A novel polyclonal antibody specific for the Na_v1.5 voltage-gated Na⁺ channel 'neonatal' splice form

Voltage-gated Na⁺ channel (VGSC) diversity is achieved through a number of mechanisms: multiple subunits, multiple genes encoding the pore-forming VGSC α-subunit and multiple gene isoforms generated by alternative splicing. A major type of VGSCα alternative splicing is in D1:S3, which has been proposed to be developmentally regulated. We recently reported a D1:S3 spliced form of Na_v1.5 in human metastatic breast cancer cells. This novel 'neonatal' isoform differs from the counterpart 'adult' form at seven amino acids (in the extracellular loop between S3-S4 of D1). Here, we generated an anti-peptide polyclonal antibody, named NESOpAb, which specifically recognised 'neonatal' but not 'adult' Na_v1.5 when tested on cells specifically over-expressing one or other of these Na_v1.5 spliced forms. The antibody was used to investigate developmental expression of 'neonatal' Na_v1.5 (nNa_v1.5) in a range of mouse tissues by immunohistochemistry. Overall, the results were consistent with nNa _v1.5 protein being more abundantly expressed in selected tissues (particularly heart and brain) from neonate as compared to adult animals. Importantly, NESOpAb blocked functional nNa_v1.5 ion conductance when applied extracellularly at concentrations as low as 0.05 ng/ml. Possible biological and clinical applications of NESOpAb are discussed.