视觉发育期大鼠视皮层锥体神经细胞被动膜特性研究

发布时间：2018-01-16 22:07

本文关键词：视觉发育期大鼠视皮层锥体神经细胞被动膜特性研究　出处：《天津医科大学》2012年硕士论文　论文类型：学位论文

【摘要】：目的研究Wistar大鼠初级视皮层第Ⅱ/Ⅲ层锥体神经元被动膜特性在视觉发育不同时期的变化规律,探讨不同饲养条件下视觉经验变化对被动膜特性参数的影响。方法选取出生后14-60天健康SPF级Wistar大鼠,雌雄不限将其分成两大组,一组在正常光照环境下饲养(12h白天/黑夜交替环境)；另一组从出生开始就放入全黑暗环境中饲养。再将每组分成五小组：视觉发育前关键期组(P13-15)；视觉发育关键期早期组(P16-20)；视觉发育关键期高峰期组(P25-27)；视觉发育关键期后期组(P30-35)；视觉发育后关键期组(P55-60)。应用红外微分干涉相差显微镜(IR-DIC)结合电耦合式摄像机(CCD-Camera),在可视法下应用全细胞膜片钳技术记录：在电流钳模式下,不做任何钳制,使细胞保持在静息电位,之后通过记录电极注入40pA、持续时间为100ms的电流,反复十次,根据诱导的超级化电压变化计算时间常数；在电压钳模式下,以-5mv为步阶,记录-50mv—-100mv及-100mv—-50mv的不同钳制电位下电流的变化,从而计算出细胞膜电阻的不同变化趋势,观察视觉发育不同阶段以及不同饲养条件下大鼠视皮层神经元被动膜特性的变化规律。结果 1.时间常数在两种饲养条件下随视觉发育的变化趋势：正常条件饲养下p13-15组的时间常数(28.79±6.68ms)明显大于p25-27组的时间常数(21.51±4.64ms)(P0.05,t=4.000)。而在暗饲养条件下,p13-15组的时间常数(38.71±6.04ms)也明显大于p25-27组的时间常数(19.13±7.35ms)(P0.05,t=9.202)。时间常数随视觉发育而下降。同时,暗饲养条件下,p13-15组的时间常数(38.71±6.04ms)明显大于正常条件饲养下p13-15组的时间常数(28.79±6.68ms)(P0.05,t=4.92)；但随着生长发育,到视觉发育关键期时,暗饲养条件下(19.13±7.35ms)与正常条件饲养下(21.51±4.64ms)的时间常数,无明显统计差异。 2.膜电阻在两种饲养条件下随视觉发育的变化趋势：正常饲养环境下,p13-15组的膜电阻(131.03±40.78MΩ)明显大与p16-20组的膜电阻(100.47±22.94MΩ)(P0.05,t=2.59)；同时,p30-35组的膜电阻(82.38±20.43MΩ)明显大于p55-60组的膜电阻(66.25±18.93MΩ)(P0.05,t=2.13),其他分组间虽无统计学差异,但膜电阻的整体趋势表现为随视觉发育而下降。而暗饲养环境下,膜电阻的整体趋势也表现为随视觉发育而下降。 3.膜电阻在不同钳制电位下的变化趋势：钳制电位从-50MV—100MV时,膜电阻都表现为由低到高的变化趋势；当钳制电位从-100MV—50MV时,膜电阻表现为由高到低的变化趋势。总之,大鼠视皮层第二层锥体神经元的膜电阻在去极化状态,电阻降低；而在超级化状态,电阻升高。 4.将视觉发育关键期前后所有年龄段的大鼠视皮层第二层锥体细胞的时间常数和膜电阻值作为两个相关变量,应用Pearson相关分析法进行相关分析：相关系数r=0.275(P=0.033),二者具有正相关性：膜电阻值越大,时间常数越大。结论 1、正常饲养和暗饲养两种条件下,大鼠初级视皮层第Ⅱ/Ⅲ层锥体神经元的被动膜特性——时间常数和膜电阻,都具有相似的变化趋势,即都表现为随视觉发育而逐渐下降。 2、本实验中同一时期暗饲养条件下的被动膜特性数值虽高于正常饲养条件下的数值,但两者的差异不具有统计学意义,提示年龄可能是影响被动膜特性的决定性因素。 3.大鼠视皮层第Ⅱ/Ⅲ层锥体神经元的膜电阻在去极化状态,电阻降低；而在超级化状态,电阻升高。 4.视觉发育关键期前后所有年龄段的大鼠视皮层第二层锥体细胞的时间常数和膜电阻值具有正相关性：膜电阻值越大,时间常数越大。
[Abstract]:objective
Objective to study the changes of passive membrane characteristics of Wistar / II pyramidal neurons in different stages of visual development, and to explore the effects of visual experience on passive membrane parameters under different feeding conditions.
Method
From postnatal day 14-60 healthy SPF Wistar rats, male and female will be divided into two groups, one group in normal light environment feeding (12h day / night alternating environment) and another group; from birth into the whole feeding dark environment. Then each group was divided into five groups: visual development front the key period group (P13-15); the critical period of visual development early group (P16-20); the peak of critical period of visual development group (P25-27); the critical period of visual development stage group (P30-35); after the critical period of visual development group (P55-60). The application of infrared differential interference contrast microscopy (IR-DIC) combined with electrical coupling camera (CCD-Camera) and using the whole cell patch clamp technique in visual method: in the current clamp mode, without any control, keeps cells in the resting potential after injected through the recording electrode of 40pA, duration of 100ms current, repeated ten times, according to the induced super Level of voltage change calculation time constant; in voltage clamp mode, -5mv step, the current changes in different holding potentials recorded -50mv - -100mv and -100mv - -50mv, to calculate the changing trend of cell membrane resistance, variation characteristics of passive film visual cortical neurons observed in different developmental stages and different visual the feeding conditions of rats.
Result
The 1. time constant in the two feeding conditions change with visual development conditions: normal feeding time constant of p13-15 group (28.79 + 6.68ms) time constant was significantly higher than that of p25-27 group (21.51 + 4.64ms) (P0.05, t=4.000). While in the dark rearing conditions, the time constant of p13-15 group (38.71. 6.04ms) time constant was significantly greater than group p25-27 (19.13 + 7.35ms) (P0.05, t=9.202). The time constant decreased with visual development. At the same time, dark rearing conditions, the time constant of the p13-15 group (38.71 + 6.04ms) time constant is significantly higher than the normal rearing conditions of group p13-15 (28.79 + 6.68ms (P0.05) t=4.92); but, with the development of the critical period of visual development, dark rearing conditions (19.13 + 7.35ms) with normal rearing conditions (21.51 + 4.64ms) time constant, no significant statistical difference.
The 2. film resistance in the two feeding conditions change with visual development environment: normal feeding, film resistance of group p13-15 (131.03 + 40.78M) significantly and p16-20 group of film resistance (100.47 + 22.94M) (P0.05, t=2.59); at the same time, the membrane resistance of p30-35 group (82.38 + 20.43M. The membrane resistance was significantly higher than that of p55-60) group (66.25 + 18.93M) (P0.05, t=2.13), although there was no statistically difference between other groups, but the overall trend of the membrane resistance is decreased with the visual development. Dark rearing environment, the overall trend of the membrane resistance as well as with the visual development and decline.
The change trend in different holding potentials of the 3. film resistance: holding potential from -50MV - 100MV, the membrane resistance from low to high trend; when the holding potential from -100MV to 50MV, the membrane resistance from high to low trend. In short, the rat visual cortical pyramidal neurons membrane resistance second in the depolarized state, reduce the resistance; and in the super state, the resistance increased.
4. before and after the critical period of visual development of all ages of rat optic time constant and the membrane resistance layer of pyramidal cells in cortex of second values as two dependent variables, using Pearson correlation analysis method for correlation analysis: correlation coefficient r=0.275 (P=0.033), two had positive correlation: the membrane resistance value is greater, the greater the time constant.
conclusion
1, under the two conditions of normal feeding and dark feeding, the passive membrane characteristics of the second / third tier pyramidal neurons of primary visual cortex in rats showed similar trend of time and membrane resistance, which all showed gradual decrease with visual development.
2, in this experiment, the passive membrane characteristics of dark feeding under the same period were higher than that of normal feeding conditions, but the difference between them was not statistically significant, suggesting that age may be the decisive factor affecting the passive membrane characteristics.
The membrane resistance of the second / third layer pyramidal neurons in the visual cortex of 3. rats was depolarized and the resistance decreased, while the resistance was elevated in the superchemical state.
4. there was a positive correlation between the time constant and the membrane resistance value of the second pyramidal cells in the visual cortex of all age groups before and after the critical period of visual development: the greater the membrane resistance, the greater the time constant.

【学位授予单位】：天津医科大学
【学位级别】：硕士
【学位授予年份】：2012
【分类号】：R77

【参考文献】