Modulation of K(v)7 potassium channels by a novel opener pyrazolo[1,5-a]pyrimidin-7(4H)-one compound QO-58.

BACKGROUND AND PURPOSE: Modulation of K(v)7/M channel function represents a relatively new strategy to treat neuronal excitability disorders such as epilepsy and neuropathic pain. We designed and synthesized a novel series of pyrazolo[1,5-a] pyrimidin-7(4H)-one compounds, which activate K(v)7 channels. Here, we characterized the effects of the lead compound, QO-58, on K(v)7 channels and investigated its mechanism of action. EXPERIMENTAL APPROACH: A perforated whole-cell patch technique was used to record K(v)7 currents expressed in mammalian cell lines and M-type currents from rat dorsal root ganglion neurons. The effects of QO-58 in a rat model of neuropathic pain, chronic constriction injury (CCI) of the sciatic nerve, were also examined. KEY RESULTS: QO-58 increased the current amplitudes, shifted the voltage-dependent activation curve in a more negative direction and slowed the deactivation of K(v)7.2/K(v)7.3 currents. QO-58 activated K(v)7.1, K(v)7.2, K(v)7.4 and K(v)7.3/K(v)7.5 channels with a more selective effect on K(v)7.2 and K(v)7.4, but little effect on K(v)7.3. The mechanism of QO-58's activation of K(v)7 channels was clearly distinct from that used by retigabine. A chain of amino acids, Val(224)Val(225)Tyr(226), in K(v)7.2 was important for QO-58 activation of this channel. QO-58 enhanced native neuronal M currents, resulting in depression of evoked action potentials. QO-58 also elevated the pain threshold of neuropathic pain in the sciatic nerve CCI model. CONCLUSIONS AND IMPLICATIONS: The results indicate that QO-58 is a potent modulator of K(v)7 channels with a mechanism of action different from those of known K(v)7 openers. Hence, QO-58 shows potential as a treatment for diseases associated with neuronal hyperexcitability.

Characteristics and molecular basis of celecoxib modulation on K(v)7 potassium channels.

BACKGROUND AND PURPOSE: Celecoxib is a selective cyclooxygenase-2 (COX-2) inhibitor used for the treatment of pain and inflammation. Emerging and accumulating evidence suggests that celecoxib can affect cellular targets other than COX, such as ion channels. In this study, we characterized the effects of celecoxib on K(v)7 K(+) channels and compared its effects with the well-established K(v)7 channel opener retigabine. EXPERIMENTAL APPROACH: A perforated whole-cell patch technique was used to record K(v)7currents expressed in HEK 293 cells and M-type currents from rat superior cervical ganglion neurons. KEY RESULTS: Celecoxib enhanced K(v)7.2-7.4, K(v)7.2/7.3 and K(v)7.3/7.5 currents but inhibited K(v)7.1 and K(v)7.1/KCNE1 currents and these effects were concentration dependent. The IC(50) value for inhibition of K(v)7.1 channels was approximately 4 µM and the EC(50) values for activation of K(v)7.2-7.4, K(v)7.2/K(v)7.3 and K(v)7.3/K(v)7.5 channels were approximately 2-5 µM. The effects of celecoxib were manifested by increasing current amplitudes, shifting the voltage-dependent activation curve in a more negative direction and slowing the deactivation of K(v)7 currents. 2,5-Dimethyl-celecoxib, a celecoxib analogue devoid of COX inhibition activity, has similar but greater effects on K(v)7currents. K(v)7.2(A235T) and K(v) 7.2(W236L) mutant channels, which have greatly attenuated responses to retigabine, showed a reversed response to celecoxib, from activation to inhibition. CONCLUSIONS AND IMPLICATIONS: These results suggest that K(v)7 channels are targets of celecoxib action and provide new mechanistic evidence for understanding the effects of celecoxib. They also provide a new approach to developing K(v)7 modulators and for studying the structure-function relationship of K(v)7 channels.

[Immunomodulatory activity and anti-tumor activity of Oldenlandia diffusa in vitro].

OBJECTIVE: To study the effect of Oldenlandia diffusa (OD) on lymphocytes and tumor cells in vitro. METHODS: Effects of OD extract (ODE) on proliferation of spleen cells of mice and the phagocytosis of monocytes to tumor cells using 3H incorporation were analysed, and effect on specific activity of natural killer (NK) cells to human and mice's tumor cells was determined using free 51 Cr experiments, effects on B cells' antibody production and cytokine of monocyte production were investigated by ELISA and biological method, the chemical composition of OD was analyzed by chromatography, protease digestion and sugar decomposition (NaIO4) test. RESULTS: ODE could vigorously promote the proliferative activity of spleen cells in mice, specific lethal activity of human and mice's NK cell to tumor cells, B cells' antibody production, monocytes' cytokine production and its phagocytosis to remove the tumor cells. By means of chromatography, protease E digestion and NaIO4 treatment, the result of chemical composition analysis showed that the component of OD was 90 kDa glycoprotein. CONCLUSION: ODE has immuno-modulating activity and anti-tumor activity in vitro through stimulating the immune system to kill or engulf tumor cells, which could be used clinically for immune function modulation and to treat tumor and other diseases.

Effect of activated charcoal on endotoxin adsorption. Part I. An in vitro study.

An in vitro study by using cross-linked agarose coated activated charcoal (CAAC-II) as adsorbent was conducted. Endotoxin was quantitatively measured by chromogenic method of limulus lysate test. Results of the study demonstrated that endotoxin was fairly efficiently removed by CAAC-II. It may therefore be possible for effective adsorption of endotoxin in vivo studies and warrant further experimental study for CAAC-II hemoperfusion in clinical endotoxemia.