Background
IAA-94 (Indanyloxyacetic acid-94) is an intracellular chloride channel blocker [1]. IAA-94 inhibited CLIC-1-dependent chloride permeability with an apparent IC50 of 8.6 μM [2]. IAA-94 depressed the K+-induced force with an IC50 of 17.0 ± 1.2 μM [3].
In cells treated with IAA-94 (30 μM) with respect to control, the amount of intracellular Aβ1-42 phagocytosis was remarkably increased after 24 hr of incubation [4]. Inhibition of Cl- channels with pan-chloride channel blocker IAA-94 (50 μM) abolished the protection induced by 200 nM CsA pre-treatment in cardiomyocytes [5]. The anion channel blocker IAA-94 (200 μM) significantly blocked glutamate and taurine release against Astrocytes in hypotonic solution [6].
IAA-94 was shown to abrogate cardioprotection mediated by IPC and cyclosporin A (CsA) [7]. Adult male rat hearts were subjected to the left coronary artery occlusion for 45min followed by 3h of reperfusion. A single intravenous bolus of phosphate buffered saline (PBS) (control) or IAA-94 (20mg/kg body weight) was administered 5min before reperfusion. The infarct size (IS) was significantly higher in the IAA-94-treated group compared to control; the ratio of IS to area at risk was 58.1±7% in IAA-94 vs. 33.5±6% in control in this rat model [1].
参考文献:
[1]. Ponnalagu D, Hussain A T, Thanawala R, et al. Chloride channel blocker IAA-94 increases myocardial infarction by reducing calcium retention capacity of the cardiac mitochondria[J]. Life sciences, 2019, 235: 116841.
[2]. Tulk B M, Schlesinger P H, Kapadia S A, et al. CLIC-1 functions as a chloride channel when expressed and purified from bacteria[J]. Journal of Biological Chemistry, 2000, 275(35): 26986-26993.
[3]. Doughty J M, Miller A L, Langton P D. Non‐specificity of chloride channel blockers in rat cerebral arteries: block of the L‐type calcium channel[J]. The Journal of physiology, 1998, 507(2): 433-439.
[4]. Paradisi S, Matteucci A, Fabrizi C, et al. Blockade of chloride intracellular ion channel 1 stimulates Aβ phagocytosis[J]. Journal of neuroscience research, 2008, 86(11): 2488-2498.
[5]. Diaz R J, Fernandes K, Lytvyn Y, et al. Enhanced cell-volume regulation in cyclosporin A cardioprotection[J]. Cardiovascular research, 2013, 98(3): 411-419.
[6]. Ye Z C, Oberheim N, Kettenmann H, et al. Pharmacological “cross‐inhibition” of connexin hemichannels and swelling activated anion channels[J]. Glia, 2009, 57(3): 258-269.
[7]. Diaz R J, Losito V A, Mao G D, et al. Chloride channel inhibition blocks the protection of ischemic preconditioning and hypo-osmotic stress in rabbit ventricular myocardium[J]. Circulation research, 1999, 84(7): 763-775.
IAA-94 (Indanyloxyacetic acid-94) 是一种细胞内氯离子通道阻滞剂[1]。 IAA-94 抑制 CLIC-1 依赖性氯离子渗透性，表观 IC50 为 8.6 μM [2]。 IAA-94 抑制 K+- 诱导力，IC50 为 17.0 ± 1.2 μM [3]。
相对于对照，在用 IAA-94 (30 μM) 处理的细胞中，细胞内 Aβ1-42 吞噬量在孵育 24 小时后显着增加 [4]。用泛氯离子通道阻滞剂 IAA-94 (50 μM) 抑制 Cl- 通道消除了 200 nM CsA 预处理对心肌细胞诱导的保护作用[5]。阴离子通道阻滞剂 IAA-94 (200 μM) 显着阻断谷氨酸和牛磺酸在低渗溶液中对星形胶质细胞的释放[6]。
IAA-94 被证明可以消除由 IPC 和环孢菌素 A (CsA) [7] 介导的心脏保护作用。对成年雄性大鼠心脏进行左冠状动脉闭塞 45 分钟，然后再灌注 3 小时。再灌注前 5 分钟给予单次静脉推注的磷酸盐缓冲盐水 (PBS)（对照）或 IAA-94（20mg/kg 体重）。与对照组相比，IAA-94 治疗组的梗塞面积 (IS) 明显更高；在该大鼠模型中，IS 与风险面积的比率在 IAA-94 中为 58.1±7%，而在对照组中为 33.5±6%[1]。
Protocol
| Cell experiment [1]: |
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Cell lines
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murine microglial cell line BV-2, Primary microglial cell
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Preparation Method
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Flow cytometry was used to measure BV-2 cell uptake of fluorescent beads after stimulation with Aβ25-35, the active fragment of Aβ, and Aβ35-25, the reverse, inactive form. Cells were treated with aggregated Aβ25-35 (50 μM) for 30 min, 90 min, and 3 hr in the presence or not of the CLIC-1 inhibitor IAA-94 and with Aβ35-25 (50 μM) for 3 hr. Treatments were performed on cells in suspension at 37°C; afterward, cells were washed with medium.
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Reaction Conditions
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30 μM for 3 h
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Applications
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Primary microglia showed a phagocytic ability that remained substantially unmodified after Aβ treatment. Treatment with IAA-94 did not modify the phagocytic ability of these cells.
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| Animal experiment [2]: |
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Animal models
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Male CD-1 mice weighing 25-30 g
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Preparation Method
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Groups of mice were treated i.t. with a selective δ-opioid receptor agonist [D-Ala2]deltorphin II (10 μg) in 10% 2-hydroxypropyl-β-cyclodextrin/saline solution (5 μl), and the antinociception induced by [D-Ala2]deltorphin II was measured for 1 h after the treatment. In order to ascertain the role of Ca2+-activated Cl- channels in the expression of δ-opioid receptor-mediated antinociception, a selective Ca2+-activated Cl- channel blocker IAA-94(2% Tween 20/saline solution, 1, 3, 10 μg in 5 μl) was pretreated i.t. 10 min prior to the [D-Ala2]deltorphin II injection.
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Dosage form
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1, 3, 10 μg in 5 μl, i.t.
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Applications
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Mice injected [D-Ala2]deltorphin II showed a marked antinociception (peak effect: 79.5±11.1% MPE). The antinociception induced by [D-Ala2]deltorphin II was attenuated by i.t.-pretreatment with IAA-94 in a dose-dependent manner, and the peak effect was significantly reduced to 44.8±10.0% MPE by 10 μg of IAA-94.
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参考文献:
[1]: Paradisi S, Matteucci A, Fabrizi C, et al. Blockade of chloride intracellular ion channel 1 stimulates Aβ phagocytosis[J]. Journal of neuroscience research, 2008, 86(11): 2488-2498.
[2]:Yamazaki M, Mizoguchi H, Ohsawa M, et al. Implications of Ca2+-activated Cl- channels in the δ-opioid receptor-mediated antinociception in the mouse spinal cord[J]. Neuroscience letters, 2000, 295(3): 113-115.
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