Electrophysiological characterization of voltage-gated Na+ current expressed in the highly metastatic Mat-LyLu cell line of rat prostate cancer

Abstract

Voltage-gated Na+ channels, classically associated with impulse conduction in excitable tissues, are also found in a variety of epithelial cell types where their possible functions are not known so well. We have previously reported expression of a voltage-gated Na+ channel specifically in the highly metastatic Mat-LyLu rat prostate cancer cell line; blockage of the current with tetrodotoxin (TTX) significantly reduced the invasiveness of the cells in vitro, suggesting that the channel may have a functional role in metastasis. The aim of the present study was to characterize this current using the whole-cell patch clamp recording technique, and compare it to Na+ currents found in various other tissues. The inward current of the Mat-LyLu cells was abolished completely, but reversibly, in Na+-free solution, confirming that Na+ was indeed the permeant ion. Activation occurred at -40 mV and currents reached a maximal amplitude at around 6 mV. Boltzmann fits to current activation and steady-state inactivation revealed that the currents were half activated at about -15 mV and half inactivated at -80 mV. Both current inactivation and recovery from inactivation followed a double-exponential time course with fast and slow components. The Na+ currents were highly sensitive to block by TTX (IC50 approximately 18 nM), whilst 1 microM mu-conotoxin GIIIA mostly had no effect. 100 microM Cd2+ also had no effect on the current, whilst 2.5 mM Cd2+, Mn2+, and Co2+ each caused a depolarizing shift in activation and a reduction in peak conductance of around 20%. In conclusion, the Na+ channel expressed in the highly metastatic Mat-LyLu cell line appeared to have electrophysiological and pharmacological properties of TTX-sensitive channels. Further work is needed, however, to elucidate the exact nature of the channel protein and the mechanism(s) of its involvement in cellular invasiveness.

Predominant expression of Kv1.3 voltage-gated K+ channel subunit in rat prostate cancer cell lines: electrophysiological, pharmacological and molecular characterisation

“…Tissue culture and electrophysiological recording Cell culture and electrophysiological recordings were performed as before [18,19], but with the following modifications. Patch pipettes (of resistance 5-15 MW) were filled with a solution containing (in mM) NaCl (5), KCl (145), MgCl 2 (2), CaCl 2 (1), HEPES (10) and EGTA (11), adjusted to pH 7.4 with 1 M KOH.…”

“…Specific details are given in the Results, the individual figure legends or have been explained previously [18,19].…”

“…Leak current subtraction was performed as described previously [18]. Equations used to calculate membrane conductance, reversal potential, steady-state activation/inactivation parameters, and the time-course of current inactivation or recovery from inactivation were as described previously [18], with the following additions:…”

“…The decay time-course of the current could be fitted by a single exponential function [18] for both cell lines and appeared to be largely independent of voltage in the range 20-60 mV. However, inactivation was significantly slower at more negative voltages.…”

Sodium Channels: Clones and Consequences

“…In contrast, no VGSC was ever recorded in the corresponding weakly metastatic cells: AT-2 (rat) and LNCaP (human). The pharmacological profile of the rodent VGSC suggested that it belonged to the tetrodotoxinsensitive (TTX -S) class of channels with an IC 50 of ~20 nM (Grimes & Djamgoz, 1998). A molecular biological study showed that adult skeletal muscle (rjhSkM1)-type VGSC mRNA was expressed in these cells (Diss et al 1998).…”