Abstract
The aim of the study was to investigate the mechanisms of Ca2+ oscillation generation upon activation of connexin-43 and regulation of the lipolysis/lipogenesis balance in white adipocytes through vesicular ATP release. With fluorescence microscopy it was revealed that a decrease in the concentration of extracellular calcium ([Ca2+]ex) results in two types of Ca2+ responses in white adipocytes: Ca2+ oscillations and transient Ca2+ signals. It was found that activation of the connexin half-channels is involved in the generation of Ca2+ oscillations, since the blockers of the connexin hemichannels—carbenoxolone, octanol, proadifen and Gap26—as well as Cx43 gene knockdown led to complete suppression of these signals. The activation of Cx43 in response to the reduction of [Ca2+]ex was confirmed by TIRF microscopy. It was shown that in response to the activation of Cx43, ATP-containing vesicles were released from the adipocytes. This process was suppressed by knockdown of the Cx43 gene and by bafilomycin A1, an inhibitor of vacuolar ATPase. At the level of intracellular signaling, the generation of Ca2+ oscillations in white adipocytes in response to a decrease in [Ca2+]ex occurred due to the mobilization of the Ca2+ ions from the thapsigargin-sensitive Ca2+ pool of IP3R as a result of activation of the purinergic P2Y1 receptors and phosphoinositide signaling pathway. After activation of Cx43 and generation of the Ca2+ oscillations, changes in the expression levels of key genes and their encoding proteins involved in the regulation of lipolysis were observed in white adipocytes. This effect was accompanied by a decrease in the number of adipocytes containing lipid droplets, while inhibition or knockdown of Cx43 led to inhibition of lipolysis and accumulation of lipid droplets. In this study, we investigated the mechanism of Ca2+ oscillation generation in white adipocytes in response to a decrease in the concentration of Ca2+ ions in the external environment and established an interplay between periodic Ca2+ modes and the regulation of the lipolysis/lipogenesis balance.
Highlights
Introduction published maps and institutional affilThere are two main types of adipose tissue: white and brown White adipose tissue (WAT) stores energy as triglycerides in lipid droplets, whereas brown adipose tissue (BAT)determines adaptive thermogenesis and dissipates energy as heat
When the complete medium was replaced with calcium-free medium supplemented with 0.5 mM EGTA (Ca2+ -free), different Ca2+ releases were observed in mature white adipocytes (9 days in vitro, DIV)
Ca2+ oscillations occurred in 45 ± 11% of cells; Ca2+ oscillations were observed in 23 ± 8% of cells after a lag period of 5.2 ± 3.5 min, and transient Ca2+ signals were recorded in 32 ± 11% of adipocytes (Figure 1A)
Summary
Introduction published maps and institutional affilThere are two main types of adipose tissue: white and brown White adipose tissue (WAT) stores energy as triglycerides in lipid droplets, whereas brown adipose tissue (BAT)determines adaptive thermogenesis and dissipates energy as heat. There are two main types of adipose tissue: white and brown White adipose tissue (WAT) stores energy as triglycerides in lipid droplets, whereas brown adipose tissue (BAT). Obesity induces a complex remodeling of adipose tissue, which expands to accommodate the excessive caloric intake, with marked changes in its structure and cellular composition, as a result leading to WAT dysfunction [1]. WAT dysfunction is accompanied by excess caloric intake, leading to type 2 diabetes, obesity-driven insulin resistance and an increased susceptibility to many pathophysiological changes, including cardiovascular disease and cancer [2,3,4]. White adipocytes form fat depots in different parts of the body and are the main type of bone marrow cells; their number increases in various pathological conditions, including radiation therapy [5]. Adipocytes—which occurs as inclusions in white fat depots and are characterized by a iations.
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