title: Critical evaluation of the expression of gastrin-releasing peptide in dorsal root ganglia and spinal cord.
abstract: There are substantial disagreements about the expression of gastrin-releasing peptide (GRP) in sensory neurons and whether GRP antibody cross-reacts with substance P (SP). These concerns necessitate a critical revaluation of GRP expression using additional approaches. Here, we show that a widely used GRP antibody specifically recognizes GRP but not SP. In the spinal cord of mice lacking SP (Tac1KO), the expression of not only GRP but also other peptides, notably neuropeptide Y (NPY), is significantly diminished. We detectedGrpmRNA in dorsal root ganglias using reverse transcription polymerase chain reaction, in situ hybridization and RNA-seq. We demonstrated thatGrpmRNA and protein are upregulated in dorsal root ganglias, but not in the spinal cord, of mice with chronic itch. Few GRP(+)immunostaining signals were detected in spinal sections following dorsal rhizotomy and GRP(+)cell bodies were not detected in dissociated dorsal horn neurons. Ultrastructural analysis further shows that substantially more GRPergic fibers form synaptic contacts with gastrin releasing peptide receptor-positive (GRPR(+)) neurons than SPergic fibers. Our comprehensive study demonstrates that a majority of GRPergic fibers are of primary afferent origin. A number of factors such as low copy number ofGrptranscripts, small percentage of cells expressingGrp, and the use of an eGFP GENSAT transgenic as a surrogate for GRP protein have contributed to the controversy. Optimization of experimental procedures facilitates the specific detection of GRP expression in dorsal root ganglia neurons.
authors.affiliation: Center for the Study of Itch, Washington University School of Medicine, St. Louis, MO, USA Departments of Anesthesiology, Washington University School of Medicine, St. Louis, MO, USA.
authors.forename: Devin M
authors.initials: DM
authors.lastname: Barry
authors.affiliation: Center for the Study of Itch, Washington University School of Medicine, St. Louis, MO, USA Department of Anatomy, K. K. Leung Brain Research Centre, The Fourth Military Medical University, Xi'an, PR China.
authors.forename: Hui
authors.initials: H
authors.lastname: Li
authors.affiliation: Center for the Study of Itch, Washington University School of Medicine, St. Louis, MO, USA Departments of Anesthesiology, Washington University School of Medicine, St. Louis, MO, USA.
authors.forename: Xian-Yu
authors.initials: XY
authors.lastname: Liu
authors.affiliation: Center for the Study of Itch, Washington University School of Medicine, St. Louis, MO, USA Department of Neurosurgery, Xinqiao Hospital, Third Military Medical University, Chongqing, PR China.
authors.forename: Kai-Feng
authors.initials: KF
authors.lastname: Shen
authors.affiliation: Center for the Study of Itch, Washington University School of Medicine, St. Louis, MO, USA Guangdong Provincial Key Laboratory of Allergy & Clinical Immunology, The State Key Laboratory of Respiratory Disease, The Second Affiliated Hospital, Guangzhou Medical University, Guangdong, PR China.
authors.forename: Xue-Ting
authors.initials: XT
authors.lastname: Liu
authors.affiliation: Department of Anatomy, K. K. Leung Brain Research Centre, The Fourth Military Medical University, Xi'an, PR China.
authors.forename: Zhen-Yu
authors.initials: ZY
authors.lastname: Wu
authors.affiliation: Center for the Study of Itch, Washington University School of Medicine, St. Louis, MO, USA Departments of Anesthesiology, Washington University School of Medicine, St. Louis, MO, USA.
authors.forename: Admire
authors.initials: A
authors.lastname: Munanairi
authors.affiliation: Institute of Neuroscience, State Key Laboratory of Neuroscience, CAS Center for Excellence in Brain Science and Intelligence Technology, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences, Shanghai, China.
authors.forename: Xiao-Jun
authors.initials: XJ
authors.lastname: Chen
authors.affiliation: Center for the Study of Itch, Washington University School of Medicine, St. Louis, MO, USA Departments of Anesthesiology, Washington University School of Medicine, St. Louis, MO, USA.
authors.forename: Jun
authors.initials: J
authors.lastname: Yin
authors.affiliation: Institute of Neuroscience, State Key Laboratory of Neuroscience, CAS Center for Excellence in Brain Science and Intelligence Technology, Shanghai Institutes for Biological Sciences, Chinese Academy of Sciences, Shanghai, China.
authors.forename: Yan-Gang
authors.initials: YG
authors.lastname: Sun
authors.affiliation: Department of Anatomy, K. K. Leung Brain Research Centre, The Fourth Military Medical University, Xi'an, PR China.
authors.forename: Yun-Qing
authors.initials: YQ
authors.lastname: Li
authors.affiliation: Center for the Study of Itch, Washington University School of Medicine, St. Louis, MO, USA Departments of Anesthesiology, Washington University School of Medicine, St. Louis, MO, USA Departments of Psychiatry, Center for the Study of Itch, Washington University School of Medicine, St. Louis, MO, USA Departments of Developmental Biology, Center for the Study of Itch, Washington University School of Medicine, St. Louis, MO, USA chenz@wustl.edu deptanat@fmmu.edu.cn.
authors.forename: Zhou-Feng
authors.initials: ZF
authors.lastname: Chen
mesh_terms: D000595:Amino Acid Sequence; D000818:Animals; D000918:Antibody Specificity; D002452:Cell Count; D002478:Cells, Cultured; D002908:Chronic Disease; D005727:Ganglia, Spinal / Q000378:metabolism* / Q000473:pathology; D019886:Gastrin-Releasing Peptide / Q000737:chemistry / Q000235:genetics / Q000378:metabolism*; D017353:Gene Deletion; D005786:Gene Expression Regulation; D008297:Male; D008810:Mice, Inbred C57BL; D018345:Mice, Knockout; D020671:Posterior Horn Cells / Q000378:metabolism / Q000473:pathology; D011537:Pruritus / Q000235:genetics / Q000473:pathology; D012333:RNA, Messenger / Q000235:genetics / Q000378:metabolism; D018004:Receptors, Bombesin / Q000378:metabolism; D013116:Spinal Cord / Q000378:metabolism*; D013373:Substance P / Q000378:metabolism
chemical_list: D012333:RNA, Messenger; D018004:Receptors, Bombesin; D013373:Substance P; D019886:Gastrin-Releasing Peptide
keywords: dorsal root ganglia; gastrin-releasing peptide; itch; sensory neurons; spinal cord; substance P