Smooth Muscle Research Centre, Dundalk Institute of Technology, Dundalk, Ireland.
Am J Physiol Cell Physiol. 2010 Nov;299(5):C1180-94. doi: 10.1152/ajpcell.00028.2010. Epub 2010 Aug 18.
Hyaluronan, a joint lubricant and regulator of synovial fluid content, is secreted by fibroblast-like synoviocytes lining the joint cavity, and secretion is greatly stimulated by Ca(2+)-dependent protein kinase C. This study aimed to define synoviocyte membrane currents and channels that may influence synoviocyte Ca(2+) dynamics. Resting membrane potential ranged from -30 mV to -66 mV (mean -45 ± 8.60 mV, n = 40). Input resistance ranged from 0.54 GΩ to 2.6 GΩ (mean 1.28 ± 0.57 GΩ; ν = 33). Cell capacitance averaged 97.97 ± 5.93 pF. Voltage clamp using C(s+) pipette solution yielded a transient inward current that disappeared in Ca(2+)-free solutions and was blocked by 1 μM nifedipine, indicating an L-type calcium current. The current was increased fourfold by the calcium channel activator FPL 64176 (300 nM). Using K(+) pipette solution, depolarizing steps positive to -40 mV evoked an outward current that showed kinetics and voltage dependence of activation and inactivation typical of the delayed rectifier potassium current. This was blocked by the nonspecific delayed rectifier blocker 4-aminopyridine. The synoviocytes expressed mRNA for four Kv1 subtypes (Kv1.1, Kv1.4, Kv1.5, and Kv1.6). Correolide (1 μM), margatoxin (100 nM), and α-dendrotoxin block these Kv1 subtypes, and all of these drugs significantly reduced synoviocyte outward current. The current was blocked most effectively by 50 nM κ-dendrotoxin, which is specific for channels containing a Kv1.1 subunit, indicating that Kv1.1 is critical, either as a homomultimeric channel or as a component of a heteromultimeric Kv1 channel. When 50 nM κ-dendrotoxin was added to current-clamped synoviocytes, the cells depolarized by >20 mV and this was accompanied by an increase in intracellular calcium concentration. Similarly, depolarization of the cells with high external potassium solution caused an increase in intracellular calcium, and this effect was greatly reduced by 1 μM nifedipine. In conclusion, fibroblast-like synoviocytes cultured from the inner synovium of the rabbit exhibit voltage-dependent inward and outward currents, including Ca(2+) currents. They thus express ion channels regulating membrane Ca(2+) permeability and electrochemical gradient. Since Ca(2+)-dependent kinases are major regulators of synovial hyaluronan secretion, the synoviocyte ion channels are likely to be important in the regulation of hyaluronan secretion.
透明质酸是一种关节润滑剂和滑液含量的调节剂，由关节腔内的成纤维细胞样滑膜细胞分泌，其分泌受 Ca(2+)-依赖性蛋白激酶 C 的强烈刺激。本研究旨在确定可能影响滑膜细胞 Ca(2+)动力学的滑膜细胞膜电流和通道。静息膜电位范围为-30 mV 至-66 mV（平均值-45 ± 8.60 mV，n = 40）。输入电阻范围为 0.54 GΩ 至 2.6 GΩ（平均值 1.28 ± 0.57 GΩ；ν = 33）。细胞电容平均为 97.97 ± 5.93 pF。使用 C(s+) 玻璃微电极溶液进行电压钳位，产生一种瞬时内向电流，该电流在无 Ca(2+)溶液中消失，并被 1 μM 硝苯地平阻断，表明存在 L 型钙电流。钙通道激活剂 FPL 64176（300 nM）使电流增加四倍。使用 K(+) 玻璃微电极溶液，正于-40 mV 以上的去极化步骤诱发一种外向电流，其激活和失活动力学和电压依赖性与延迟整流钾电流典型特征一致。这被非特异性延迟整流钾通道阻滞剂 4-氨基吡啶阻断。滑膜细胞表达四种 Kv1 亚型（Kv1.1、Kv1.4、Kv1.5 和 Kv1.6）的 mRNA。Correolide（1 μM）、margaritoxin（100 nM）和α-dendrotoxin 阻断这些 Kv1 亚型，所有这些药物均显著降低滑膜细胞外向电流。50 nM κ-dendrotoxin 最有效地阻断电流，κ-dendrotoxin 特异性针对含有 Kv1.1 亚基的通道，表明 Kv1.1 作为同源多聚体通道或作为异源多聚体 Kv1 通道的组成部分至关重要。当将 50 nM κ-dendrotoxin 添加到电流钳制的滑膜细胞中时，细胞超极化超过 20 mV，这伴随着细胞内钙浓度的增加。同样，用高钾溶液去极化细胞会导致细胞内钙增加，而 1 μM 硝苯地平大大降低了这种效应。总之，从兔滑膜内层培养的成纤维细胞样滑膜细胞表现出电压依赖性内向和外向电流，包括 Ca(2+)电流。因此，它们表达调节膜 Ca(2+)通透性和电化学梯度的离子通道。由于 Ca(2+)-依赖性激酶是滑膜透明质酸分泌的主要调节剂，因此滑膜细胞离子通道可能在透明质酸分泌的调节中起重要作用。
