Cytoplasmic polyamines as permeant blockers and modulators of the voltage-gated sodium channel

We report that voltage-gated Na+ channels (Na-V) from rat muscle (mu1) expr essed in HEK293 cells exhibit anomalous rectification of whole-cell outward current under conditions of symmetrical Na+. This behavior gradually fades with time after membrane break-in, as if a diffusible blocking substance i n the cytoplasm is slowly diluted by the pipette solution. The degree of su ch block and rectification is markedly altered by various mutations of the conserved Lys(III) residue in Domain III of the Na, channel selectivity fil ter (DEKA locus), a principal determinant of inorganic ion selectivity and organic cation permeation. Using whole-cell and macropatch recording techni ques, we show that two ubiquitous polyamines, spermine and spermidine, are potent voltage-dependent cytoplasmic blockers of mu1 Na-V current that exhi bit relief of block at high positive voltage, a phenomenon that is also enh anced by certain mutations of the Lys(III) residue. In addition, we find th at:polyamines alter the apparent rate of macroscopic inactivation and exhib it a use-dependent blocking phenomenon reminiscent of the action of local a nesthetics. In the presence of a physiological Na+/K+ gradient, spermine al so inhibits inward Na-V current and shifts the voltage dependence of activa tion and inactivation. Similarities between the endogenous blocking phenome non observed in whole cells and polyamine blocle characterized in excised p atches suggest that polyamines or related metabolites may function as endog enous modulators of Na, channel activity.