Adaptive thermogenesis in brown adipose tissue involves activation of pannexin-1 channels,Molecular Metabolism

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Adaptive thermogenesis in brown adipose tissue involves activation of pannexin-1 channels
Molecular Metabolism ( IF 8.1 ) Pub Date : 2020-11-25 , DOI: 10.1016/j.molmet.2020.101130
Subramanian Senthivinayagam ₁ , Vlad Serbulea ₁ , Clint M Upchurch ₁ , Renata Polanowska-Grabowska ₁ , Suresh K Mendu ₁ , Srabani Sahu ₁ , Prathiba Jayaguru ₂ , Kevin W Aylor ₃ , Mahendra D Chordia ₄ , Limor Steinberg ₁ , Nathaniel Oberholtzer ₁ , Seichii Uchiyama ₅ , Noriko Inada ₆ , Ulrike M Lorenz ₇ , Thurl E Harris ₁ , Susanna R Keller ₃ , Akshaya K Meher ₁ , Alexandra Kadl ₂ , Bimal N Desai ₁ , Bijoy K Kundu ₈ , Norbert Leitinger ₉

Affiliation

Department of Pharmacology, University of Virginia School of Medicine, Charlottesville, VA, 22908, USA.
Division of Pulmonary and Critical Care Medicine, Department of Medicine, University of Virginia, Charlottesville, VA, 22908, USA.
Division of Endocrinology and Metabolism, Department of Medicine, University of Virginia, Charlottesville, VA, 22908, USA.
Department of Radiology and Medical Imaging, University of Virginia School of Medicine, Charlottesville, VA, 22908, USA.
Graduate School of Pharmaceutical Sciences, The University of Tokyo, Tokyo, Japan.
Graduate School of Life and Environmental Sciences, Osaka Prefecture University, Osaka, Japan.
Department of Microbiology, Immunology and Cancer Biology, Center for Cell Clearance, the Beirne B. Carter Center for Immunology Research, USA.
Department of Radiology and Medical Imaging, University of Virginia School of Medicine, Charlottesville, VA, 22908, USA; Department of Biomedical Engineering, University of Virginia School of Medicine, Charlottesville, VA, 22908, USA.
Department of Pharmacology, University of Virginia School of Medicine, Charlottesville, VA, 22908, USA; Robert M Berne Cardiovascular Research Center, University of Virginia, Charlottesville, VA, 22908, USA.

Objective

Brown adipose tissue (BAT) is specialized in thermogenesis. The conversion of energy into heat in brown adipocytes proceeds via stimulation of β-adrenergic receptor (βAR)-dependent signaling and activation of mitochondrial uncoupling protein 1 (UCP1). We have previously demonstrated a functional role for pannexin-1 (Panx1) channels in white adipose tissue; however, it is not known whether Panx1 channels play a role in the regulation of brown adipocyte function. Here, we tested the hypothesis that Panx1 channels are involved in brown adipocyte activation and thermogenesis.

Methods

In an immortalized brown pre-adipocytes cell line, Panx1 currents were measured using patch-clamp electrophysiology. Flow cytometry was used for assessment of dye uptake and luminescence assays for adenosine triphosphate (ATP) release, and cellular temperature measurement was performed using a ratiometric fluorescence thermometer. We used RNA interference and expression plasmids to manipulate expression of wild-type and mutant Panx1. We used previously described adipocyte-specific Panx1 knockout mice (Panx1^Adip-/-) and generated brown adipocyte-specific Panx1 knockout mice (Panx1^BAT-/-) to study pharmacological or cold-induced thermogenesis. Glucose uptake into brown adipose tissue was quantified by positron emission tomography (PET) analysis of ¹⁸F-fluorodeoxyglucose (¹⁸F-FDG) content. BAT temperature was measured using an implantable telemetric temperature probe.

Results

In brown adipocytes, Panx1 channel activity was induced either by apoptosis-dependent caspase activation or by β3AR stimulation via a novel mechanism that involves Gβγ subunit binding to Panx1. Inactivation of Panx1 channels in cultured brown adipocytes resulted in inhibition of β3AR-induced lipolysis, UCP-1 expression, and cellular thermogenesis. In mice, adiponectin-Cre-dependent genetic deletion of Panx1 in all adipose tissue depots resulted in defective β3AR agonist- or cold-induced thermogenesis in BAT and suppressed beigeing of white adipose tissue. UCP1-Cre-dependent Panx1 deletion specifically in brown adipocytes reduced the capacity for adaptive thermogenesis without affecting beigeing of white adipose tissue and aggravated diet-induced obesity and insulin resistance.

Conclusions

These data demonstrate that Gβγ-dependent Panx1 channel activation is involved in β3AR-induced thermogenic regulation in brown adipocytes. Identification of Panx1 channels in BAT as novel thermo-regulatory elements downstream of β3AR activation may have therapeutic implications.

中文翻译：

棕色脂肪组织的适应性产热涉及 pannexin-1 通道的激活

客观的

棕色脂肪组织（BAT）专门负责产热。棕色脂肪细胞中能量转化为热量是通过刺激 β-肾上腺素能受体 (βAR) 依赖性信号传导和激活线粒体解偶联蛋白 1 (UCP1) 进行的。我们之前已经证明了 pannexin-1 (Panx1) 通道在白色脂肪组织中的功能作用；然而，尚不清楚 Panx1 通道是否在棕色脂肪细胞功能的调节中发挥作用。在这里，我们测试了 Panx1 通道参与棕色脂肪细胞活化和产热的假设。

方法

在永生化的棕色前脂肪细胞中s使用膜片钳电生理学测量细胞系 Panx1 电流。使用流式细胞术评估染料摄取和三磷酸腺苷 (ATP) 释放的发光测定，并使用比率荧光温度计进行细胞温度测量。我们使用 RNA 干扰和表达质粒来操纵野生型和突变型 Panx1 的表达。我们使用先前描述的脂肪细胞特异性 Panx1 敲除小鼠 (Panx1 ^Adip-/- ) 并生成棕色脂肪细胞特异性 Panx1 敲除小鼠 (Panx1 ^BAT-/- ) 来研究药理学或冷诱导的生热作用。通过正电子发射断层扫描（PET）分析¹⁸ F-氟脱氧葡萄糖（¹⁸ F-FDG）含量来量化棕色脂肪组织中的葡萄糖摄取。使用植入式遥测温度探头测量 BAT 温度。

结果

在棕色脂肪细胞中，Panx1 通道活性是由凋亡依赖性 caspase 激活或通过涉及 Gβγ 亚基与 Panx1 结合的新机制刺激 β3AR 诱导的。培养的棕色脂肪细胞中 Panx1 通道的失活导致 β3AR 诱导的脂肪分解、UCP-1 表达和细胞产热的抑制。在小鼠中，所有脂肪组织库中 Panx1 的脂联素-Cre 依赖性基因缺失导致 BAT 中 β3AR 激动剂或冷诱导的生热作用缺陷，并抑制白色脂肪组织的米色化。UCP1-Cre依赖性Panx1缺失（特别是在棕色脂肪细胞中）降低了适应性产热的能力，但不影响白色脂肪组织的米色化并加剧饮食诱导的肥胖和胰岛素抵抗。