The Evidence for Sparsentan-Mediated Inhibition of INa and IK(erg): Possibly Unlinked to Its Antagonism of Angiotensin II or Endothelin Type a Receptor

Abstract

Sparsentan is viewed as a dual antagonist of endothelin type A (ETA) receptor and angiotensin II (AngII) receptor and it could be beneficial in patients with focal segmental glomerulosclerosis. Moreover, it could improve glomerular filtration rate and augment protective tissue remodeling in mouse models of focal segmental glomerulosclerosis. The ionic mechanisms through which it interacts with the magnitude and/or gating kinetics of ionic currents in excitable cells were not thoroughly investigated. Herein, we aimed to examine the effects of varying sparsentan concentrations on ionic currents residing in pituitary GH3 somatolactotrophs. From whole-cell current recordings made in GH3 cells, sparsentan (0.3–100 μM) differentially inhibited the peak and late components of voltage-gated Na+ current (INa). The IC50 value of sparsentan required to exert a reduction in peak and late INa in GH3 cells was 15.04 and 1.21 μM, respectively; meanwhile, the KD value estimated from its shortening in the slow component of INa inactivation time constant was 2.09 μM. The sparsentan (10 μM) presence did not change the overall current–voltage relationship of INa; however, the steady-state inactivation curve of the current was shifted to more negative potential in its presence (10 μM), with no change in the gating charge of the curve. The window INa activated by a brief upsloping ramp was decreased during exposure to sparsentan (10 μM); moreover, recovery of peak INa became slowed in its presence. The Tefluthrin (Tef)-stimulated resurgent INa activated in response to abrupt depolarization followed by the descending ramp pulse was additionally attenuated by subsequent application of sparsentan. In continued presence of Tef (3 μM) or β-pompilidotoxin (3 μM), further application of sparsentan (3 μM) reversed their stimulation of INa. However, sparsentan-induced inhibition of INa failed to be overcome by subsequent application of either endothelin 1 (1 μM) or angiotensin II (1 μM); moreover, in continued presence of endothelin (1 μM) or angiotensin II (1 μM), further addition of sparsentan (3 μM) effectively decreased peak INa. Additionally, the application of sparsentan (3 μM) inhibited the peak and late components of erg-mediated K+ current in GH3 cells, although it mildly decreased the amplitude of delayed-rectifier K+ current. Altogether, this study provides a distinct yet unidentified finding that sparsentan may perturb the amplitude or gating of varying ionic currents in excitable cells.

The Evidence for Effective Inhibition of INa Produced by Mirogabalin ((1R,5S,6S)-6-(aminomethyl)-3-ethyl-bicyclo [3.2.0] hept-3-ene-6-acetic acid), a Known Blocker of CaV Channels

“…The presence of instantaneous I Na(W) evoked by the ascending (or upsloping) ramp voltage (V ramp ) was revealed earlier in a variety of excitable cells [ 37 , 38 , 39 , 40 , 41 ]. Next, we explored whether the MGB presence in GH 3 cells could modify the magnitude of I Na(W) activated in response to the rapid ascending V ramp .…”

“…Next, we explored whether the MGB presence in GH 3 cells could modify the magnitude of I Na(W) activated in response to the rapid ascending V ramp . In order to conduct these experiments, the tested cell was voltage-clamped at −80 mV, and we then applied an ascending V ramp from −110 to +50 mV for a duration of 50 ms to evoke I Na(W ) [ 37 ]. As disclosed in Figure 5 A,B, within one minute of exposing cells to MGB (10 or 30 μM), the amplitude of I Na(W) achieved by the 50-ms upsloping V ramp decreased strikingly.…”

“…As the electrodes were filled with the different internal solutions described above, their resistance was measured to range between 3 and 5 MΩ, for the purpose of avoiding excessive damage to the cell. Patch-clamp recordings were carried out in whole-cell configuration using either an RK-400 (Bio-Logic, Claix, France) or an Axopatch-200B amplifier (Molecular Devices; Bestgen Biotech, New Taipei City, Taiwan), as described elsewhere [ 31 , 37 , 52 , 67 ]. Whole-cell recording was achieved by rupturing the patch of membrane isolated with GΩ sealing by the patch pipet, which brings the cell interior into contact with the pipet interior.…”

“…The Na V 1.7 and Na V 1.8 subtypes have emerged as key molecules involved in peripheral pain processing and in the development of an increased pain sensitivity associated with inflammation and tissue injury [ 26 , 27 , 28 , 29 , 30 ]. Several activators or inhibitors have been increasingly reported to preferentially modify the late component of the voltage-gated Na + current (i.e., I Na(L) ) [ 24 , 31 , 32 , 33 , 34 , 35 , 36 , 37 ]. However, to date, the issue of whether or how MGB could perturb the magnitude, kinetic gating, or hysteresis of membrane ionic currents (e.g., I Na ) is poorly characterized.…”

Characterization in Inhibitory Effectiveness of Carbamazepine in Voltage-Gated Na+ and Erg-Mediated K+ Currents in a Mouse Neural Crest-Derived (Neuro-2a) Cell Line

“…The activity of these channels is to depolarize the cell and to generate the upstroke of the action potential, thereby controlling the firing amplitude, frequency, and pattern inherently in electrically excitable cells [ 36 , 37 , 38 ]. Of additional note, some inhibitors of Na V channels (e.g., esaxerenone, ranolazine, and sparsentan) were recognized to increase the inactivation rate of voltage-gated Na + current ( I Na ) [ 37 , 38 , 39 , 40 , 41 , 42 ], whereas several activators of Na V channels (e.g., tefluthrin [Tef]) could preferentially slow the inactivation rate as well as increase the late component of I Na ( I Na(L) ) [ 43 , 44 , 45 ].…”