Characterization of Perturbing Actions by Verteporfin, a Benzoporphyrin Photosensitizer, on Membrane Ionic Currents

Abstract

Verteporfin (VP), a benzoporphyrin derivative, has been clinically tailored as a photosensitizer and recently known to suppress YAP-TEAD complex accompanied by suppression of the growth in an array of neoplastic cells. However, the detailed information is little available regarding possible modifications of it and its related compounds on transmembrane ionic currents, despite its growing use in clinical settings. In this study, from whole cell recordings, VP (0.3–100 μM) increased the amplitude of Ca 2+ -activated K + currents ( I K(Ca) ) in pituitary tumor (GH 3 ) cells in a concentration-dependent manner with an EC 50 value of 2.4 μM. VP-stimulated I K(Ca) in these cells was suppressed by further addition of either paxilline, iberiotoxin, or dithiothreitol, but not by that of tobultamide or TRAM-39. VP at a concentration of 10 μM mildly suppressed the amplitude of delayed-rectifier K + current; however, it had minimal effects on M-type K + current. In cell-attached current recordings, addition of VP to the recording medium enhanced the activity of large-conductance Ca 2+ -activated K + (BK Ca ) channels. In the presence of VP, additional illumination with light intensity of 5.5 mW/cm 2 raised the probability of BK Ca -channel openings further. Addition of VP decreased the peak amplitude of L-type Ca 2+ current together with slowed inactivation time course of the current; however, it failed to modify voltage-gated Na + current. Illumination of GH 3 cells in continued presence of VP also induced a non-selective cation current. Additionally, VP increased the activity of BK Ca channels in human 13-06-MG glioma cells with an EC 50 value of 1.9 μM. Therefore, the effects of VP on ionic currents described herein tend to be upstream of its inhibition of YAP-TEAD complex and they are conceivably likely to contribute to the underlying mechanisms through which it and its structurally similar compounds effect the modifications in functional activities of pituitary or glial neoplastic cells, if the in vivo findings occur.

High Efficacy by GAL-021: A Known Intravenous Peripheral Chemoreceptor Modulator that Suppresses BKCa-Channel Activity and Inhibits IK(M) or Ih

“…GAL-021-mediated suppression of I K(Ca) observed in GH 3 cells was found to be attenuated by further application of either verteporfin or ionomycin, yet not by diazoxide, an activator of K ATP channels [51]. Vertiporfin was recently reported to activate BK Ca channels [35], while ionomycin is a Ca 2+ ionophore. In keeping with the present observations and previous studies [5,6], the inside-out current recordings revealed the effectiveness of GAL-021 in decreasing the open-state probability of BK Ca channels that would be open.…”

“…After cells were left to adhere to the bottom for several minutes, the recordings were performed. Patch-clamp experiments under either whole-cell, cell-attached, or inside-out mode were achieved with either an RK-400 (Biologic, Claix, France) or an Axopatch-200B amplifier (Molecular Devices, Sunnyvale, CA) [16,35]. We fabricated patch pipettes, the resistance of which was around 3 to 5 MΩ, from Kimax-51 borosilicate capillaries (#34500; Kimble; Dogger, New Taipei City, Taiwan) pulled on either a PP-830 vertical puller (Narishige, Major Instruments, New Taipei City, Taiwan) or a P-97 programmable horizontal puller (Sutter Instruments, Novato, CA, USA), and the pipettes were then fire polished with an MF-83 microforge (Narishige).…”

“…As whole-cell model, that is, when membrane patch was broken by suction, was achieved, we maintained the examined cell at the level of 0 mV, a level which was delivered to inactivate other types of outwardly rectifying K + currents [36], and ionic currents were then evoked by a series of voltage pulses ranging between 0 and +80 mV. This type of macroscopic outward currents with a markedly noisy and outwardly rectifying property has been regarded as I K(Ca) which is particularly subject to inhibition by tremorgenic mycotoxins (e.g., paxilline or verroculogen) [16,35,37]. As illustrated in Figure 1A,B, the presence of GAL-021 (1 or 3 µM) produced a progressive decrease in I K(Ca) amplitude measured at the entire voltage-clamp steps.…”

“…We further tested whether the suppressive effect of GAL-021 on I K(Ca) was perturbed by diazoxide, a stimulator of ATP-sensitive K + (K ATP ) channels [38]; verteporfin, an activator of I K(Ca) [35]; or ionomycin, a Ca 2+ ionophore [39]. As shown in Figure 2, when the cells were depolarized from 0 to +50 mV, exposure to GAL-021 (3 µM) significantly suppressed the amplitude of I K(Ca) .…”

Inhibitory Effective Perturbations of Cilobradine (DK-AH269), A Blocker of HCN Channels, on the Amplitude and Gating of Both Hyperpolarization-Activated Cation and Delayed-Rectifier Potassium Currents

“…The tip resistance was in the range of 3-5 MΩ when the electrode was backfilled up with different internal solutions detailed above. Ion currents were measured in the whole-cell mode of standard patch-clamp technique by using either an Axopatch 200B (Molecular Devices, Sunnyvale, CA, USA) or an RK-400 (Bio-Logic, Claix, France) patch amplifier [15,27,47]. The recorded area on the vibration-free table was shielded by using a Faraday cage (Scitech, Seoul, South Korea) to minimize mechanical noise.…”