Limits...
The Cyclooctadepsipeptide Anthelmintic Emodepside Differentially Modulates Nematode, Insect and Human Calcium-Activated Potassium (SLO) Channel Alpha Subunits.

Crisford A, Ebbinghaus-Kintscher U, Schoenhense E, Harder A, Raming K, O'Kelly I, Ndukwe K, O'Connor V, Walker RJ, Holden-Dye L - PLoS Negl Trop Dis (2015)

Bottom Line: To test this at the molecular level we compared the actions of emodepside at heterologously expressed SLO-1 alpha subunit orthologues from nematode (Caenorhabditis elegans), Drosophila melanogaster and human using whole cell voltage clamp.Intriguingly we found that emodepside modulated nematode (Ce slo-1), insect (Drosophila, Dm slo) and human (hum kcnma1)SLO channels but that there are discrete differences in the features of the modulation that are consistent with its anthelmintic efficacy.These data provide an important benchmark for the wider deployment of emodepside as an anthelmintic treatment.

View Article: PubMed Central - PubMed

Affiliation: Centre for Biological Sciences, University of Southampton, Southampton, United Kingdom.

ABSTRACT
The anthelmintic emodepside paralyses adult filarial worms, via a mode of action distinct from previous anthelmintics and has recently garnered interest as a new treatment for onchocerciasis. Whole organism data suggest its anthelmintic action is underpinned by a selective activation of the nematode isoform of an evolutionary conserved Ca2+-activated K+ channel, SLO-1. To test this at the molecular level we compared the actions of emodepside at heterologously expressed SLO-1 alpha subunit orthologues from nematode (Caenorhabditis elegans), Drosophila melanogaster and human using whole cell voltage clamp. Intriguingly we found that emodepside modulated nematode (Ce slo-1), insect (Drosophila, Dm slo) and human (hum kcnma1)SLO channels but that there are discrete differences in the features of the modulation that are consistent with its anthelmintic efficacy. Nematode SLO-1 currents required 100 μM intracellular Ca2+ and were strongly facilitated by emodepside (100 nM; +73.0 ± 17.4%; n = 9; p < 0.001). Drosophila Slo currents on the other hand were activated by emodepside (10 μM) in the presence of 52 nM Ca2+ but were inhibited in the presence of 290 nM Ca2+ and exhibited a characteristic loss of rectification. Human Slo required 300 nM Ca2+ and emodepside transiently facilitated currents (100 nM; +33.5 ± 9%; n = 8; p<0.05) followed by a sustained inhibition (-52.6 ± 9.8%; n = 8; p < 0.001). This first cross phyla comparison of the actions of emodepside at nematode, insect and human channels provides new mechanistic insight into the compound's complex modulation of SLO channels. Consistent with whole organism behavioural studies on C. elegans, it indicates its anthelmintic action derives from a strong activation of SLO current, not observed in the human channel. These data provide an important benchmark for the wider deployment of emodepside as an anthelmintic treatment.

No MeSH data available.


Related in: MedlinePlus

The effect of emodepside on voltage-activation of Ce SLO-1 and hum KCNMA1.Analysis of the current-voltage relationship of the data sets shown in Fig 6 and Fig 7 for Ce SLO-1 and hum KCNMA1 whole cell currents expressed in HEK293 cells before and after treatment with emodepside (100nM). Ce SLO-1 currents were recorded before and 12 min after emodepside application. Hum KCNMA1 currents were recorded before and 5 min after emodepside application. Membrane potential was held at -60mV and stepped to between -100 and +90mV in 10mV increments for 50ms. Data points are the mean ± s.e.mean of n = 9 for Ce SLO-1 and n = 8 for hum KCNMA1. The amplitude of Ce SLO-1 currents in the presence of 100nM emodepside was significantly greater than currents recorded in the absence of emodepside between +20 and +90mV and for hum KCNMA1 was significantly greater than currents recorded in the absence of emodepside between +70 and +90mV; p<0.05; two-way ANOVA with Bonferroni post-hoc tests.
© Copyright Policy
Related In: Results  -  Collection

License
getmorefigures.php?uid=PMC4593646&req=5

pntd.0004062.g007: The effect of emodepside on voltage-activation of Ce SLO-1 and hum KCNMA1.Analysis of the current-voltage relationship of the data sets shown in Fig 6 and Fig 7 for Ce SLO-1 and hum KCNMA1 whole cell currents expressed in HEK293 cells before and after treatment with emodepside (100nM). Ce SLO-1 currents were recorded before and 12 min after emodepside application. Hum KCNMA1 currents were recorded before and 5 min after emodepside application. Membrane potential was held at -60mV and stepped to between -100 and +90mV in 10mV increments for 50ms. Data points are the mean ± s.e.mean of n = 9 for Ce SLO-1 and n = 8 for hum KCNMA1. The amplitude of Ce SLO-1 currents in the presence of 100nM emodepside was significantly greater than currents recorded in the absence of emodepside between +20 and +90mV and for hum KCNMA1 was significantly greater than currents recorded in the absence of emodepside between +70 and +90mV; p<0.05; two-way ANOVA with Bonferroni post-hoc tests.

Mentions: The voltage activation of Ce SLO-1 and hum KCNMA1 was tested before and during the peak of the emodepside facilitation (i.e. after 12 min application of 100 nM emodepside for Ce SLO-1 and after 5 min application of 100nM emodepside for hum KCNMA1; Fig 7). A significant shift in the voltage activation curve was observed for Ce SLO-1 (p<0.05) after emodepside treatment and a smaller significant shift for hum KCNMA1 (p<0.05). The data from Fig 7 were subjected to Boltzmann analysis to estimate V50, the membrane potential for half-activation of the channels using Gmax values of 0.0174, 0.0258, 0.0177 and 0.0253 μS for Ce SLO-1 control, Ce SLO-1 with emodepside, hum KCNMA1 control and hum KCNMA1 with emodepside, respectively. For Ce SLO-1 V50 was +49.74 mV (95% confidence intervals 45.46 to 54.01; V slope 20.66) and this was shifted in the presence of emodepside to +32.95 mV (95% confidence intervals 28.82 to 37.07; Vslope 28.36). For hum KCNMA1 the V50 was +56.51 mV (95% confidence intervals 52.82 to 60.20; Vslope 17.45) which was unchanged in the presence of emodepside, + 57.88 mV (95% confidence intervals 54.09 to 61.68; Vslope 17.12). This analysis confirms the selective effect of emodepside on the voltage-activation for Ce SLO-1.


The Cyclooctadepsipeptide Anthelmintic Emodepside Differentially Modulates Nematode, Insect and Human Calcium-Activated Potassium (SLO) Channel Alpha Subunits.

Crisford A, Ebbinghaus-Kintscher U, Schoenhense E, Harder A, Raming K, O'Kelly I, Ndukwe K, O'Connor V, Walker RJ, Holden-Dye L - PLoS Negl Trop Dis (2015)

The effect of emodepside on voltage-activation of Ce SLO-1 and hum KCNMA1.Analysis of the current-voltage relationship of the data sets shown in Fig 6 and Fig 7 for Ce SLO-1 and hum KCNMA1 whole cell currents expressed in HEK293 cells before and after treatment with emodepside (100nM). Ce SLO-1 currents were recorded before and 12 min after emodepside application. Hum KCNMA1 currents were recorded before and 5 min after emodepside application. Membrane potential was held at -60mV and stepped to between -100 and +90mV in 10mV increments for 50ms. Data points are the mean ± s.e.mean of n = 9 for Ce SLO-1 and n = 8 for hum KCNMA1. The amplitude of Ce SLO-1 currents in the presence of 100nM emodepside was significantly greater than currents recorded in the absence of emodepside between +20 and +90mV and for hum KCNMA1 was significantly greater than currents recorded in the absence of emodepside between +70 and +90mV; p<0.05; two-way ANOVA with Bonferroni post-hoc tests.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4593646&req=5

pntd.0004062.g007: The effect of emodepside on voltage-activation of Ce SLO-1 and hum KCNMA1.Analysis of the current-voltage relationship of the data sets shown in Fig 6 and Fig 7 for Ce SLO-1 and hum KCNMA1 whole cell currents expressed in HEK293 cells before and after treatment with emodepside (100nM). Ce SLO-1 currents were recorded before and 12 min after emodepside application. Hum KCNMA1 currents were recorded before and 5 min after emodepside application. Membrane potential was held at -60mV and stepped to between -100 and +90mV in 10mV increments for 50ms. Data points are the mean ± s.e.mean of n = 9 for Ce SLO-1 and n = 8 for hum KCNMA1. The amplitude of Ce SLO-1 currents in the presence of 100nM emodepside was significantly greater than currents recorded in the absence of emodepside between +20 and +90mV and for hum KCNMA1 was significantly greater than currents recorded in the absence of emodepside between +70 and +90mV; p<0.05; two-way ANOVA with Bonferroni post-hoc tests.
Mentions: The voltage activation of Ce SLO-1 and hum KCNMA1 was tested before and during the peak of the emodepside facilitation (i.e. after 12 min application of 100 nM emodepside for Ce SLO-1 and after 5 min application of 100nM emodepside for hum KCNMA1; Fig 7). A significant shift in the voltage activation curve was observed for Ce SLO-1 (p<0.05) after emodepside treatment and a smaller significant shift for hum KCNMA1 (p<0.05). The data from Fig 7 were subjected to Boltzmann analysis to estimate V50, the membrane potential for half-activation of the channels using Gmax values of 0.0174, 0.0258, 0.0177 and 0.0253 μS for Ce SLO-1 control, Ce SLO-1 with emodepside, hum KCNMA1 control and hum KCNMA1 with emodepside, respectively. For Ce SLO-1 V50 was +49.74 mV (95% confidence intervals 45.46 to 54.01; V slope 20.66) and this was shifted in the presence of emodepside to +32.95 mV (95% confidence intervals 28.82 to 37.07; Vslope 28.36). For hum KCNMA1 the V50 was +56.51 mV (95% confidence intervals 52.82 to 60.20; Vslope 17.45) which was unchanged in the presence of emodepside, + 57.88 mV (95% confidence intervals 54.09 to 61.68; Vslope 17.12). This analysis confirms the selective effect of emodepside on the voltage-activation for Ce SLO-1.

Bottom Line: To test this at the molecular level we compared the actions of emodepside at heterologously expressed SLO-1 alpha subunit orthologues from nematode (Caenorhabditis elegans), Drosophila melanogaster and human using whole cell voltage clamp.Intriguingly we found that emodepside modulated nematode (Ce slo-1), insect (Drosophila, Dm slo) and human (hum kcnma1)SLO channels but that there are discrete differences in the features of the modulation that are consistent with its anthelmintic efficacy.These data provide an important benchmark for the wider deployment of emodepside as an anthelmintic treatment.

View Article: PubMed Central - PubMed

Affiliation: Centre for Biological Sciences, University of Southampton, Southampton, United Kingdom.

ABSTRACT
The anthelmintic emodepside paralyses adult filarial worms, via a mode of action distinct from previous anthelmintics and has recently garnered interest as a new treatment for onchocerciasis. Whole organism data suggest its anthelmintic action is underpinned by a selective activation of the nematode isoform of an evolutionary conserved Ca2+-activated K+ channel, SLO-1. To test this at the molecular level we compared the actions of emodepside at heterologously expressed SLO-1 alpha subunit orthologues from nematode (Caenorhabditis elegans), Drosophila melanogaster and human using whole cell voltage clamp. Intriguingly we found that emodepside modulated nematode (Ce slo-1), insect (Drosophila, Dm slo) and human (hum kcnma1)SLO channels but that there are discrete differences in the features of the modulation that are consistent with its anthelmintic efficacy. Nematode SLO-1 currents required 100 μM intracellular Ca2+ and were strongly facilitated by emodepside (100 nM; +73.0 ± 17.4%; n = 9; p < 0.001). Drosophila Slo currents on the other hand were activated by emodepside (10 μM) in the presence of 52 nM Ca2+ but were inhibited in the presence of 290 nM Ca2+ and exhibited a characteristic loss of rectification. Human Slo required 300 nM Ca2+ and emodepside transiently facilitated currents (100 nM; +33.5 ± 9%; n = 8; p<0.05) followed by a sustained inhibition (-52.6 ± 9.8%; n = 8; p < 0.001). This first cross phyla comparison of the actions of emodepside at nematode, insect and human channels provides new mechanistic insight into the compound's complex modulation of SLO channels. Consistent with whole organism behavioural studies on C. elegans, it indicates its anthelmintic action derives from a strong activation of SLO current, not observed in the human channel. These data provide an important benchmark for the wider deployment of emodepside as an anthelmintic treatment.

No MeSH data available.


Related in: MedlinePlus