We provide here detailed electrophysiological protocols to study voltage-gated sodium channels and to investigate how wild-type and mutant channels influence firing properties of transfected mammalian dorsal root ganglion (DRG) neurons. Whole-cell voltage-clamp recordings permit us to analyze kinetic and voltage-dependence properties of ion channels and to determine the effect and mode of action of pharmaceuticals on specific channel isoforms. They also permit us to analyze the role of individual sodium channels and their mutant derivatives in regulating firing of DRG neurons. Five to ten cells can be recorded daily, depending on the extent of analysis that is required. Because of different internal solutions that are used in voltage-clamp and current-clamp recordings, only limited information can be obtained from recording the same neuron in both modes. These electrophysiological studies help to elucidate the role of specific channels in setting threshold and suprathreshold responses of neurons, under normal and pathological conditions.
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Raman, I.M., Sprunger, L.K., Meisler, M.H. & Bean, B.P. Altered subthreshold sodium currents and disrupted firing patterns in Purkinje neurons of Scn8a mutant mice. Neuron 19, 881–891 (1997).
Raman, I.M. & Bean, B.P. Resurgent sodium current and action potential formation in dissociated cerebellar Purkinje neurons. J. Neurosci. 17, 4517–4526 (1997).
Cummins, T.R., Dib-Hajj, S.D., Herzog, R.I. & Waxman, S.G. Nav1.6 channels generate resurgent sodium currents in spinal sensory neurons. FEBS Lett. 579, 2166–2170 (2005).
Akopian, A.N., Sivilotti, L. & Wood, J.N. A tetrodotoxin-resistant voltage-gated sodium channel expressed by sensory neurons. Nature 379, 257–262 (1996).
Djouhri, L. et al. The TTX-resistant sodium channel Nav1.8 (SNS/PN3): expression and correlation with membrane properties in rat nociceptive primary afferent neurons. J. Physiol. (Lond.) 550, 739–752 (2003).
Sangameswaran, L. et al. Structure and function of a novel voltage-gated, tetrodotoxin-resistant sodium channel specific to sensory neurons. J. Biol. Chem. 271, 5953–5956 (1996).
Choi, J.S., Dib-Hajj, S.D. & Waxman, S. Differential slow inactivation and use-dependent inhibition of Nav1.8 channels contribute to distinct firing properties in IB4+ and IB4− DRG neurons. J. Neurophysiol. 97, 1258–1265 (2007).
Dib-Hajj, S.D., Cummins, T.R., Black, J.A. & Waxman, S.G. From genes to pain: Nav1.7 and human pain disorders. Trends Neurosci. 30, 555–563 (2007).
Rush, A.M. et al. A single sodium channel mutation produces hyper- or hypoexcitability in different types of neurons. Proc. Natl. Acad. Sci. USA 103, 8245–8250 (2006).
Dib-Hajj, S. et al. Transfection of rat or mouse neurons by biolistics or electroporation. Nat. Protoc 4, 1118–1127 (2009); DOI: 10.1038/nprot.2009.90.
Marty, A. & Neher, E. Tight-Seal Whole-cell Recording (Plenum, New York, 1995).
Hamill, O.P., Marty, A., Neher, E., Sakmann, B. & Sigworth, F.J. Improved patch-clamp techniques for high-resolution current recording from cells and cell-free membrane patches. Pflugers Arch. 391, 85–100 (1981).
Molleman, A. Patch Clamping: An Introductory Guide to Patch Clamp Electrophysiology (John Wiley, West Sussex, UK, 2003).
Cummins, T.R. et al. Nav1.3 sodium channels: rapid repriming and slow closed-state inactivation display quantitative differences after expression in a mammalian cell line and in spinal sensory neurons. J. Neurosci. 21, 5952–5961 (2001).
Akopian, A.N. et al. The tetrodotoxin-resistant sodium channel SNS has a specialized function in pain pathways. Nat. Neurosci. 2, 541–548 (1999).
Fjell, J. et al. Differential role of GDNF and NGF in the maintenance of two TTX-resistant sodium channels in adult DRG neurons. Mol. Brain Res. 67, 267–282 (1999).
Leffler, A. et al. GDNF and NGF reverse changes in repriming of TTX-sensitive Na(+) currents following axotomy of dorsal root ganglion neurons. J. Neurophysiol. 88, 650–658 (2002).
Cummins, T.R. et al. A novel persistent tetrodotoxin-resistant sodium current in SNS-null and wild-type small primary sensory neurons. J. Neurosci. 19, RC43 (1999).
Herzog, R.I., Cummins, T.R., Ghassemi, F., Dib-Hajj, S.D. & Waxman, S.G. Distinct repriming and closed-state inactivation kinetics of Nav1.6 and Nav1.7 sodium channels in mouse spinal sensory neurons. J. Physiol. (Lond.) 551, 741–750 (2003).
Campbell, D.T. Single-channel current/voltage relationships of two kinds of Na+ channel in vertebrate sensory neurons. Pflugers Arch. 423, 492–496 (1993).
Motomura, H., Fujikawa, S., Tashiro, N., Ito, Y. & Ogata, N. Single-channel analysis of two types of Na+ currents in rat dorsal root ganglia. Pflugers Arch. 431, 221–229 (1995).
Rush, A.M., Brau, M.E., Elliott, A.A. & Elliott, J.R. Electrophysiological properties of sodium current subtypes in small cells from adult rat dorsal root ganglia. J. Physiol. (Lond.) 511, 771–789 (1998).
Roy, M.L., Reuveny, E. & Narahashi, T. Single-channel analysis of tetrodotoxin-sensitive and tetrodotoxin-resistant sodium channels in rat dorsal root ganglion neurons. Brain Res. 650, 341–346 (1994).
Coste, B., Osorio, N., Padilla, F., Crest, M. & Delmas, P. Gating and modulation of presumptive NaV1.9 channels in enteric and spinal sensory neurons. Mol. Cell Neurosci. 26, 123–134 (2004).
Saab, C.Y., Cummins, T.R. & Waxman, S.G. GTP(gammaS) increases Nav1.8 current in small-diameter dorsal root ganglia neurons. Exp. Brain Res. 152, 415–419 (2003).
Chen, Y. & Penington, N.J. Competition between internal AlF(4)(−) and receptor-mediated stimulation of dorsal raphe neuron G-proteins coupled to calcium current inhibition. J. Neurophysiol. 83, 1273–1282 (2000).
Coleman, D.E. et al. Structures of active conformations of Gi alpha 1 and the mechanism of GTP hydrolysis. Science 265, 1405–1412 (1994).
Murphy, A.J. & Hoover, J.C. Inhibition of the Na,K-ATPase by fluoride. Parallels with its inhibition of the sarcoplasmic reticulum CaATPase. J. Biol. Chem. 267, 16995–16700 (1992).
Ono, K. & Arita, M. Mechanism of fluoride action on the L-type calcium channel in cardiac ventricular myocytes. Cell Calcium 26, 37–47 (1999).
Partanen, S. Inhibition of human renal acid phosphatases by nephrotoxic micromolar concentrations of fluoride. Exp. Toxicol. Pathol. 54, 231–237 (2002).
Zeng, H. et al. Improved throughput of PatchXpress hERG assay using intracellular potassium fluoride. Assay Drug Dev. Technol. 6, 235–241 (2008).
Toledo-Aral, J.J., Brehm, P., Halegoua, S. & Mandel, G. A single pulse of nerve growth factor triggers long-term neuronal excitability through sodium channel gene induction. Neuron 14, 607–611 (1995).
Toledo-Aral, J.J. et al. Identification of PN1, a predominant voltage-dependent sodium channel expressed principally in peripheral neurons. Proc. Natl. Acad. Sci. USA 94, 1527–1532 (1997).
Rush, A.M., Cummins, T.R. & Waxman, S.G. Multiple sodium channels and their roles in electrogenesis within dorsal root ganglion neurons. J. Physiol. (Lond.) 579 (Part 1): 1–14 (2007).
Cummins, T.R., Dib-Hajj, S.D. & Waxman, S.G. Electrophysiological properties of mutant Nav1.7 sodium channels in a painful inherited neuropathy. J. Neurosci. 24, 8232–8236 (2004).
Cummins, T.R. & Waxman, S.G. Downregulation of tetrodotoxin-resistant sodium currents and upregulation of a rapidly repriming tetrodotoxin-sensitive sodium current in small spinal sensory neurons after nerve injury. J. Neurosci. 17, 3503–3514 (1997).
Dib-Hajj, S.D. et al. Gain-of-function mutation in Nav1.7 in familial erythromelalgia induces bursting of sensory neurons. Brain 128, 1847–1854 (2005).
We thank Dr. Jin Sung Choi, Dr. Xiaoyang Cheng, Lynda Tyrrell, Larry Macala, Shujun Liu, Rachel Blackman, Bart Toftness and other members of our groups for valuable assistance in development and refinement of the techniques described in this article. Work in S.G.W. Laboratory is supported in part by grants from the National Multiple Sclerosis Society and the Rehabilitation Research and Development Service and Medical Research Service, Department of Veterans Affairs. T.R.C. was supported by research grant NS053422 from the National Institutes of Health. The Center for Neuroscience and Regeneration Research is a Collaboration of the Paralyzed Veterans of America and the United Spinal Association with Yale University.
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Cummins, T., Rush, A., Estacion, M. et al. Voltage-clamp and current-clamp recordings from mammalian DRG neurons. Nat Protoc 4, 1103–1112 (2009) doi:10.1038/nprot.2009.91
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