Stilbene derivatives inhibit the activity of the inner mitochondrial membrane chloride channels

Abstract

Ion channels selective for chloride ions are present in all biological membranes, where they regulate the cell volume or membrane potential. Various chloride channels from mitochondrial membranes have been described in recent years. The aim of our study was to characterize the effect of stilbene derivatives on single-chloride channel activity in the inner mitochondrial membrane. The measurements were performed after the reconstitution into a planar lipid bilayer of the inner mitochondrial membranes from rat skeletal muscle (SMM), rat brain (BM) and heart (HM) mitochondria. After incorporation in a symmetric 450/450 mM KCl solution (cis/trans), the chloride channels were recorded with a mean conductance of 155 ± 5 pS (rat skeletal muscle) and 120 ± 16 pS (rat brain). The conductances of the chloride channels from the rat heart mitochondria in 250/50 mM KCl (cis/trans) gradient solutions were within the 70-130 pS range. The chloride channels were inhibited by these two stilbene derivatives: 4,4'-diisothiocyanostilbene-2,2'-disulfonic acid (DIDS) and 4-acetamido-4'-isothiocyanostilbene-2,2'-disulfonic acid (SITS). The skeletal muscle mitochondrial chloride channel was blocked after the addition of 1 mM DIDS or SITS, whereas the brain mitochondrial channel was blocked by 300 µM DIDS or SITS. The chloride channel from the rat heart mitochondria was inhibited by 50-100 μM DIDS. The inhibitory effect of DIDS was irreversible. Our results confirm the presence of chloride channels sensitive to stilbene derivatives in the inner mitochondrial membrane from rat skeletal muscle, brain and heart cells.

Mitochondrial chloride channels – What are they for?

“…This was supported by observations that chloride channel blockers inhibited apoptosis, including ROS‐dependent apoptosis [6,102]. The chloride channel blockers DIDS, NPPB and phloretin inhibited both H 2 O 2 ‐induced apoptosis of cardiomyocytes and the activities of intracellular chloride channels derived from mitochondrial and lysosomal membranes [26,48,49]. Further evidence of a role for mtCl channels in apoptosis include a suggestion of a pro‐apoptotic role of the mtCLIC in p53‐mediated apoptosis [73,75] and the formation of a Cl − selective channel (Cl − /K + = ∼3) by pro‐apoptotic Bax protein in BLM.…”

“…Conductances of chloride channels in BLM (symmetrical 450 mmol/l KCl) from rat skeletal muscle and brain were 155 ± 5 pS and 120 ± 16 pS, respectively [48]. The channels were blocked by the chloride channel inhibitors SITS and DIDS [48]. A bell‐shaped dependence of P open on voltage (−60 mV to +100 mV) was observed for chloride channels under asymmetrical (250/50 mmol/l KCl) and symmetrical conditions (150 mmol/l KCl, 92 ± 14 pS).…”

“…A two-substate anion channel ($40 pS or $85 pS in symmetric 300 or 550 mmol/l choline chloride, respectively) displayed Ca 2+ and Mg 2+ sensitive rectification [47]. Conductances of chloride channels in BLM (symmetrical 450 mmol/l KCl) from rat skeletal muscle and brain were 155 ± 5 pS and 120 ± 16 pS, respectively [48]. The channels were blocked by the chloride channel inhibitors SITS and DIDS [48].…”

Identifying an uptake mechanism for the antiepileptic and bipolar disorder treatment valproic acid using the simple biomedical modelDictyostelium

“…Note that these bicarbonate inhibitors are not specific to SLC4 transporters and potentially block a range of anion channels and exchangers (Izumi et al, 2003;Utoguchi and Audus, 2000); however, their functional overlap allows identification of the most likely target reducing the uptake of VPA. DIDS, SITS and tenidap are known to inhibit chloride channels, as well as SLC4 (Koszela-Piotrowska et al, 2007;McNiff et al, 1994). However, chloride channels and monocarboxylate transporters of the MCT family (Goncalves et al, 2009) are also blocked by NPPB, but this inhibitor did not block the effects of VPA on development.…”

A Joint Theoretical and Experimental Study of the Behavior of the DIDS Inhibitor and its Derivatives

“…For more than three decades, 4,4′‐diisothiocyanostilbene‐2,2′‐disulfonic acid (DIDS, see Figure ) has been recognized as an inhibitor of anion transport proteins . Acting as a nonselective inhibitor, DIDS modulates membrane transporters and, consequently, regulates associated diseases .…”