ION CHANNELS, TRANSPORTERS Andreas Leffler Æ Raimund I. Herzog Sulayman D. Dib-Hajj Æ Stephen G. Waxman Theodore R. Cummins Pharmacological properties of neuronal TTX-resistant sodium channels and the role of a critical serine pore residue Received: 15 February 2005 / Accepted: 29 April 2005 / Published online: 25 June 2005 Ó Springer-Verlag 2005 Abstract Voltage-gated sodium channels can be char- acterized by their sensitivity to inhibitors. Na v 1.5 is sensitive to block by cadmium and extracellular QX-314, but relatively insensitive to tetrodotoxin and saxitoxin. Na v 1.4 is tetrodotoxin- and saxitoxin-sensitive but resistant to cadmium and extracellular QX-314. Na v 1.8 and Na v 1.9 generate slowly inactivating (I TTXr-Slow ) and persistent (I TTXr-Per ) currents in sensory neurons that are tetrodotoxin-resistant. Tetrodotoxin sensitivity is largely determined by the identity of a single residue; tyrosine 401 in Na v 1.4, cysteine 374 in Na v 1.5 and serine 356 and 355 in Na v 1.8 and Na v 1.9. We asked whether Na v 1.8 and Na v 1.9 share other pharmacological properties as a result of this serine residue. I TTXr-Slow and I TTXr-Per were saxitoxin-resistant and resistant to internal QX-314. I TTXr-Slow was also resistant to external QX-314 and displayed a approximately fourfold higher sensitivity than I TTXr-Per to cadmium. The impact of the serine residue was investigated by replacing tyrosine 401 in Na v 1.4 with serine (Y401S) or cysteine (Y401C). Both mutants were resistant to tetrodotoxin and saxitoxin. Whereas Na v 1.4-Y401C displayed an increased sensi- tivity to cadmium and extracellular QX-314, the serine substitution did not alter the sensitivity of Na v 1.4 to cadmium or QX-314. Our data indicates that while the serine residue determines the sensitivity of I TTXr-Slow and I TTXr-Per to tetrodotoxin and saxitoxin, it does not determine their insensitivity to QX-314 or their differ- ential sensitivities to cadmium. Keywords Sodium channel antagonist Æ Sodium current Æ Saxitoxin Æ Tetrodotoxin Æ Tetrodotoxin- resistant channels Æ Sodium channel Abbreviations Na v 1.4: Voltage-gated sodium channel a-subunit from skeletal muscle Æ Na v 1.8: SNS or PN3 Æ Na v 1.9: NaN or SNS2 Æ NGF: Nerve growth factor Æ BDNF: Brain-derived neurotrophic factor Æ NT4/5: Neurotrophin 4/5 Æ TrkB: Tyrosine kinase B Æ TTX: tetrodotoxin Æ STX: Saxitoxin Introduction The TTX-resistant (TTXr) voltage-gated sodium chan- nels Na v 1.8 and Na v 1.9 are predominantly expressed in spinal sensory neurons [1, 13, 30]. Electrophysiological studies on cultured dorsal root ganglion (DRG) neurons from adult rodents and humans have revealed two dis- tinct TTXr sodium currents, a slowly inactivating cur- rent (I TTXr-Slow ) and a persistent current (I TTXr-Per )[8, 9, 11, 14, 24]. Studies on transgenic mice lacking Na v 1.8 and Na v 1.9 respectively have revealed the molecular determinant for I TTXr-Slow as Na v 1.8 [2, 9] and Na v 1.9 for I TTXr-Per [20]. The expression and functional prop- erties of both I TTXr-Slow and I TTXr-Per suggest that they can substantially contribute to the electro-responsive- ness of nociceptive neurons and thus they are both considered to be attractive targets for the development of new analgesic drugs [4, 12, 18, 23, 33]. This work was supported in part by grants from the Medical Re- search Service and the Rehabilitation Research Service, Depart- ment of Veterans Affairs, and by grants from The Eastern Paralyzed Veterans Association and the Paralyzed Veterans of America. A. Leffler Klinik fu¨r Ana¨sthesiologie, Friedrich-Alexander-Universita¨t Erlangen-Nuremberg, Krankenhausstr. 12, 91054 Erlangen, Germany R. I. Herzog Æ S. D. Dib-Hajj Æ S. G. Waxman Department of Neurology, Yale University School of Medicine, New Haven, CT, USA T. R. Cummins (&) Department of Pharmacology and Toxicology, Stark Neurosciences Research Center, Indiana University School of Medicine, R2-402 950 W Walnut Street, Indianapolis, IN 46202, USA E-mail: trcummin@iupui.edu Tel.: +1-317-2789342 Fax: +1-317-2785849 Pflugers Arch – Eur J Physiol (2005) 451: 454–463 DOI 10.1007/s00424-005-1463-x