Hayato R, Matsumoto T, Higure Y
Laboratory of Anatomy and Physiology, School of Nutritional Sciences, Nagoya University of Arts and Sciences, Takenoyama, Nissin-City, Aichi, Japan.
Physiol Res. 2024 Mar 11;73(1):69-80. doi: 10.33549/physiolres.935071.
beta3-adrenergic activation causes Ca2+ release from the mitochondria and subsequent Ca2+ release from the endoplasmic reticulum (ER), evoking store-operated Ca2+ entry (SOCE) due to Ca2+ depletion from the ER in mouse brown adipocytes. In this study, we investigated how Ca2+ depletion from the ER elicits SOCE in mouse brown adipocytes using fluorometry of intracellular Ca2+ concentration ([Ca2+]i). The administration of cyclopiazonic acid (CPA), a reversible sarcoplasmic/endoplasmic reticulum calcium ATPase (SERCA) pump blocker in the ER, caused an increase in [Ca2+]i. Moreover, CPA induced SOCE was suppressed by the administration of a Ca2+ free Krebs solution and the transient receptor potential canonical 6 (TRPC6) selective blockers 2-APB, ML-9 and GsMTx-4 but not Pico145, which blocks TRPC1/4/5. Administration of TRPC6 channel agonist 1-oleoyl-2-acetyl-sn-glycerol (OAG) and flufenamic acid elicited Ca2+ entry. Moreover, our RT-PCR analyses detected mRNAs for TRPC6 in brown adipose tissues. In addition, western blot analyses showed the expression of the TRPC6 protein. Thus, TRPC6 is one of the Ca2+ pathways involved in SOCE. These modes of Ca2+ entry provide the basis for heat production via activation of Ca2+-dependent dehydrogenase and the expression of uncoupling protein 1 (UCP1). Enhancing thermogenic metabolism in brown adipocytes may serve as broad therapeutic utility to reduce obesity and metabolic syndrome.
β3-肾上腺素能激活导致线粒体释放Ca2+，随后内质网（ER）释放Ca2+，由于小鼠棕色脂肪细胞内质网Ca2+耗竭，引发了储存-操作性Ca2+内流（SOCE）。在本研究中，我们使用细胞内Ca2+浓度（[Ca2+]i）荧光测定法，研究了内质网Ca2+耗竭如何在小鼠棕色脂肪细胞中引发SOCE。给予环匹阿尼酸（CPA），一种内质网中可逆的肌浆网/内质网钙ATP酶（SERCA）泵阻滞剂，导致[Ca2+]i升高。此外，给予无Ca2+的Krebs溶液以及瞬时受体电位香草酸亚型6（TRPC6）选择性阻滞剂2-氨基乙氧基二苯硼酸（2-APB）、ML-9和GsMTx-4可抑制CPA诱导的SOCE，但阻断TRPC1/4/5的Pico145则不能。给予TRPC6通道激动剂1-油酰-2-乙酰-sn-甘油（OAG）和氟芬那酸可引发Ca2+内流。此外，我们的逆转录-聚合酶链反应（RT-PCR）分析在棕色脂肪组织中检测到了TRPC6的信使核糖核酸（mRNA）。另外，蛋白质免疫印迹分析显示了TRPC6蛋白的表达。因此，TRPC6是参与SOCE的Ca2+途径之一。这些Ca2+内流模式为通过激活Ca2+依赖性脱氢酶和表达解偶联蛋白1（UCP1）产热提供了基础。增强棕色脂肪细胞的产热代谢可能具有广泛的治疗用途，以减轻肥胖和代谢综合征。
