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Anesthetic drug midazolam inhibits cardiac human ether-à-go-go-related gene channels: mode of action.

Vonderlin N, Fischer F, Zitron E, Seyler C, Scherer D, Thomas D, Katus HA, Scholz EP - Drug Des Devel Ther (2015)

Bottom Line: However, steady-state inactivation was not significantly affected.Analyzing the subacute effects of midazolam on hERG channel trafficking, we further found that midazolam does not affect channel surface expression.Taken together, we show that the anesthetic midazolam is a low-affinity inhibitor of cardiac hERG channels without additional effects on channel surface expression.

View Article: PubMed Central - PubMed

Affiliation: Department of Internal Medicine III, University Hospital Heidelberg, Heidelberg, Germany.

ABSTRACT
Midazolam is a short-acting benzodiazepine that is in wide clinical use as an anxiolytic, sedative, hypnotic, and anticonvulsant. Midazolam has been shown to inhibit ion channels, including calcium and potassium channels. So far, the effects of midazolam on cardiac human ether-à-go-go-related gene (hERG) channels have not been analyzed. The inhibitory effects of midazolam on heterologously expressed hERG channels were analyzed in Xenopus oocytes using the double-electrode voltage clamp technique. We found that midazolam inhibits hERG channels in a concentration-dependent manner, yielding an IC50 of 170 μM in Xenopus oocytes. When analyzed in a HEK 293 cell line using the patch-clamp technique, the IC50 was 13.6 μM. Midazolam resulted in a small negative shift of the activation curve of hERG channels. However, steady-state inactivation was not significantly affected. We further show that inhibition is state-dependent, occurring within the open and inactivated but not in the closed state. There was no frequency dependence of block. Using the hERG pore mutants F656A and Y652A we provide evidence that midazolam uses a classical binding site within the channel pore. Analyzing the subacute effects of midazolam on hERG channel trafficking, we further found that midazolam does not affect channel surface expression. Taken together, we show that the anesthetic midazolam is a low-affinity inhibitor of cardiac hERG channels without additional effects on channel surface expression. These data add to the current understanding of the pharmacological profile of the anesthetic midazolam.

No MeSH data available.


Related in: MedlinePlus

Effects of midazolam on channel inactivation.Notes: (A) In order to analyze the effects of midazolam on steady-state inactivation, a double-step voltage protocol was used (see inset). A typical family of current traces is displayed for control conditions (A) and after application of 200 μM midazolam (B). Current traces were corrected for channel deactivation by extrapolation to the beginning of the second voltage step (see dashed lines and arrows in A). (C) Current-voltage relationship of tail current amplitude. Steady-state inactivation was obtained by dividing the current amplitude by the electrochemical driving force (D). Midazolam did not significantly affect the half-maximal inactivation voltage of hERG channels. Protocol: holding potential −80 mV, first test pulse to +40 mV (one second), second test pulse between −140 to +40 mV (20 mV increment, 500 msec).Abbreviation: hERG, human ether-à-go-go-related gene.
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f3-dddt-9-867: Effects of midazolam on channel inactivation.Notes: (A) In order to analyze the effects of midazolam on steady-state inactivation, a double-step voltage protocol was used (see inset). A typical family of current traces is displayed for control conditions (A) and after application of 200 μM midazolam (B). Current traces were corrected for channel deactivation by extrapolation to the beginning of the second voltage step (see dashed lines and arrows in A). (C) Current-voltage relationship of tail current amplitude. Steady-state inactivation was obtained by dividing the current amplitude by the electrochemical driving force (D). Midazolam did not significantly affect the half-maximal inactivation voltage of hERG channels. Protocol: holding potential −80 mV, first test pulse to +40 mV (one second), second test pulse between −140 to +40 mV (20 mV increment, 500 msec).Abbreviation: hERG, human ether-à-go-go-related gene.

Mentions: The effects of midazolam on channel inactivation were analyzed using a double-step voltage protocol as reported previously.21,22 A first depolarizing step to +40 mV served to fully activate and inactivate the channels. This step was followed by a variable return pulse to potentials between −140 mV and +80 mV (500 msec duration, 20 mV increment; Figure 3A, inset). During this second voltage step, channels quickly redistributed to their activated and inactivated states (Figure 3A and B). In order to correct for channel deactivation, single exponentials or double exponentials were fitted to the deactivating current traces and extrapolated to the beginning of the second voltage step (see dashed lines and arrows in Figure 3A). Figure 3C displays the current-voltage relationship of corrected tail current amplitudes under control conditions and after incubation with 200 μM midazolam. When dividing the current amplitude by the corresponding electrochemical driving force, steady-state inactivation curves could be established.22 Inactivation curves for control conditions and after midazolam incubation are displayed in Figure 3D. When fitted with a Boltzmann function, the half-maximal inactivation voltage could be obtained, yielding a V1/2 of −29.2±3.8 mV for control conditions and −31.3±13.4 mV for midazolam incubation. There was no significant difference between these values (n=6, P<0.05).


Anesthetic drug midazolam inhibits cardiac human ether-à-go-go-related gene channels: mode of action.

Vonderlin N, Fischer F, Zitron E, Seyler C, Scherer D, Thomas D, Katus HA, Scholz EP - Drug Des Devel Ther (2015)

Effects of midazolam on channel inactivation.Notes: (A) In order to analyze the effects of midazolam on steady-state inactivation, a double-step voltage protocol was used (see inset). A typical family of current traces is displayed for control conditions (A) and after application of 200 μM midazolam (B). Current traces were corrected for channel deactivation by extrapolation to the beginning of the second voltage step (see dashed lines and arrows in A). (C) Current-voltage relationship of tail current amplitude. Steady-state inactivation was obtained by dividing the current amplitude by the electrochemical driving force (D). Midazolam did not significantly affect the half-maximal inactivation voltage of hERG channels. Protocol: holding potential −80 mV, first test pulse to +40 mV (one second), second test pulse between −140 to +40 mV (20 mV increment, 500 msec).Abbreviation: hERG, human ether-à-go-go-related gene.
© Copyright Policy
Related In: Results  -  Collection

License
Show All Figures
getmorefigures.php?uid=PMC4338257&req=5

f3-dddt-9-867: Effects of midazolam on channel inactivation.Notes: (A) In order to analyze the effects of midazolam on steady-state inactivation, a double-step voltage protocol was used (see inset). A typical family of current traces is displayed for control conditions (A) and after application of 200 μM midazolam (B). Current traces were corrected for channel deactivation by extrapolation to the beginning of the second voltage step (see dashed lines and arrows in A). (C) Current-voltage relationship of tail current amplitude. Steady-state inactivation was obtained by dividing the current amplitude by the electrochemical driving force (D). Midazolam did not significantly affect the half-maximal inactivation voltage of hERG channels. Protocol: holding potential −80 mV, first test pulse to +40 mV (one second), second test pulse between −140 to +40 mV (20 mV increment, 500 msec).Abbreviation: hERG, human ether-à-go-go-related gene.
Mentions: The effects of midazolam on channel inactivation were analyzed using a double-step voltage protocol as reported previously.21,22 A first depolarizing step to +40 mV served to fully activate and inactivate the channels. This step was followed by a variable return pulse to potentials between −140 mV and +80 mV (500 msec duration, 20 mV increment; Figure 3A, inset). During this second voltage step, channels quickly redistributed to their activated and inactivated states (Figure 3A and B). In order to correct for channel deactivation, single exponentials or double exponentials were fitted to the deactivating current traces and extrapolated to the beginning of the second voltage step (see dashed lines and arrows in Figure 3A). Figure 3C displays the current-voltage relationship of corrected tail current amplitudes under control conditions and after incubation with 200 μM midazolam. When dividing the current amplitude by the corresponding electrochemical driving force, steady-state inactivation curves could be established.22 Inactivation curves for control conditions and after midazolam incubation are displayed in Figure 3D. When fitted with a Boltzmann function, the half-maximal inactivation voltage could be obtained, yielding a V1/2 of −29.2±3.8 mV for control conditions and −31.3±13.4 mV for midazolam incubation. There was no significant difference between these values (n=6, P<0.05).

Bottom Line: However, steady-state inactivation was not significantly affected.Analyzing the subacute effects of midazolam on hERG channel trafficking, we further found that midazolam does not affect channel surface expression.Taken together, we show that the anesthetic midazolam is a low-affinity inhibitor of cardiac hERG channels without additional effects on channel surface expression.

View Article: PubMed Central - PubMed

Affiliation: Department of Internal Medicine III, University Hospital Heidelberg, Heidelberg, Germany.

ABSTRACT
Midazolam is a short-acting benzodiazepine that is in wide clinical use as an anxiolytic, sedative, hypnotic, and anticonvulsant. Midazolam has been shown to inhibit ion channels, including calcium and potassium channels. So far, the effects of midazolam on cardiac human ether-à-go-go-related gene (hERG) channels have not been analyzed. The inhibitory effects of midazolam on heterologously expressed hERG channels were analyzed in Xenopus oocytes using the double-electrode voltage clamp technique. We found that midazolam inhibits hERG channels in a concentration-dependent manner, yielding an IC50 of 170 μM in Xenopus oocytes. When analyzed in a HEK 293 cell line using the patch-clamp technique, the IC50 was 13.6 μM. Midazolam resulted in a small negative shift of the activation curve of hERG channels. However, steady-state inactivation was not significantly affected. We further show that inhibition is state-dependent, occurring within the open and inactivated but not in the closed state. There was no frequency dependence of block. Using the hERG pore mutants F656A and Y652A we provide evidence that midazolam uses a classical binding site within the channel pore. Analyzing the subacute effects of midazolam on hERG channel trafficking, we further found that midazolam does not affect channel surface expression. Taken together, we show that the anesthetic midazolam is a low-affinity inhibitor of cardiac hERG channels without additional effects on channel surface expression. These data add to the current understanding of the pharmacological profile of the anesthetic midazolam.

No MeSH data available.


Related in: MedlinePlus