维司力农结构改造物FPA对豚鼠心肌细胞钙电流的影响

Effects of Derivatives of Vesnarinone FPA on L-type Calcium Current in Isolated Ventricular Myocytes of Guinea Pig

目的用膜片钳全细胞记录法观察FPA对分离的单个豚鼠心室肌细胞L型钙电流(Ica-L)的影响。方法采用急性酶分离法(胶原酶+白蛋白)分离单个豚鼠心室肌细胞,在生物倒置显微镜下对豚鼠心室肌细胞进行膜片钳全细胞记录。实验采用空白试验组,加药组(FPA组),二甲亚砜组进行对造,在实验中FPA以三种浓度给药(0.75,1.0和1.25 mmol.L-1)。实验中保持电位为-40 mV,刺激频率为0.1 Hz,刺激时间200 ms,以10 mV为一阶跃逐级去极化到+60 mV,通道电流经膜片钳负反馈放大器放大,再用pClamp 7.0软件的数据采集系统采样和最后分析。结果FPA增加Ica-L。0.75 mmol.L-1FPA使Ica-L最大峰值电流从(4.1181±1.404)pA/pF增至(4.907±1.208)pA/pF(n=6,P均<0.05),增加率为19.16%;1.0 mmol.L-1FPA使Ica-L最大峰值电流从(4.519±1.106)pA/pF增至(7.483±2.154)pA/pF(n=6,P均<0.05),增加率为65.58%;1.25 mmol.L-1FPA使Ica-L最大峰值电流从(3.288±1.038)pA/pF增至(8.345±0.172)pA/pF(n=4,P均<0.05),增加率为153.80%。FPA使Ica-L的电流-电压曲线下移,但不改变其激活、峰值和反转电位,亦不改变细胞的静息电位。结论FPA对Ica-L的促进作用为其强心效应的离子基础之一。

OBJECTIVE To investigate the effects of vesnarinone derivatives FPA on L-type calcium channel current（Ica-L） of isolated single ventricular cells from guinea pig hearts using patch clamp whole-cell recording.METHODS Single ventricular myocytes were obtained by an enzymatic dissociation procedure.Ica-L was activated by voltage –clamp steps commands applied from a holding potential（-40 mV） to various depolarizing levels.0.75,1.0 and 1.25 mmol·L^-1 FPA were tested in different groups.Channel current was amplified by path-clamp amplifier,then imported to computer through A/D.D/A converter.PClamp 7.0 software was used in data recording and analysis.RESULTS Ica-L was increased from（4.118±1.404）pA/pF to（4.907±1.208）pA/pF（n=6,P〈0.05 respectively） by 0.75mM FPA,from（4.519±1.106）pA/pF to（7.483±2.154）pA/pF（n=6,P〈0.05 respectively） by 1.0 mmol·L^-1 FPA,and from（3.288±1.038）pA/pF to（8.345±0.172）pA/pF（n=4,P〈0.05 respectively） by 1.25 mmol·L^-1 FPA.Their raise rates were 19.16%,65.58% and 153.80%,respectively.FPA shifts down the I-V curve of Ica-L,while its active,peak value,reverse potentials and resting potential did not changed.The results suggest that FPA increases Ica-L,which may contribute to its inotropic action.CONCLUSION FPA directly acts on calcium channel and increases Ca2＋ inward to cause positive inotropic action,The results indicated that FPA had evident positive inotropic action.