Department of Pharmacology and Toxicology, Dokkyo Medical University, 880 Kitakobayashi, Mibu, Tochigi 321-0293, Japan.
Department of Electrical and Mechanical Engineering, Graduate School of Engineering, Nagoya Institute of Technology, Nagoya 466-8555, Japan.
Int J Mol Sci. 2023 May 17;24(10):8905. doi: 10.3390/ijms24108905.
Previous studies demonstrated that hamster sperm hyperactivation is suppressed by extracellular Na by lowering intracellular Ca levels, and Na/Ca-exchanger (NCX) specific inhibitors canceled the suppressive effects of extracellular Na. These results suggest the involvement of NCX in the regulation of hyperactivation. However, direct evidence of the presence and functionality of NCX in hamster spermatozoa is still lacking. This study aimed to reveal that NCX is present and is functional in hamster spermatozoa. First, NCX1 and NCX2 transcripts were detected via RNA-seq analyses of hamster testis mRNAs, but only the NCX1 protein was detected. Next, NCX activity was determined by measuring the Na-dependent Ca influx using the Ca indicator Fura-2. The Na-dependent Ca influx was detected in hamster spermatozoa, notably in the tail region. The Na-dependent Ca influx was inhibited by the NCX inhibitor SEA0400 at NCX1-specific concentrations. NCX1 activity was reduced after 3 h of incubation in capacitating conditions. These results, together with authors' previous study, showed that hamster spermatozoa possesses functional NCX1 and that its activity was downregulated upon capacitation to trigger hyperactivation. This is the first study to successfully reveal the presence of NCX1 and its physiological function as a hyperactivation brake.
先前的研究表明，通过降低细胞内钙水平，细胞外钠离子抑制了仓鼠精子的超激活，而钠离子/钙交换器（NCX）特异性抑制剂则取消了细胞外钠离子的抑制作用。这些结果表明 NCX 参与了对超激活的调节。然而，仓鼠精子中 NCX 的存在和功能的直接证据仍然缺乏。本研究旨在揭示 NCX 存在于并在仓鼠精子中具有功能性。首先，通过对仓鼠睾丸 mRNA 的 RNA-seq 分析检测到了 NCX1 和 NCX2 的转录本，但仅检测到了 NCX1 蛋白。接下来，通过使用 Ca 指示剂 Fura-2 测量 Na 依赖性 Ca 内流来确定 NCX 活性。在仓鼠精子中检测到了 Na 依赖性 Ca 内流，尤其是在尾部区域。NCX1 特异性浓度的 NCX 抑制剂 SEA0400 抑制了 Na 依赖性 Ca 内流。在获能条件下孵育 3 小时后，NCX1 活性降低。这些结果与作者之前的研究一起表明，仓鼠精子具有功能性的 NCX1，其活性在获能时下调以引发超激活。这是首次成功揭示 NCX1 的存在及其作为超激活制动器的生理功能的研究。
