X. Early infarct growth rate is associated with symptomatic intracranial hemorrhage after endovascular thrombectomy. Ther Adv Neurol Disord. 2024;17:17562864241306561. doi: 10.1177/17562864241306561. PubMed DOI PMC

Alexander TH, Hennigan A, Harrison P, Plotkin G. Transcranial Doppler: The Fifth Decade. J Vasc Ultrasound. 2021;45:15–24. doi: 10.1177/1544316720976210. DOI

Soares MS, Andrade AF, Brasil S, DE-Lima-Oliveira M, Belon AR, Bor-Seng-Shu E, Nogueira RC, et al. Evaluation of cerebral hemodynamics by transcranial Doppler ultrasonography and its correlation with intracranial pressure in an animal model of intracranial hypertension. Arq Neuropsiquiatr. 2022;80:344–352. doi: 10.1590/0004-282x-anp-2020-0591. PubMed DOI PMC

Mulder KJ, Mulder JB. Ketamine and xylazine anesthesia in the mouse. Vet Med Small Anim Clin. 1979;74:569–570. PubMed

Scheer P, Kosáková D, Doubek J. Střednědobá injekční anestezie po éterové indukci u laboratorního potkana. (Article in Czech) Cesk Fysiol. 2003;52:172–178. PubMed

Hama N, Ito SI, Hirota A. Optical imaging of the propagation patterns of neural responses in the rat sensory cortex: comparison under two different anesthetic conditions. Neuroscience. 2015;284:125–133. doi: 10.1016/j.neuroscience.2014.08.059. PubMed DOI

Brožíčková C, Otáhal J. Effect of an inhibitor of neuronal nitric oxide synthase 7-nitroindazole on cerebral hemodynamic response and brain excitability in urethane-anesthetized rats. Physiol Res. 2013;62(Suppl 1):S57–S66. doi: 10.33549/physiolres.932564. PubMed DOI

Zagvazdin Y, Sancesario G, Fitzgerald ME, Reiner A. Effects of halothane and urethane-chloralose anaesthesia on the pressor and cerebrovascular responses to 7-NITROINDAZOLE, an inhibitor of nitric oxide synthase. Pharmacol Res. 1998;38:339–346. doi: 10.1006/phrs.1998.0374. PubMed DOI

Gómez-de Frutos MC, García-Suárez I, Laso-García F, Diekhorst L, Otero-Ortega L, Alonso-López E, Díez-Tejedor E, et al. Identification of brain structures and blood vessels by conventional ultrasound in rats. J Neurosci Methods. 2020;346:108935. doi: 10.1016/j.jneumeth.2020.108935. PubMed DOI

Bathala L, Mehndiratta MM, Sharma VK. Transcranial doppler: Technique and common findings (Part 1) Ann Indian Acad Neurol. 2013;16:174–179. doi: 10.4103/0972-2327.112460. PubMed DOI PMC

Bunting KV, Steeds RP, Slater K, Rogers JK, Gkoutos GV, Kotecha D. A Practical Guide to Assess the Reproducibility of Echocardiographic Measurements. J Am Soc Echocardiogr. 2019;32:1505–1515. doi: 10.1016/j.echo.2019.08.015. PubMed DOI

Kim J, Park JE, Nahrendorf M, Kim D-E. Direct Thrombus Imaging in Stroke. J Stroke. 2016;18:286–296. doi: 10.5853/jos.2016.00906. PubMed DOI PMC

Bourquin Ch, Porée J, Lesage F, Provost J. In Vivo Pulsatility Measurement of Cerebral Microcirculation in Rodents Using Dynamic Ultrasound Localization Microscopy. IEEE Transac Med Imaging. 2022;41:782–791. doi: 10.1109/TMI.2021.3123912. PubMed DOI

Samary CS, Ramos AB, Maia LA, Rocha NN, Santos CL, Magalhães RF, Clevelario AL, et al. Focal ischemic stroke leads to lung injury and reduces alveolar macrophage phagocytic capability in rats. Crit Care. 2018;22:249. doi: 10.1186/s13054-018-2164-0. PubMed DOI PMC

Ma RF, Xue LL, Liu JX, Chen L, Xiong LL, Wang TH, Liu F. Transcranial Doppler Ultrasonography detection on cerebral infarction and blood vessels to evaluate hypoxic ischemic encephalopathy modeling. Brain Res. 2023;1822:148580. doi: 10.1016/j.brainres.2023.148580. PubMed DOI

Dolgorukova A, Potapenko AV, Murzina AA, Lyubashina OA, Sokolov AY. The implementation of transcranial Doppler ultrasonography for preclinical study of migraine. Can J Physiol Pharmacol. 2022;100:553–561. doi: 10.1139/cjpp-2021-0626. PubMed DOI

Giustetto P, Filippi M, Castano M, Terreno E. Non-invasive parenchymal, vascular and metabolic high-frequency ultrasound and photoacoustic rat deep brain imaging. J Vis Exp. 2015;(97):52162. doi: 10.3791/52162. PubMed DOI PMC

Errico C, Osmanski BF, Pezet S, Couture O, Lenkei Z, Tanter M. Transcranial functional ultrasound imaging of the brain using microbubble-enhanced ultrasensitive Doppler. Neuroimage. 2016;124:752–761. doi: 10.1016/j.neuroimage.2015.09.037. PubMed DOI PMC

Bonnin P, Debbabi H, Mariani J, Charriaut-Marlangue C, Renolleau S. Ultrasonic assessment of cerebral blood flow changes during ischemia-reperfusion in 7-day-old rats. Ultrasound Med Biol. 2008;34:913–922. doi: 10.1016/j.ultrasmedbio.2007.11.018. PubMed DOI

Kreis D, Schulz D, Stein M, Preuss M, Nestler U. Assessment of parameters influencing the blood flow velocities in cerebral arteries of the rat using ultrasonographic examination. Neurol Res. 2011;33:389–395. doi: 10.1179/1743132810Y.0000000010. PubMed DOI

Li L, Ke Z, Tong KY, Ying M. Evaluation of cerebral blood flow changes in focal cerebral ischemia rats by using transcranial Doppler ultrasonography. Ultrasound Med Biol. 2010;36:595–603. doi: 10.1016/j.ultrasmedbio.2010.01.005. PubMed DOI

Ke Z, Ying M, Li L, Zhang S, Tong KY. Evaluation of transcranial Doppler flow velocity changes in intracerebral hemorrhage rats using ultrasonography. J Neurosci Methods. 2012;210:272–280. doi: 10.1016/j.jneumeth.2012.07.024. PubMed DOI

Flecknell PA. Laboratory Animal Anaesthesia. Third Edition. Academic Press; 2009. Anaesthesia; pp. 19–78. DOI

Reinhart KM, Morton RA, Brennan KC, Carlson AP, Shuttleworth CW. Ketamine improves neuronal recovery following spreading depolarization in peri-infarct tissues. J Neurochem. 2024;168:855–867. doi: 10.1111/jnc.15923. PubMed DOI PMC

Sumitra M, Manikandan P, Rao KV, Nayeem M, Manohar BM, Puvanakrishnan R. Cardiorespiratory effects of diazepam-ketamine, xylazine-ketamine and thiopentone anesthesia in male Wistar rats--a comparative analysis. Life Sci. 2004;75:1887–1896. doi: 10.1016/j.lfs.2004.05.009. PubMed DOI

Dembovska K, Svoboda P, Scheer P, Kantorová I, Tomenedálová J, Řeháková K, Doubek J. Model of septic shock induced by live E. coli (O18) in a laboratory rat. Vet Medicina. 2018;53:153–164. doi: 10.17221/1942-VETMED. DOI

Schmittner MD, Vajkoczy SL, Horn P, Bertsch T, Quintel M, Vajkoczy P, Muench E. Effects of fentanyl and S(+)-ketamine on cerebral hemodynamics, gastrointestinal motility, and need of vasopressors in patients with intracranial pathologies: a pilot study. J Neurosurg Anesthesiol. 2007;19:257–262. doi: 10.1097/ANA.0b013e31811f3feb. PubMed DOI

Lei H, Grinberg O, Nwaigwe CI, Hou HG, Williams H, Swartz HM, Dunn JF. The effects of ketamine-xylazine anesthesia on cerebral blood flow and oxygenation observed using nuclear magnetic resonance perfusion imaging and electron paramagnetic resonance oximetry. Brain Res. 2001;913:174–179. doi: 10.1016/S0006-8993(01)02786-X. PubMed DOI

Thalerová S, Vítečková Wünschová A, Kittová P, Vašátková L, Pešková M, Volný O, et al. A collateral circulation in ischemic stroke accelerates recanalization due to lower clot compaction. PLoS One. 2024;19:e0314079. doi: 10.1371/journal.pone.0314079. PubMed DOI PMC

Low LA, Bauer LC, Klaunberg BA. Comparing the Effects of Isoflurane and Alpha Chloralose upon Mouse Physiology. PLoS One. 2016;11:e0154936. doi: 10.1371/journal.pone.0154936. PubMed DOI PMC

Silver NRG, Ward-Flanagan R, Dickson CT. Long-term stability of physiological signals within fluctuations of brain state under urethane anesthesia. PLoS One. 2021;16:e0258939. doi: 10.1371/journal.pone.0258939. PubMed DOI PMC

Zehendner CM, Luhmann HJ, Yang JW. A simple and novel method to monitor breathing and heart rate in awake and urethane-anesthetized newborn rodents. PLoS One. 2013;8:e62628. doi: 10.1371/journal.pone.0062628. PubMed DOI PMC

Silverman J, Muir WW., 3rd A review of laboratory animal anesthesia with chloral hydrate and chloralose. Lab Anim Sci. 1993;43:210–216. PubMed

Field KJ, Lang CM. Hazards of urethane (ethyl carbamate): a review of the literature. Lab Anim. 1988;22:255–262. doi: 10.1258/002367788780746331. PubMed DOI

Tejima E, Katayama Y, Suzuki Y, Kano T, Lo EH. Hemorrhagic transformation after fibrinolysis with tissue plasminogen activator: evaluation of role of hypertension with rat thromboembolic stroke model. Stroke. 2001;32:1336–1340. doi: 10.1161/01.STR.32.6.1336. PubMed DOI

Klein KU, Glaser M, Reisch R, Tresch A, Werner C, Engelhard K. The effects of arterial carbon dioxide partial pressure and sevoflurane on capillary venous cerebral blood flow and oxygen saturation during craniotomy. Anesth Analg. 2009;109:199–204. doi: 10.1213/ane.0b013e3181a800e5. PubMed DOI

Yeo LL, Arnberg F, Chireh A, Sharma V, Tan B, Gontu V, Little P, Holmin S. The Role of Carbon Dioxide in the Rat Acute Stroke Penumbra. Front Digit Health. 2022;3:824334. doi: 10.3389/fdgth.2021.824334. PubMed DOI PMC