The cardiac sodium current Na_v1.5 is functionally expressed in rabbit bronchial smooth muscle cells

Abstract

Bradley E, Webb TI, Hollywood MA, Sergeant GP, McHale NG, Thornbury KD. The cardiac sodium current Nav1.5 is functionally expressed in rabbit bronchial smooth muscle cells. Am J Physiol Cell Physiol 305: C427-C435, 2013. First published June 19, 2013 doi:10.1152/ajpcell.00034.2013.-A collagenase-proteinase mixture was used to isolate airway smooth muscle cells (ASMC) from rabbit bronchi, and membrane currents were recorded using the whole cell patch-clamp technique. Stepping from Ϫ100 mV to a test potential of Ϫ40 mV evoked a fast voltage-dependent Na ϩ current, sometimes with an amplitude of several nanoamperes. The current disappeared within 15 min of exposure to papain ϩ DTT (n ϭ 6). Comparison of the current in ASMC with current mediated by NaV1.5 ␣-subunits expressed in human embryonic kidney cells revealed similar voltage dependences of activation (V1/2 ϭ Ϫ42 mV for NaV1.5) and sensitivities to TTX (IC50 ϭ 1.1 and 1.2 M for ASMC and NaV1.5, respectively). The current in ASMC was also blocked by lidocaine (IC50 ϭ 160 M). Although veratridine, an agonist of voltage-gated Na ϩ channels, reduced the peak current by 33%, it slowed inactivation, resulting in a fourfold increase in sustained current (measured at 25 ms after onset). In current-clamp mode, veratridine prolonged evoked action potentials from 37 Ϯ 9 to 1,053 Ϯ 410 ms (n ϭ 8). Primers for NaV1.2-1.9 were used to amplify mRNA from groups of ϳ20 isolated ASMC and from whole bronchial tissue by RT-PCR. Transcripts for NaV1.2, NaV1.3, and NaV1.5-1.9 were detected in whole tissue, but only NaV1.2 and NaV1.5 were detected in single cells. We conclude that freshly dispersed rabbit ASMC express a fast voltage-gated Na ϩ current that is mediated mainly by the NaV1.5 subtype. sodium current; Nav1.5; airway smooth muscle; patch clamp VOLTAGE-GATED SODIUM CHANNELS (VGSC) are normally associated with excitable cells, such as neurons and skeletal and cardiac myocytes, where they are responsible for the initiation and propagation of the action potential. They are classified according to their pore-forming ␣-subunits, of which there are nine known subtypes, designated Na V 1.1-1.9, encoded by the SCN gene family (SCN1A-5A and SCN8A-11A) (5). Perhaps surprisingly, VGSC have also been found in some nonexcitable cell types, including Schwann cells, cultured fibroblasts, cultured vascular endothelial cells, and metastatic cancer cells, although their role in these cells is poorly understood (1,4,6,8,11,12,19,26). In freshly dispersed smooth muscle cells, VGSC were previously regarded as a rarity, but evidence for their existence in this cell type has been gradually accumulating. In phasic tissues, such as uterine, lymphatic, vas deferens, portal vein, and gastrointestinal smooth muscle, they may play a part in the generation and/or propagation of spontaneous electrical activity underlying myogenic contractions (14,15,29,31,38). However, they have also been found in a variety of tonic arterial smooth muscles (7,9,21,27), although mostly only in cultured cells, suggestin...

Veratridine modifies the gating of human voltage-gated sodium channel Nav1.7

“…Veratridine at 50-100 µM inhibits the peak current of Nav1.4, Nav1.5, Nav1.7, Nav1.2, and Nav1.6 in HEK293 cells to different extents and enhances the sustained inward current and tail current in all of these channels [11,41,42]. Our data confirmed these effects of veratridine on Nav1.7, suggesting that veratridine has a general effect on these three parameters in different subtypes of VGSCs.…”

Late cardiac sodium current can be assessed using automated patch-clamp

“…The APD90 was increased over twofold by veratridine 1 μM, with a good reversibility upon wash-out ( Table 2), and an even higher prolongation with 5 μM ( Figure 4), although in this case we were unable to accurately compute APD90 values for externally paced APs, due to their overlap with the next stimulus. This effect is consistent with the findings of previous studies, showing APD prolongation in bronchial smooth muscle cells upon veratridine application 32 , and the enhancement of AP burst generation by veratridine in rat trigeminal sensory neurons 33 . Furthermore, we have demonstrated that hERG inhibition does not play any role in the observed APD prolongation by veratridine at 1 or 5 μM, since significant hERG inhibition occurred with higher veratridine concentrations ( Figure 5).…”

Cellular Na+ handling mechanisms involved in airway smooth muscle contraction (Review)

“…The recent evidence presented by Bradley et al (65) regarding the expression of a voltage-dependent Na + current in freshly dispersed rabbit bronchial myocytes seemingly mediated by the Nav1.5 α subunit, cannot be overlooked. When this current in bronchial myocytes was pharmacologically altered (blocked with veratridine), their action potential was prolonged.…”