Thiele I, Warth R, Bleich M, Waldegger S, Lang F, Greger R
Physiologische Institute, Albert-Ludwigs-Universität, Freiburg, Deutschland.
Kidney Blood Press Res. 1998;21(5):317-24. doi: 10.1159/000025888.
The rectal gland of Squalus acanthias is critically involved in the homeostasis of NaCl and water metabolism and hence in overall osmoregulation. In the present study, we have examined the acute responses of rectal gland slices and in vitro perfused rectal gland tubule (RGT) cells to the exposure to dilute and hypertonic peritubule solutions. Five series were performed. (i) With changes in osmolality, Western blots to monitor tyrosine, threonine and serine phosphorylation in rectal gland slices did not reveal clear-cut changes in phosphorylation patterns. All other series were performed in in vitro perfused RGT. (ii) Relative cell volume was estimated by fura-2 fluorescence using the emission at the isosbestic excitation wavelength of 360 nm. Hypotonic solution (-100 mmol/l NaCl) reduced fura-2 fluorescence by 16% and hypertonic solution (+100 mmol/l NaCl) had the opposite effect (+12%). (iii) Transepithelial resistance was increased markedly by hypotonic solution, probably by cell swelling, and the opposite was seen with hypertonic solutions. (iv) Whole-cell patch clamp experiments indicated that hypotonic solution hyperpolarized the cells, and increased membrane conductance and membrane capacitance. The latter two changes correlated significantly with each other. Hypertonic solution had the opposite effect. (v) Measurements of the fura-2 fluorescence ratio (340/380 nm) revealed that hypotonic solution (-NaCl) increased cytosolic Ca2+ activtiy ([Ca2+]i). Hypertonic solution had no detectable effect on [Ca2+]i. These data indicate that RGT cells are swollen by removal of NaCl from the bath solution. This causes an increase in [Ca2+]i and a predominant increase in K+ conductance and hyperpolarization. Urea apparently permeates these cells quite well and its addition (+U) or its removal (-U) had only moderate osmotic effects. The removal of urea and replacement by mannitol produced effects similar to those seen with hypertonic NaCl solution.
棘鲛的直肠腺在氯化钠和水代谢的稳态中起着关键作用，因此在整体渗透压调节中也至关重要。在本研究中，我们检测了直肠腺切片和体外灌注的直肠腺小管（RGT）细胞对暴露于稀释和高渗的肾小管周溶液的急性反应。进行了五个系列的实验。（i）随着渗透压的变化，用于监测直肠腺切片中酪氨酸、苏氨酸和丝氨酸磷酸化的蛋白质免疫印迹未显示磷酸化模式有明显变化。所有其他系列实验均在体外灌注的RGT中进行。（ii）使用360 nm等吸收激发波长处的发射光，通过fura-2荧光估计相对细胞体积。低渗溶液（-100 mmol/l氯化钠）使fura-2荧光降低了16%，而高渗溶液（+100 mmol/l氯化钠）则产生相反的效果（+12%）。（iii）低渗溶液显著增加了跨上皮电阻，可能是由于细胞肿胀，而高渗溶液则产生相反的效果。（iv）全细胞膜片钳实验表明，低渗溶液使细胞超极化，并增加了膜电导和膜电容。后两种变化彼此显著相关。高渗溶液则产生相反的效果。（v）fura-2荧光比率（340/380 nm）的测量结果显示，低渗溶液（-氯化钠）增加了胞质钙活性（[Ca2+]i）。高渗溶液对[Ca2+]i没有可检测到的影响。这些数据表明，从浴液中去除氯化钠会使RGT细胞肿胀。这会导致[Ca2+]i增加，钾电导和超极化显著增加。尿素显然能很好地透过这些细胞，添加（+尿素）或去除（-尿素）尿素仅产生适度的渗透作用。去除尿素并用甘露醇替代产生的效果与高渗氯化钠溶液相似。
