Surgical extraction of human dorsal root ganglia from organ donors and preparation of primary sensory neuron cultures.
Primary cultures of rodent sensory neurons are widely used to investigate the cellular and molecular mechanisms involved in pain, itch, nerve injury and regeneration. However, translation of these preclinical findings may be greatly improved by direct validation in human tissues. We have developed an approach to extract and culture human sensory neurons in collaboration with a local organ procurement organization (OPO). Here we describe the surgical procedure for extraction of human dorsal root ganglia (hDRG) and the necessary modifications to existing culture techniques to prepare viable adult human sensory neurons for functional studies. Dissociated sensory neurons can be maintained in culture for >10 d, and they are amenable to electrophysiological recording, calcium imaging and viral gene transfer. The entire process of extraction and culturing can be completed in <7 h, and it can be performed by trained graduate students. This approach can be applied at any institution with access to organ donors consenting to tissue donation for research, and is an invaluable resource for improving translational research.
Washington University Pain Center and Department of Anesthesiology, Washington University School of Medicine, St. Louis, Missouri, USA.|Medical Scientist Training Program, Washington University in St. Louis, St. Louis, Missouri, USA.|Neurosciences Program, Washington University in St. Louis, St. Louis, Missouri, USA.
Manouela V
MV
Valtcheva
Washington University Pain Center and Department of Anesthesiology, Washington University School of Medicine, St. Louis, Missouri, USA.
Bryan A
BA
Copits
Washington University Pain Center and Department of Anesthesiology, Washington University School of Medicine, St. Louis, Missouri, USA.|Pain Research Center, Department of Anesthesiology, University of Cincinnati College of Medicine, Cincinnati, Ohio, USA.
Steve
S
Davidson
Washington University Pain Center and Department of Anesthesiology, Washington University School of Medicine, St. Louis, Missouri, USA.|Neurosciences Program, Washington University in St. Louis, St. Louis, Missouri, USA.
Tayler D
TD
Sheahan
Washington University Pain Center and Department of Anesthesiology, Washington University School of Medicine, St. Louis, Missouri, USA.|Developmental, Regenerative, and Stem Cell Biology Program, Washington University in St. Louis, St. Louis, Missouri, USA.
Melanie Y
MY
Pullen
Washington University Pain Center and Department of Anesthesiology, Washington University School of Medicine, St. Louis, Missouri, USA.
Jordan G
0000-0001-8295-0664
JG
McCall
Department of Neuroscience, Washington University School of Medicine, St. Louis, Missouri, USA.
Krikor
K
Dikranian
Washington University Pain Center and Department of Anesthesiology, Washington University School of Medicine, St. Louis, Missouri, USA.
Robert W
0000-0002-5428-4251
RW
Gereau
D002118:Calcium / Q000378:metabolism; D018929:Cell Culture Techniques / Q000379:methods; D002469:Cell Separation / Q000379:methods; D002478:Cells, Cultured; D005727:Ganglia, Spinal / Q000166:cytology* / Q000601:surgery*; D018014:Gene Transfer Techniques; D006801:Humans; D011984:Sensory Receptor Cells / Q000166:cytology* / Q000378:metabolism; D009927:Tissue and Organ Procurement / Q000379:methods*; D014161:Transduction, Genetic / Q000379:methods; D014780:Viruses / Q000235:genetics
D002118:Calcium