The regulation factor of intracellular Ca^<2+> level in taste transduction

牛蛙（Rana catesbeiana）用于实验。从断头和去髓动物的舌中分离味细胞。在全细胞配置中进行电压钳记录。膜电位保持在-50 mV。通过电压斜坡获得电流-电压（I-V）关系（167 mV/s）从-100至100 mV。当移液管充满含有1 mM EGTA的K^+-内部溶液时，在静息状态下保持Ca^<2+>水平为17 nM，味觉细胞对3 μ M离子霉素的Ca^2+-离子载体的反应显示出-50 mV的持续内向电流。同时去除细胞外Na^+和Ca^<2+>可完全消除离子霉素诱导的电流，表明细胞内Ca^<2+>的升高可促进非选择性阳离子通道的开放。用1 mM Ca^2+或50 μ M三磷酸肌醇（IP_3）的K^+内溶液进行细胞内透析，也能诱导类似的-50 mV内向电流。用104 mM Cl^-，GTP_<gamma>S（0.5 mM）透析，在约40%的视杆细胞中诱导了-50 mV的快速瞬时和缓慢持续电流。当内部Cl^-浓度从104 mM降至10 mM时，慢电流成分消失，但仍能观察到快电流，表明快电流是阳离子性的。向移液器中的GTP_ S添加10 muMU 73211（磷脂酶C抑制剂）<gamma>可将GTP_<gamma>S诱导的电流降低至对照的23%。结果表明，在蛙味觉信号转导中，IP_3-Ca^&lt;2+&gt;级联反应与G蛋白的兴奋电流相耦合。

The bullfrogs (Rana catesbeiana) were used for the experiments. Taste cells were isolated from the tongue of decapitated and pithed animals. Voltage clamp recording was performed in the whole-cell configuration. The membrane potentials were held to -50 mV.The current-voltage (I-V) relationships were obtained by a voltage ramp (167 mV/s) from -100 to 100 mV.When the pipette was filled with a K^+-internal solution containing 1 mM EGTA which kept Ca^<2+> level of 17 nM at resting state, taste cells displayd sustained inward current at -50 mV in response to a Ca^<2+>-ionophore 3 muM ionomycin. Simultaneous removals of external Na^+ and Ca^<2+> eliminated the ionomycin-induced current completely, suggesting that the rise of intracellular Ca^<2+> elicits the opening of nonselective cation channels. Intracellular dialysis with 1 mM Ca^<2+> or 50 muM inositol 1,4,5-trisphosphate (IP_3) in K^+-internal solution also induced an similar inward current at -50 mV.With internal 104 mM Cl^-, GTP_<gamma>S (0.5 mM) dialysis induced fast transient and slow sustained currents at -50 mV in about 40% of rod cells examined. When internal Cl^- was reduced from 104 to 10 mM,slow component disappeared, but fast one was still observed, suggesting that fast current is cationic. Addition of 10 muMU73211 (inhibitor of phospholipase C) to GTP_<gamma>S in the pipette decreased the GTP_<gamma>S-induced current to 23% of the control. Tne results suggest that the IP_3-Ca^<2+> cascade coupled to G protein elicits cation current in frog taste transduction.