Improving Stereotaxic Neurosurgery Techniques and Procedures Greatly Reduces the Number of Rats Used per Experimental Group—A Practice Report

Simple Summary Stereotaxic surgery techniques are commonly used today in research laboratories by a range of students, technicians, and researchers. Over the past twenty years, technical and scientific progress has been made in neurosurgery to meet the evolving requirements imposed by international legislation, and to promote the implementation of 3R rules. These improvements were motivated by a greater awareness of animal welfare and the necessary effort in the reduction of the number of animals used in experiments. The data presented in the present study show that technical and methodological improvements brought to our surgical procedures from 1992 resulted in reproducible stereotaxic neurosurgeries and in a significant reduction in experimental errors and animal morbidity. The effects of these improvements include a decrease in the final number of animals used in our experiments as well as better management of pain during and after surgery and the use of appropriate aseptic techniques. Correct stereotaxic surgical approaches are precisely described throughout the text. Abstract Techniques of stereotaxic surgery are commonly used in research laboratories by a range of students, technicians, and researchers. To meet the evolving requirements imposed by international legislation, and to promote the implementation of 3R rules (replacement, reduction, and refinement) by reducing experimental error, animal morbidity, and mortality, it is essential that standard operating procedures and proper conduct following such complex surgeries be precisely described and respected. The present report shows how refinements of our own neurosurgical techniques over decades, have significantly reduced the number of animals (rats) used in experiments and improved the animals’ well-being during the post-surgical recovery period. The current pre-, per-, and post-surgical procedures used in our laboratory are detailed. We describe the practical aspects of stereotaxic neurosurgery that have been refined in our laboratory since 1992 and that cover various areas including appropriate anesthesia and pain management during and after surgery, methods to determine the stereotaxic coordinates, and the best approach to the target brain structure. The application of these optimal surgical methods that combine reliable and reproducible results with an acute awareness of ethics and animal welfare leads to a significant reduction in the number of animals included in experimental research in accordance with ethical and regulatory rules as required by the European Directive on laboratory animal welfare.

[1]  T. Steckler,et al.  Good Research Practice: Lessons from Animal Care and Use. , 2019, Handbook of experimental pharmacology.

[2]  P. Duchamp-Viret,et al.  Microdialysis Unveils the Role of the α2-Adrenergic System in the Basolateral Amygdala during Acquisition of Conditioned Odor Aversion in the Rat. , 2018, ACS chemical neuroscience.

[3]  Maree T. Smith,et al.  In vivo profiling of four centrally administered opioids for antinociception, constipation and respiratory depression: Between‐colony differences in Sprague Dawley rats , 2018, Clinical and experimental pharmacology & physiology.

[4]  B. Nuttin,et al.  Stereotaxy in rat models: Current state of the art, proposals to improve targeting accuracy and reporting guideline , 2017, Behavioural Brain Research.

[5]  Nima Azimi,et al.  Discrepancies in stereotaxic coordinate publications and improving precision using an animal-specific atlas , 2017, Journal of Neuroscience Methods.

[6]  B. Tyler,et al.  Association of nausea with buprenorphine analgesia for rats , 2017, Lab Animal.

[7]  R. Gervais,et al.  Respective role of the dorsal hippocampus and the entorhinal cortex during the recombination of previously learned olfactory–tactile associations in the rat , 2017, Learning & memory.

[8]  M. Majchrzak,et al.  The entorhinal cortex is involved in conditioned odor and context aversions , 2015, Front. Neurosci..

[9]  J. Henke,et al.  Influence of repeated anaesthesia on physiological parameters in male Wistar rats: a telemetric study about isoflurane, ketamine-xylazine and a combination of medetomidine, midazolam and fentanyl , 2014, BMC Veterinary Research.

[10]  J. D. McGaugh,et al.  Noradrenergic influences in the basolateral amygdala on inhibitory avoidance memory are mediated by an action on α2-adrenoceptors , 2014, Psychoneuroendocrinology.

[11]  R. Gervais,et al.  Involvement of the lateral entorhinal cortex for the formation of cross‐modal olfactory‐tactile associations in the rat , 2014, Hippocampus.

[12]  P. Duchamp-Viret,et al.  The orexin component of fasting triggers memory processes underlying conditioned food selection in the rat , 2014, Learning & memory.

[13]  Benno Roozendaal,et al.  Rodent stereotaxic surgery and animal welfare outcome improvements for behavioral neuroscience. , 2012, Journal of visualized experiments : JoVE.

[14]  Michael R. Dashwood,et al.  Endothelin-1 as a neuropeptide: neurotransmitter or neurovascular effects? , 2009, Journal of Cell Communication and Signaling.

[15]  P. Flecknell,et al.  Reported analgesic and anaesthetic administration to rodents undergoing experimental surgical procedures , 2009, Laboratory animals.

[16]  B. Ferry,et al.  Differential effects of β-adrenergic receptor blockade in basolateral amygdala or insular cortex on incidental and associative taste learning , 2008, Neurobiology of Learning and Memory.

[17]  James L McGaugh,et al.  Involvement of basolateral amygdala alpha2-adrenoceptors in modulating consolidation of inhibitory avoidance memory. , 2008, Learning & memory.

[18]  J. Hekmatpanah Cerebral microvessel perfusion and pathologic alteration of the brain during drowsiness and coma caused by brain tumor: a laboratory study on rats. , 2007, Surgical neurology.

[19]  M. Majchrzak,et al.  Selective involvement of the lateral entorhinal cortex in the control of the olfactory memory trace during conditioned odor aversion in the rat. , 2006, Behavioral neuroscience.

[20]  A. Marchand,et al.  Entorhinal cortex lesions disrupt fear conditioning to background context but spare fear conditioning to a tone in the rat , 2006, Hippocampus.

[21]  H. Litvan,et al.  Pitfalls and challenges when assessing the depth of hypnosis during general anaesthesia by clinical signs and electronic indices , 2004, Acta anaesthesiologica Scandinavica.

[22]  N. Harrison Knockin' on the door of general anesthetic mechanisms: but will U.S. researchers be shut out? , 2003, Anesthesia and analgesia.

[23]  B. Antkowiak,et al.  General anesthetic actions in vivo strongly attenuated by a point mutation in the GABAA receptor β3 subunit , 2003, FASEB journal : official publication of the Federation of American Societies for Experimental Biology.

[24]  P A Flecknell,et al.  Buprenorphine: a reappraisal of its antinociceptive effects and therapeutic use in alleviating post-operative pain in animals , 2002, Laboratory animals.

[25]  P. Rehbinder,et al.  Recommendations for the health monitoring of rodent and rabbit colonies in breeding and experimental units , 2002, Laboratory animals.

[26]  J. D. McGaugh,et al.  Basolateral Amygdala–Nucleus Accumbens Interactions in Mediating Glucocorticoid Enhancement of Memory Consolidation , 2001, The Journal of Neuroscience.

[27]  J. D. McGaugh,et al.  Clenbuterol Administration into the Basolateral Amygdala Post-training Enhances Retention in an Inhibitory Avoidance Task , 1999, Neurobiology of Learning and Memory.

[28]  Claude Messier,et al.  New Techniques in Stereotaxic Surgery and Anesthesia in the Mouse , 1999, Pharmacology Biochemistry and Behavior.

[29]  J. D. McGaugh,et al.  Involvement of alpha1-adrenoceptors in the basolateral amygdala in modulation of memory storage. , 1999, European journal of pharmacology.

[30]  P. Luppi,et al.  Electrophysiological evidence that noradrenergic neurons of the rat locus coeruleus are tonically inhibited by GABA during sleep , 1998, The European journal of neuroscience.

[31]  L. Jarrard,et al.  Facilitation of conditioned odor aversion by entorhinal cortex lesions in the rat. , 1996, Behavioral neuroscience.

[32]  B. Ferry,et al.  Neuroanatomical and Functional Specificity of the Basolateral Amygdaloid Nucleus in Taste-Potentiated Odor Aversion , 1995, Neurobiology of Learning and Memory.

[33]  P A Flecknell,et al.  The effects of buprenorphine, nalbuphine and butorphanol alone or following halothane anaesthesia on food and water consumption and locomotor movement in rats , 1992, Laboratory animals.

[34]  A. Cowan,et al.  THE ANIMAL PHARMACOLOGY OF BUPRENORPHINE, AN ORIPAVINE ANALGESIC AGENT , 1977, British journal of pharmacology.

[35]  W. Russell,et al.  The Principles of Humane Experimental Technique , 1960 .

[36]  J. Galligan,et al.  The Rat in Neuroscience Research , 2020 .

[37]  Matthias Stefan Eggel,et al.  The logic, methodological and practical limitations in the current benefit concept in the harm-benefit-analysis : an investigation in the context of the EU directive 2010/63 on the protection of animals used for scientific purposes , 2018 .

[38]  A. Foundas,et al.  The Cerebral Vascular System , 2008 .

[39]  J L McGaugh,et al.  Basolateral amygdala noradrenergic influences on memory storage are mediated by an interaction between beta- and alpha1-adrenoceptors. , 1999, The Journal of neuroscience : the official journal of the Society for Neuroscience.

[40]  T. Scheck,et al.  The Vascular System of the Rat Cerebral Cortex—Basic Pattern of Leptomeningeal Vessels and Numerical Densities of Neocortical Arteries and Veins , 1983 .

[41]  G. Paxinos,et al.  The Rat Brain in Stereotaxic Coordinates , 1983 .