Voltage-Gated Sodium Channels in Human Aortic Smooth Muscle Cells

Abstract

Whole-cell, voltage clamp methods were used to study inward currents in human aortic smooth muscle cells in culture. Cells were plated on glass coverslips, cultured in supplemented M-199 media with 5% serum and studied as primary cells and at passages 2–5. Inward currents were measured with a pipette solution containing Cs⁺ and TEA⁺ to block K⁺ currents and with 2.5 mM [Ca²⁺]₀ in the perfusate. Inward currents activated at about –50 mV, peaked at about –15 mV and reversed at about +30 mV. Values of peak inward current averaged 14.7 ± 3.3 pA/pF and cell capacitance averaged 124 ± 10 pF (n = 35). These currents activated rapidly with a time-to-peak current of 2.4 ± 0.3 ms at a test potential of –10 mV from a holding potential of –80 mV. The current also inactivated rapidly with a time course that could be described by two components with time constants of 1.8 ± 0.2 and 17.8 ± 3.5 ms at –10 mV. The currents decreased when extracellular Na⁺ was reduced and were completely inhibited by 50 nM tetrodotoxin (TTX), suggesting that they represented voltage-gated Na⁺ currents (I_Na). Activation curves were characterized with a V_0.5 = –16.6 ± 2.4 mV and a slope factor k = –5.2 ± 0.2 mV while inactivation curves were characterized with a V_0.5 = –60.9 ± 1.7 mV and a slope factor k = 8.8 ± 0.4 mV. Lowering external [Ca²⁺] to zero increased the maximum I_Na, shifted its voltage dependence in the hyperpolarizing direction and increased the rate of I_Na inactivation. Increasing external [Ca²⁺] or [Mg²⁺]decreased I_Na and slowed its rate of inactivation. These studies demonstrate the presence of voltage-gated Na⁺ channels with high TTX sensitivity that are modulated by extracellular divalent cations in human aortic smooth muscle cells maintained in cell culture. Window currents were found in the voltage range of –50 to –20 mV, suggesting that these channels could contribute to the resting membrane potential.

Contribution of Kv Channels to Phenotypic Remodeling of Human Uterine Artery Smooth Muscle Cells

“…14,15 Figure 1 shows average I/V curves and sample records (insets) obtained in acutely dispersed cells ( Figure 1A) and in cultured VSMCs ( Figure 1B) before and after paxilline application. As previously reported, 16 cultured cells exhibited tetrodotoxinsensitive inward Na ϩ currents, which were absent in acutely dispersed cells ( Figure 1B, inset). Depolarization elicited outward currents with an apparent threshold for activation positive to Ϫ40 mV, and paxilline decreased current amplitudes at potentials positive to ϩ30 mV.…”

Identification of functional voltage-gated Na+ channels in cultured human pulmonary artery smooth muscle cells

“…The inward I Na we observed in human PASMC possessed biophysical and pharmacological characteristics similar to those previously identified in other human VSMC [4,5,7,16,28]: sensitivity to TTX (EC 50 17.5 nM), -60 to -50 mV activation threshold, -15 to 0 mV peak amplitude potential, -70 to -65 mV half-inactivation voltage, -25 to -15 mV half-activation voltage, and activation (τ act ∼1 ms) and inactivation time (τ inact ≤ 4 ms) constants. Similarities in the window currents, -60 to -10 mV in 3human PASMC and other VSMC [4,7], also suggest that Na + channels are active over a wide range of physiological E m , and that they may contribute to the regulation of resting E m in human PASMC. However, in unpublished observations, we found that TTX did not significantly alter human PASMC E m (-41.2±2.6 mV, -46.2±3.1 mVin TTX, n=4, data not shown) indicating that Na + channel activity does not modulate E m in cultured normal human PASMC.…”

Characterization of a voltage-dependent Na+current in human esophageal smooth muscle

“…Expression studies in oocytes showed that, much like neuronal Na ϩ channels, transient and sustained currents can be mediated by a single population of skeletal muscle Na ϩ channels (9). Therefore, although sustained currents have been observed in several smooth muscle systems (10,28), their presence cannot be used to distinguish between neuronal and skeletal muscle channel isoforms.…”

“…Voltage-dependent Na ϩ currents have been recorded in a number of smooth muscle tissues, including arterial (10), lymphatic (19), uterine (36,37), and vas deferens smooth muscle (6). In the gastrointestinal tract, Na ϩ current has been observed in the ileum (30), large intestine (35), and fundus (25).…”