School of Life Sciences, University of Nottingham, Nottingham, UK.
Faculty of Pharmacy, Mahasarakham University, Mahasarakham, Thailand.
J Endocrinol. 2020 Feb;244(2):369-381. doi: 10.1530/JOE-19-0493.
L-type channel antagonists are of therapeutic benefit in the treatment of hyperlipidaemia and insulin resistance. Our aim was to identify L-type voltage-gated Ca2+ channels in white fat adipocytes, and determine if they affect intracellular Ca2+, lipolysis and lipogenesis. We used a multidisciplinary approach of molecular biology, confocal microscopy, Ca2+ imaging and metabolic assays to explore this problem using adipocytes isolated from adult rat epididymal fat pads. CaV1.2, CaV1.3 and CaV1.1 alpha1, beta and alpha2delta subunits were detected at the gene expression level. The CaV1.2 and CaV1.3 alpha1 subunits were identified in the plasma membrane at the protein level. Confocal microscopy with fluorescent antibodies labelled CaV1.2 in the plasma membrane. Ca2+ imaging revealed that the intracellular Ca2+ concentration, [Ca2 +]i was reversibly decreased by removal of extracellular Ca2+, an effect mimicked by verapamil, nifedipine and Co2+, all blockers of L-type channels, whereas the Ca2+ channel agonist BAY-K8644 increased [Ca2+]i. The finding that the magnitude of these effects correlated with basal [Ca2+]i suggests that adipocyte [Ca2+]i is controlled by L-type Ca2+ channels that are constitutively active at the adipocyte depolarized membrane potential. Pharmacological manipulation of L-type channel activity modulated both basal and catecholamine-stimulated lipolysis but not insulin-induced glucose uptake or lipogenesis. We conclude that white adipocytes have constitutively active L-type Ca2+ channels which explains their sensitivity of lipolysis to Ca2+ channel modulators. Our data suggest CaV1.2 as a potential novel therapeutic target in the treatment of obesity.
L 型通道拮抗剂在治疗高脂血症和胰岛素抵抗方面具有治疗益处。我们的目的是鉴定白色脂肪细胞中的 L 型电压门控 Ca2+通道，并确定它们是否影响细胞内 Ca2+、脂肪分解和脂肪生成。我们使用分子生物学、共聚焦显微镜、Ca2+成像和代谢测定的多学科方法，使用从成年大鼠附睾脂肪垫中分离的脂肪细胞来探索这个问题。在基因表达水平检测到 Cav1.2、Cav1.3 和 Cav1.1α1、β和α2δ亚基。在蛋白质水平上鉴定出 Cav1.2 和 Cav1.3α1 亚基在质膜中。用荧光抗体进行共聚焦显微镜检查，标记质膜中的 Cav1.2。Ca2+成像显示，细胞外 Ca2+的去除可使细胞内 Ca2+浓度[Ca2+]i可逆性降低，维拉帕米、硝苯地平和 Co2+（均为 L 型通道阻滞剂）模拟了这种作用，而 Ca2+通道激动剂 BAY-K8644 增加了[Ca2+]i。这些作用的幅度与基础[Ca2+]i 相关的发现表明，脂肪细胞[Ca2+]i 由 L 型 Ca2+通道控制，L 型 Ca2+通道在脂肪细胞去极化膜电位下持续活跃。L 型通道活性的药理学操纵调节了基础和儿茶酚胺刺激的脂肪分解，但不调节胰岛素诱导的葡萄糖摄取或脂肪生成。我们得出结论，白色脂肪细胞具有持续活跃的 L 型 Ca2+通道，这解释了它们对 Ca2+通道调节剂的脂肪分解敏感性。我们的数据表明 Cav1.2 可能是肥胖治疗的潜在新治疗靶点。
