Abstract-The excitability of pulmonary artery smooth muscle cells (PASMC) is regulated by potassium (K ϩ ) conductances. Although studies suggest that background K ϩ currents carried by 2-pore domain K ϩ channels are important regulators of resting membrane potential in PASMC, their role in human PASMC is unknown. Our study tested the hypothesis that TASK-1 leak K ϩ channels contribute to the K ϩ current and resting membrane potential in human PASMC. We used the whole-cell patch-clamp technique and TASK-1 small interfering RNA (siRNA). Noninactivating K ϩ current performed by TASK-1 K ϩ channels were identified by current characteristics and inhibition by anandamide and acidosis (pH 6.3), each resulting in significant membrane depolarization. Moreover, we showed that TASK-1 is blocked by moderate hypoxia and activated by treprostinil at clinically relevant concentrations. This is mediated via protein kinase A (PKA)-dependent phosphorylation of TASK-1. To further confirm the role of TASK-1 channels in regulation of resting membrane potential, we knocked down TASK-1 expression using TASK-1 siRNA. The knockdown of TASK-1 was reflected by a significant depolarization of resting membrane potential. Treatment of human PASMC with TASK-1 siRNA resulted in loss of sensitivity to anandamide, acidosis, alkalosis, hypoxia, and treprostinil. These results suggest that (1) TASK-1 is expressed in human PASMC; (2) TASK-1 is hypoxia-sensitive and controls the resting membrane potential, thus implicating an important role for TASK-1 K ϩ channels in the regulation of pulmonary vascular tone; and (3) treprostinil activates TASK-1 at clinically relevant concentrations via PKA, which might represent an important mechanism underlying the vasorelaxing properties of prostanoids and their beneficial effect in vivo. Key Words: pulmonary circulation Ⅲ potassium channels Ⅲ TASK-1 Ⅲ treprostinil Ⅲ hypoxic pulmonary vasoconstriction T he membrane potential of pulmonary artery smooth muscle cells (PASMC) is an important regulator of arterial tone. These cells have a resting membrane potential of approximately Ϫ65 to Ϫ50 mV in vitro, close to the predicted equilibrium potential for potassium (K ϩ ) ions. The opening of K ϩ channels in the PASMC membrane increases K ϩ efflux, which causes membrane hyperpolarization. This closes voltage-dependent Ca 2ϩ channels, decreasing Ca 2ϩ entry and leading to vasodilatation. Conversely, inhibition of K ϩ channels causes membrane depolarization, Ca 2ϩ entry, cell contraction, and vasoconstriction.Background or leak K ϩ -selective channels, as defined by a lack of time and voltage dependency, play an essential role in setting the resting membrane potential and input resistance in excitable cells. Two-pore domain K ϩ (2-PK) channels have been shown to conduct several leak K ϩ currents. The activity of 2-PK channels is strongly regulated by protons, protein kinases, and hypoxia. Alteration of K ϩ conductance can influence cellular activity via membrane potential changes.Both RT-PCR and Northern blot analyses p...

Sensory neurones with their cell bodies in dorsal root ganglia (DRG neurones) transmit various types of sensory information from the periphery to the spinal cord. DRG neurones are a physically and functionally heterogeneous population, with cell bodies in adult rats ranging in size from less than 20 to over 50 ìm diameter. During the last 20 years there has been increasing acceptance that cells of different size and sensory modality also express different mixes of voltage-gated ion channels, including pharmacologically and electrophysiologically distinct subtypes of voltage-gated Na¤ channels. The initial DRG Na¤ channel classification related to the effects of the Na¤ channel blocker tetrodotoxin (TTX) and included TTX-sensitive (TTX-S) and TTXresistant (TTX-R) currents (Kostyuk et al. 1981). The first TTX-R currents to be described were slower to activate and inactivate than their TTX-S counterparts and had higher voltage thresholds for both activation and inactivation (Kostyuk et al. 1981;Schwartz et al. 1990; Journal of Physiology (1998) 1. Whole-cell and single-channel Na¤ currents were recorded from small (ca. 20 ìm diameter) cells isolated from adult rat dorsal root ganglia (DRG). Currents were classified by their sensitivity to 0·3 ìÒ tetrodotoxin (TTX), electrophysiological properties and single-channel amplitude. Cells were classified according to the types of current recorded from them. 2. Type A cells expressed essentially pure TTX-sensitive (TTX-S) currents. Availability experiments with prepulse durations between 50 ms and 1 s gave a half-available voltage (Vh) of around −65 mV but the availability curves often had a complex shape, consistent with multiple inactivation processes. Measured inactivation time constants ranged from less than 1 ms to over 100 s, depending on the protocol used. 3. Cell types B and C each had, in addition to TTX-S currents, substantial and different TTXresistant (TTX-R) currents that we have designated TTX-R1 and TTX-R2, respectively. TTX-R1 currents had a 1 s Vh of −29 mV, showed little 1 Hz use dependence at −67 mV and recovered from the inactivation induced by a 60 ms depolarizing pulse with time constants of 1·6 ms (91%) and 908 ms. They also exhibited slow inactivation processes with component time constants around 10 and 100 s. TTX-R2 currents activated and inactivated at more negative potentials (1 s Vh = −46 mV), showed substantial 1 Hz use dependence and had inactivation (60 ms pulse) recovery time constants at −67 mV of 3·3 ms (58%) and 902 ms. 4. Type D cells had little or no current in 0·3 ìÒ TTX at a holding potential of −67 mV.Current amplitude increased on changing the holding potential to −107 mV. Type D cell currents had more hyperpolarized availability and I-V curves than even TTX-R2 currents and suggest the existence of TTX-R3 channels. 5. In outside-out patches with 250 mÒ external NaCl, the single-channel conductance (ã) of TTX-S channels was 19·5 pS and the potential for half-maximal activation (Va) was −45 mV. One population of TTX-R channels had...

Less negative membrane potential and repetitive firing have little effect on tetrodotoxin-resistant Na(+) current amplitude but increase their sensitivity to lidocaine, mexiletine, and amitriptyline so that concentrations after intravenous administration of these drugs can impair channel function. This may explain alleviation from pain by reducing firing frequency in ectopic sites without depressing central nervous or cardiac excitability.