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Characterisation of Nav1.7 functional expression in rat dorsal root ganglia neurons by using an electrical field stimulation assay.
Mol Pain. 2017 Jan-Dec; 13:1744806917745179.MP

Abstract

Background The Nav1.7 subtype of voltage-gated sodium channels is specifically expressed in sensory and sympathetic ganglia neurons where it plays an important role in the generation and transmission of information related to pain sensation. Human loss or gain-of-function mutations in the gene encoding Nav1.7 channels (SCN9A) are associated with either absence of pain, as reported for congenital insensitivity to pain, or with exacerbation of pain, as reported for primary erythromelalgia and paroxysmal extreme pain disorder. Based on this important human genetic evidence, numerous drug discovery efforts are ongoing in search for Nav1.7 blockers as a novel therapeutic strategy to treat pain conditions. Results We are reporting here a novel approach to study Nav1.7 function in cultured rat sensory neurons. We used live cell imaging combined with electrical field stimulation to evoke and record action potential-driven calcium transients in the neurons. We have shown that the tarantula venom peptide Protoxin-II, a known Nav1.7 subtype selective blocker, inhibited electrical field stimulation-evoked calcium responses in dorsal root ganglia neurons with an IC50 of 72 nM, while it had no activity in embryonic hippocampal neurons. The results obtained in the live cell imaging assay were supported by patch-clamp studies as well as by quantitative PCR and Western blotting experiments that confirmed the presence of Nav1.7 mRNA and protein in dorsal root ganglia but not in embryonic hippocampal neurons. Conclusions The findings presented here point to a selective effect of Protoxin-II in sensory neurons and helped to validate a new method for investigating and comparing Nav1.7 pharmacology in sensory versus central nervous system neurons. This will help in the characterisation of the selectivity of novel Nav1.7 modulators using native ion channels and will provide the basis for the development of higher throughput models for enabling pain-relevant phenotypic screening.

Authors+Show Affiliations

1 Lilly Research Centre, 1539 Eli Lilly and Company , Windlesham, UK.1 Lilly Research Centre, 1539 Eli Lilly and Company , Windlesham, UK.2 Lilly Research Laboratories, 1539 Eli Lilly and Company , IN, USA.2 Lilly Research Laboratories, 1539 Eli Lilly and Company , IN, USA.2 Lilly Research Laboratories, 1539 Eli Lilly and Company , IN, USA.2 Lilly Research Laboratories, 1539 Eli Lilly and Company , IN, USA.2 Lilly Research Laboratories, 1539 Eli Lilly and Company , IN, USA.1 Lilly Research Centre, 1539 Eli Lilly and Company , Windlesham, UK.1 Lilly Research Centre, 1539 Eli Lilly and Company , Windlesham, UK.

Pub Type(s)

Journal Article

Language

eng

PubMed ID

29166836

Citation

Fouillet, Antoine, et al. "Characterisation of Nav1.7 Functional Expression in Rat Dorsal Root Ganglia Neurons By Using an Electrical Field Stimulation Assay." Molecular Pain, vol. 13, 2017, p. 1744806917745179.
Fouillet A, Watson JF, Piekarz AD, et al. Characterisation of Nav1.7 functional expression in rat dorsal root ganglia neurons by using an electrical field stimulation assay. Mol Pain. 2017;13:1744806917745179.
Fouillet, A., Watson, J. F., Piekarz, A. D., Huang, X., Li, B., Priest, B., Nisenbaum, E., Sher, E., & Ursu, D. (2017). Characterisation of Nav1.7 functional expression in rat dorsal root ganglia neurons by using an electrical field stimulation assay. Molecular Pain, 13, 1744806917745179. https://doi.org/10.1177/1744806917745179
Fouillet A, et al. Characterisation of Nav1.7 Functional Expression in Rat Dorsal Root Ganglia Neurons By Using an Electrical Field Stimulation Assay. Mol Pain. 2017 Jan-Dec;13:1744806917745179. PubMed PMID: 29166836.
* Article titles in AMA citation format should be in sentence-case
TY - JOUR T1 - Characterisation of Nav1.7 functional expression in rat dorsal root ganglia neurons by using an electrical field stimulation assay. AU - Fouillet,Antoine, AU - Watson,Jake F, AU - Piekarz,Andrew D, AU - Huang,Xiaofang, AU - Li,Baolin, AU - Priest,Birgit, AU - Nisenbaum,Eric, AU - Sher,Emanuele, AU - Ursu,Daniel, Y1 - 2017/11/22/ PY - 2017/11/24/pubmed PY - 2018/7/24/medline PY - 2017/11/24/entrez KW - Nav1.7 KW - Sodium channels KW - calcium imaging KW - dorsal root ganglia neurons KW - electrical field stimulation KW - protoxin-II SP - 1744806917745179 EP - 1744806917745179 JF - Molecular pain JO - Mol Pain VL - 13 N2 - Background The Nav1.7 subtype of voltage-gated sodium channels is specifically expressed in sensory and sympathetic ganglia neurons where it plays an important role in the generation and transmission of information related to pain sensation. Human loss or gain-of-function mutations in the gene encoding Nav1.7 channels (SCN9A) are associated with either absence of pain, as reported for congenital insensitivity to pain, or with exacerbation of pain, as reported for primary erythromelalgia and paroxysmal extreme pain disorder. Based on this important human genetic evidence, numerous drug discovery efforts are ongoing in search for Nav1.7 blockers as a novel therapeutic strategy to treat pain conditions. Results We are reporting here a novel approach to study Nav1.7 function in cultured rat sensory neurons. We used live cell imaging combined with electrical field stimulation to evoke and record action potential-driven calcium transients in the neurons. We have shown that the tarantula venom peptide Protoxin-II, a known Nav1.7 subtype selective blocker, inhibited electrical field stimulation-evoked calcium responses in dorsal root ganglia neurons with an IC50 of 72 nM, while it had no activity in embryonic hippocampal neurons. The results obtained in the live cell imaging assay were supported by patch-clamp studies as well as by quantitative PCR and Western blotting experiments that confirmed the presence of Nav1.7 mRNA and protein in dorsal root ganglia but not in embryonic hippocampal neurons. Conclusions The findings presented here point to a selective effect of Protoxin-II in sensory neurons and helped to validate a new method for investigating and comparing Nav1.7 pharmacology in sensory versus central nervous system neurons. This will help in the characterisation of the selectivity of novel Nav1.7 modulators using native ion channels and will provide the basis for the development of higher throughput models for enabling pain-relevant phenotypic screening. SN - 1744-8069 UR - https://www.unboundmedicine.com/medline/citation/29166836/Characterisation_of_Nav1_7_functional_expression_in_rat_dorsal_root_ganglia_neurons_by_using_an_electrical_field_stimulation_assay_ L2 - http://journals.sagepub.com/doi/full/10.1177/1744806917745179?url_ver=Z39.88-2003&rfr_id=ori:rid:crossref.org&rfr_dat=cr_pub=pubmed DB - PRIME DP - Unbound Medicine ER -