Inhibition Of Voltage-gated Potassium Channels Research Articles

Marine compound rhizochalinin shows high in vitro and in vivo efficacy in castration resistant prostate cancer.

Development of drug resistance is an inevitable phenomenon in castration-resistant prostate cancer (CRPC) cells requiring novel therapeutic approaches. In this study, efficacy and toxicity of Rhizochalinin (Rhiz) – a novel sphingolipid-like marine compound – was evaluated in prostate cancer models, resistant to currently approved standard therapies. In vitro activity and mechanism of action of Rhiz were examined in the human prostate cancer cell lines PC-3, DU145, LNCaP, 22Rv1, and VCaP. Rhiz significantly reduced cell viability at low micromolar concentrations showing most pronounced effects in enzalutamide and abiraterone resistant AR-V7 positive cells. Caspase-dependent apoptosis, inhibition of pro-survival autophagy, downregulation of AR-V7, PSA and IGF-1 expression as well as inhibition of voltage-gated potassium channels were identified as mechanisms of action. Remarkably, Rhiz re-sensitized AR-V7 positive cells to enzalutamide and increased efficacy of taxanes.In vivo activity and toxicity were evaluated in PC-3 and 22Rv1 NOD SCID mouse xenograft models using i.p. administration. Rhiz significantly reduced growth of PC-3 and 22Rv1 tumor xenografts by 27.0% (p = 0.0156) and 46.8% (p = 0.047) compared with controls with an increased fraction of tumor cells showing apoptosis secondary to Rhiz exposure. In line with the in vitro data, Rhiz was most active in AR-V7 positive xenografts in vivo. In animals, no severe side effects were observed.In conclusion, Rhiz is a promising novel marine-derived compound characterized by a unique combination of anticancer properties. Its further clinical development is of high impact for patients suffering from drug resistant prostate cancer especially those harboring AR-V7 mediated resistance to enzalutamide and abiraterone.

Inhibition of voltage-gated potassium channels mediates uncarboxylated osteocalcin-regulated insulin secretion in rat pancreatic β cells

Insulin secretion from pancreatic β cells is important to maintain glucose homeostasis and is regulated by electrical activities. Uncarboxylated osteocalcin, a bone-derived protein, has been reported to regulate glucose metabolism by increasing insulin secretion, stimulating β cell proliferation and improving insulin sensitivity. But the underlying mechanisms of uncarboxylated osteocalcin-modulated insulin secretion remain unclear. In the present study, we investigated the relationship of uncarboxylated osteocalcin-regulated insulin secretion and voltage-gated potassium (KV) channels, voltage-gated calcium channels in rat β cells. Insulin secretion was measured by radioimmunoassay. Channel currents and membrane action potentials were recorded using the conventional whole-cell patch-clamp technique. Calcium imaging system was used to analyze intracellular Ca2+ concentration ([Ca2+]i). The data show that under 16.7mmol/l glucose conditions uncarboxylated osteocalcin alone increased insulin secretion and [Ca2+]i, but with no such effects on insulin secretion and [Ca2+]i in the presence of a KV channel blocker, tetraethylammonium chloride. In the patch-clamp experiments, uncarboxylated osteocalcin lengthened action potential duration and significantly inhibited KV currents, but had no influence on the characteristics of voltage-gated calcium channels. These results indicate that KV channels are involved in uncarboxylated osteocalcin-regulated insulin secretion in rat pancreatic β cells. By inhibiting KV channels, uncarboxylated osteocalcin prolongs action potential duration, increases intracellular Ca2+ concentration and finally promotes insulin secretion. This finding provides new insight into the mechanisms of osteocalcin-modulated insulin secretion.

Distinct and complementary roles for α and β isoenzymes of PKC in mediating vasoconstrictor responses to acutely elevated glucose.

We investigated the hypothesis that elevated glucose increases contractile responses in vascular smooth muscle and that this enhanced constriction occurs due to the glucose-induced PKC-dependent inhibition of voltage-gated potassium channels. Patch-clamp electrophysiology in rat isolated mesenteric arterial myocytes was performed to investigate the glucose-induced inhibition of voltage-gated potassium (Kv ) current. To determine the effects of glucose in whole vessel, wire myography was performed in rat mesenteric, porcine coronary and human internal mammary arteries. Glucose-induced inhibition of Kv was PKC-dependent and could be pharmacologically dissected using PKC isoenzyme-specific inhibitors to reveal a PKCβ-dependent component of Kv inhibition dominating between 0 and 10 mM glucose with an additional PKCα-dependent component becoming evident at concentrations greater than 10 mM. These findings were supported using wire myography in all artery types used, where contractile responses to vessel depolarization and vasoconstrictors were enhanced by increasing bathing glucose concentration, again with evidence for distinct and complementary PKCα/PKCβ-mediated components. Our results provide compelling evidence that glucose-induced PKCα/PKCβ-mediated inhibition of Kv current in vascular smooth muscle causes an enhanced constrictor response. Inhibition of Kv current causes a significant depolarization of vascular myocytes leading to marked vasoconstriction. The PKC dependence of this enhanced constrictor response may present a potential therapeutic target for improving microvascular perfusion following percutaneous coronary intervention after myocardial infarction in hyperglycaemic patients.

Different approaches for breast cancer: voltage gated potassıum channels and micrornas

Micro RNAs and voltage-gated potassium channels (VGPCs) both play critical roles in the development of cancer. We aimed to reveal the diversity of miR-126/126*, which effects angiogenesis and vascular development through the inhibition of VGPCs. In this study, potassium channel inhibitors, including tetraethylammonium (5mM), 4-aminopyridine (5mM), margatoxin (1µM) and astemizole (2µM), were applied to MCF-7 and MDA-MB-231 breast cancer cell lines. After totally isolating RNA from the cells, Real-Time Polymerase Chain Reaction was used in order to identify gene expressions. OneWay ANOVA was used for variation analyses, while Tukey HSD and Tamhane were used to assess whether multiple comparisons were statistically significant (P < 0.001). Our results showed an increase in miR-126/126* expressions after the channel inhibition of MCF-7 and MDA-MB-231 cell lines (P < 0.001). miR-126/126* expressions were increased using TEA, 4-AP and astemizole in both cell lines. miR-126/126* expressions were only increased through the use of margatoxin in MCF-7. miR-126/126* may interact with voltage-gated potassium channels. In our study, the inhibition of K channels using K channel blockers resulted in an increase of miR-126/126* expression. Therefore, our data suggested that there could be another perspective between K channels and non-coding RNAs in the development of breast cancer

Mechanism of Inhibition of Voltage-Gated Potassium Channels by Guanidine Compounds

Inhibition of presynaptic voltage-gated potassium (Kv) channels by guanidine underlies its use in the therapy for the neuromuscular diseases, myasthenic syndrome of Lambert-Eaton and botulism. The therapeutic use of guanidine is limited, however, due to side effects that accompany its administration. Therefore, the design of guanidine analogs with improved therapeutic indices is desirable. Progress towards this goal is hindered by the lack of knowledge of the mechanism by which these molecules inhibit Kv channels. We examined an array of possible mechanisms of inhibition of the Shaker Kv channel by guanidine, methyl guanidine and N,N-dimethyl guanidine, including charge screening, disruption of the protein-lipid interfaces, direct interaction with the voltage sensors and pore-binding. Our results demonstrate that guanidines bind within the intracellular pore of the channel, and perturb a hydrophobic subunit interface to stabilize a closed state of the channel. This mechanism provides a foundation for the design of guanidine analogs for the therapeutic intervention of neuromuscular diseases.

The glucagon-producing alpha cell: an electrophysiologically exceptional cell

Activation of potassium channels normally serves to reduce cellular activity but recent data indicate that the glucagon-secreting alpha cells are different in this respect and that inhibition of voltage-gated potassium channels results in a paradoxical inhibition of glucagon secretion. Here we discuss these findings and attempt to provide a model for the regulation of glucagon secretion that incorporates these observations.

Low potassium dextran lung preservation solution reduces reactive oxygen species production

Background Low potassium dextran lung preservation solution has reduced primary graft failure in animal and human studies. Though the mechanism of reducing primary graft failure is unknown, low potassium dextran differs most significantly from solutions such as Euro-Collins (EC) and University of Wisconsin in its potassium concentration. The aim of this study was to investigate the impact that potassium concentration in lung preservation solutions had on pulmonary arterial smooth muscle cell depolarization and production of reactive oxygen species. Methods Using isolated pulmonary artery smooth muscle cells from Sprague-Dawley rats, the patch-clamp technique was used to measure resting cellular membrane potential and whole cell potassium current. Measurements were recorded at base line and after exposure to low potassium dextran, EC, and University of Wisconsin solutions. Pulmonary arteries from rats were isolated from the main pulmonary artery to the fourth segmental branch. Arteries were placed into vials containing low potassium dextran, EC, low potassium EC, Celsior, and University of Wisconsin solutions with reactive oxygen species measured by lucigenin-enhanced chemiluminescence. Results Pulmonary artery smooth muscle cell membrane potentials had a significant depolarization when placed in the University of Wisconsin or EC solutions, with changes probably related to inhibition of voltage-gated potassium channels. Low potassium dextran solution did not alter the membrane potential. Production of reactive oxygen species as measured by chemiluminescence was significantly higher when pulmonary arteries were exposed to University of Wisconsin or EC solutions (51,289 ± 5,615 and 35,702 ± 4353 counts/0.1 minute, respectively) compared with low potassium dextran, Celsior, and low potassium EC (12,537 ± 3623, 13,717 ± 3,844 and 15,187 ± 3,792 counts/0.1 minute, respectively). Conclusions Preservation solutions with high potassium concentration are clearly able to depolarize the pulmonary artery smooth muscle cells and increase pulmonary artery reactive oxygen species production. Low potassium preservations solutions may limit reactive oxygen species production and thus reduce the incidence of primary graft failure in lung transplantation.

Rapid neuromodulation by cortisol in the rat paraventricular nucleus: an in vitro study.

1. We have used a range of in vitro electrophysiological techniques to investigate the mechanism of rapid cortisol neuromodulation of parvocellular neurones in the rat paraventricular nucleus. 2. In our study, we found that cortisol (10 microM) increased spontaneous action-current firing frequency to 193%. This effect was insensitive to the glucocorticoid intracellular-receptor antagonist mifepristone. 3. Cortisol (0.1-10 microM) had no detectable effects on whole-cell GABA current amplitudes, or GABA(A) single-channel kinetics. 4. Cortisol (10 microM) inhibited whole-cell potassium currents in parvocellular neurones by shifting the steady-state activation curve by 14 mV to the right. 5. Additionally, in a cell line expressing both the glucocorticoid intracellular receptor and recombinant, fast inactivating potassium channels (hKv1.3), cortisol (1 and 10 microM) inhibited potassium currents by shifting their steady-state activation curves to the right by 12 mV (10 microM cortisol). This effect was also insensitive to the cortisol antagonist, mifepristone. 6. These data suggest that inhibition of voltage-gated potassium channels may contribute to the rapid neuromodulatory effects of cortisol, possibly by direct interaction with the ion channel itself.

Voltage-gated potassium channels in human ductus arteriosus

We studied tone in the human ductus arteriosus and show that the constriction to oxygen is due to inhibition of voltage-gated potassium channels and, in the acute phase, is independent of endothelin-1.

Metabotropic glutamate receptor inhibition of visceral afferent potassium currents

The effects of metabotropic glutamate receptor activation (mGluR) on voltage-gated potassium currents have been characterized in visceral sensory afferent neurons. l-Glutamate is known to be a primary neurotransmitter in visceral afferents which terminate at the level of the nucleus of the solitary tract (NTS). Synaptic communication between these afferents and the NTS has been shown to involve both postsynaptic ionotropic and presynaptic metabotropic glutamate receptor activation. The purpose of the present study was to determine the effects of mGluR activation on voltage-gated potassium currents in visceral sensory neurons. Application of mGluR agonist t-ACPD inhibited both the peak and the steady state voltage-gated potassium current in 39 out of 56 visceral afferent neurons tested (70%) by 22.0 ± 3 and 22.8 ± 2%, respectively. Voltage and pharmacological protocols were utilized to isolate the potassium current affected by mGluR activation. Increasing the holding potential from −100 mV to −30 mV only partially attenuated the inhibitory effects of t-ACPD (decreased effect by 11%), suggesting that t-ACPD modulates both a voltage insensitive and a voltage-sensitive potassium current. In addition, 4-aminopyridine (5 μM) was applied to eliminate the 4-AP sensitive transient current. Also, this protocol only partially attenuated the inhibitory effects of t-ACPD (decreased effect by 6.3%), suggesting that mGluR activation inhibits both a 4-AP-sensitive and 4-AP-insensitive potassium current in visceral afferent neurons. Results from this study suggest that mGluRs may regulate visceral sensory afferent neuronal activity through inhibition of voltage-gated potassium channels.