Cardiac sodium channels expressed in a peripheral neurotumor derived cell line, rt4-b8. Zeng, Dewan, John W. Kyle, Ruth L. Martin, Kelly S. Ambler, and Dorothy A. Hanck. Departments of Medicine and the Pharmacological and Physiological Sciences, The University of Chicago
APStracts 2:0423C, 1995.
RT4-B is one of several cell lines derived from a multipotent stem -cell line, RT4-AC, which originated from a rat peripheral neurotumor. Based on Northern blot and RNase protection experiments RT4-B8 cells have been proposed to express rat cardiac sodium channel mRNA as the major isoform. We report here direct electrophysiological evidence that the expressed voltage-gated Na channels in the RT4-B8 cell line are of the cardiac phenotype with no evidence for subpopulations expressing other Na channel isoforms. Current activation half-point (conductance) was -41 +/- 5 mV (n=7) and the steady-state voltage -dependent availability half-point was -89 +/- 1 mV. As expected for cardiac Na channels, the ED50 for tetrodotoxin block was 0.74 [mu]M, for saxitoxin (STX) 0.15 [mu]M, and for the class 2B divalent cation Cd2+ 67 [mu]M. Block was well described by single-site dose-response relationships with no indication of a subpopulation with "neuronal" affinity. Single channel conductance (140 mM Na+) was 10?pS and predicted the average number of channels open at peak INa to be 3 channels [mu]m-2. 3H-STX binding data were also consistent with a single population of low affinity STX binding sites and predicted channel density to be 11 sites [mu]m-2. No inwardly or outwardly rectifying K currents or Ca currents were detected electrophysiologically, although in some cells a small time -independent Cl- current was detected. RT-PCR of mRNA isolated from RT4-B8 cells demonstrated the presence of rat cardiac (rH1) and brain IIa [alpha]-subunit mRNA, as well as mRNA for the Na channel [beta]1 -subunit. Northern blot analysis confirmed the predominance of the rat cardiac Na mRNA compared to brain IIa. The [beta]1-subunit mRNA levels were significantly lower than detected in rat brain and rat heart mRNA, but were comparable to the low level of [beta]1-subunit mRNA detected in isolated rat ventricular myocytes. 

Received 24 August 1994; accepted in final form 6 November 1995.
APS Manuscript Number C503-4.
Article publication pending Am. J. Physiol. (Cell Physiology).
ISSN 1080-4757 Copyright 1995 The American Physiological Society.
Published in APStracts on 8 December 95