Localization of proopiomelanocortin mRNA in functional subsets of neurons defined by their axonal projections

In situ cDNA:mRNA hybridization is a technique that has been developed for the visualization of cDNA:mRNA hybrids in individual cells. To use this technique to answer questions of regulation in heterogeneous populations of cells in the brain, it must be combined with other procedures allowing for the identification of functional subgroups of neurons. We report here a procedure by which in situ cDNA:mRNA hybridization may be combined with retrograde axonal tracing using the fluorescent tracer fast blue. Using this technique, it now becomes possible to measure mRNA regulation in functional subsets of cells defined by their axonal projections.     

Nucleus A10 dopaminergic neurons in inbred mouse strains: Firing rate and autoreceptor sensitivity are independent of the number of cells in the nucleus

Inbred mouse strains have different numbers of midbrain dopaminergic neurons; for example, BALB/cJ mice have 20–25% more neurons than CBA/J mice. As the number of cells decrease, for example in Parkinson's disease and in animals with midbrain dopaminergic cell lesions, the activity of their remaining cells increases. The purpose of the present experiment was to determine whether the functional properties of dopaminergic neurons in the ventral tegmental area (nucleus A10) differ in inbred mouse strains which possess different numbers of cells. The firing rate and autoreceptor sensitivity of A10 dopaminergic cells were examined in the in vitro slice preparation in BALB/cJ, C3H/HeJ, CBA/J, and DBA/2J mouse strains. It was observed that the autoreceptors on mouse dopaminergic neurons exhibit pharmacological properties of dopamine autoreceptors; activation of the autoreceptor produced a marked inhibition (50–70%) in cell firing rate by quinpirole (10⁻⁸ M), LY-141865 (10⁻⁷ M), (+)-3-(3-hydroxyphenyl)-N-n-propyl-piperidine (10⁻⁶ M), propyl-norapomorphine (10⁻⁵ M) and dopamine (10⁻⁴ M), and this inhibition was blocked or reversed by specific dopamine D2 receptor antagonists [(−) sulphide and spiroperidol, 10⁻⁶ M]. The baseline firing rates of the A10 cells did not differ among the four inbred strains [range 2.5 ± 0.2 (C3H/HeJ)−3.4 ± 0.3 (CBA/J) spikes/s ±SEM], and there was no significant difference in autoreceptor sensitivity among the mouse strains as assessed either by superfused dopamine (inhibitory dose 50% ≈150 μM), or by superfused quinpirole (inhibitory dose 50% ≈10 nM). These data indicate that differences in A10 cell numbers of 30% do not significantly influence the baseline firing rate or autoreceptor sensitivity of the cells.     

急性缺氧对大鼠海马神经元钙激活钾通道的作用

目的:本实验采用膜片钳单通道技术研究缺氧条件下对培养新生大鼠海马神经元钙激活钾通道的作用;方法:取1 ～3天的SD大鼠海马神经元组织,制成细胞悬液,加入含80 %DMEM(dulbecco' s modified eagle' s medium)、20 %小牛血清培养基进行培养,将培养3 ～5天的形态典型的神经元置于对称性高钾溶液中,用膜钳技术记录单通道电流,其通道电流通过膜片钳放大器(CEZ -2200)放大,滤波(3KHz)后,经A/D、D/A转换器输入计算机,采用pClamp6 .0 .6软件采样分析;结果:在细胞贴附式下,应用氰化钠(20μmol/L)造成细胞急性缺氧,在缺氧早期(5 ～20 min)通道开放概率增加(P<0 .01或P<0 .05 ,n=5) ,而缺氧后期(20 ～30 min) ,通道开放概率明显降低,表明缺氧时间对通道的开放概率有明显影响。结论:缺氧早期神经元钙激活钾通道有自身激活作用,当钙激活钾通道激活时,钾离子外流增加,产生膜的复极化,使去极化不易产生,从而稳定细胞膜及降低细胞的兴奋性,这可能是缺氧早期神经元自身的一种代偿机制。     

A neosynthesis of sodium channels is involved in the evolution of the sodium current in isolated adult DUM neurons

Short-term culture of isolated adult dorsal unpaired median (DUM) neurons of the cockroach Periplaneta americana has been used to study the evolution of the sodium current during the time in culture after axotomy and deafferentation treatment. An increase in the maximum peak amplitude of the sodium current recorded under voltage-clamp conditions with the patch-clamp technique in the whole-cell recording configuration, was only observed between 24h and 72h (75%) without any modification of the kinetics and the voltage-dependence of the current. A decrease in the level of foetal calf serum in the culture medium reduces the amplitude of the sodium current on all days but does not affect its time-course of development which was on the contrary completely abolished by both protein synthesis inhibitors, actinomycin D and cycloheximide. The results obtained in these neurons strongly suggest that a neosynthesis of sodium channel proteins is involved in the evolution of the sodium current induced by axotomy and deafferentation.     

胫骨前肌疲劳时比目鱼肌诱发肌电图H波的变化及其机制探讨

为了解主动肌疲劳时拮抗肌脊髓运动神经元兴奋性变化的规律 ,本研究采用踝关节背屈运动形式 ,对胫骨前肌 (主动肌 )疲劳状态下的比目鱼肌 (拮抗肌 )诱发肌电图H波成分进行了观察。并以压迫阻断胫骨前肌Ⅰa类神经纤维传导的方法 ,对比目鱼肌H波变化机制进行了分析探讨。结果发现 :(1)胫骨前肌疲劳后 ,比目鱼肌H波明显受到抑制 ,与安静时比较呈非常显著性差异 ;(2 )胫骨前肌Ⅰa类神经纤维传导被阻断后 ,比目鱼肌H波的抑制现象没有解除。表明 ,胫骨前肌疲劳时比目鱼肌H波被抑制的原因 ,可能是由于主动肌内的代谢产物激活了Ⅲ·Ⅳ类神经纤维的感受器 ,Ⅲ·Ⅳ类神经纤维的传入冲动增加 ,使Ⅰa抑制性中间神经元被激活 ,导致拮抗肌脊髓运动神经元的兴奋性受到了抑制     

基质金属蛋白酶与肺癌转移的关系

基质金属蛋白酶(matrix metalloproteinases，MMPs)是一组锌离子依赖的蛋白水解酶，在ECM降解中起着重要作用。目前研究表明，MMPs活性异常不仅能促进肿瘤血管形成，还是人类多种肿瘤侵袭和转移的必要条件。本文就与肺癌侵袭和转移有关的MMPs研究作一综述。     

活性氧对神经母细胞株SH-SY5Y细胞L型钙通道电流的影响

①目的 探讨氧自由基导致神经元钙超载的发生机制。②方法 在神经母细胞SH—SY5Y细胞体外培养的基础上，采用全细胞膜片钳技术，观察H2O2对SH—SY5Y细胞L型钙通道电流(ICa,L)的影响。③结果 H2O2能够明显增加峰值ICa,L，且该作用具有浓度依赖性。H2O2使ICa,L的I-V相关曲线下移，但对其形状和最大激活电位无明显影响。④结论 H2O2对L型钙通道有明显增强作用，此可能是其导致神经元钙超载的机制之一。     

大鼠周龄及切片厚度对脊髓片培养神经细胞活性的影响

目的探讨离体培养脊髓片的理想厚度和大鼠的最佳周龄。方法通过不同周龄的大鼠(1周、2周、3周、4周龄各10只,共40只),以及不同厚度(切片厚度分为:100μm、200μm、300μm以及400μm 4种)的脊髓组织切片培养,采用MTT技术进行脊髓片培养细胞的活性鉴定,观察脊髓片培养不同时间点其神经细胞的活性。结果1~7 d鼠龄的脊髓组织片培养1~14 d时细胞活性最高,但其神经细胞结构形态容易受到破坏,不利于观察,且在培养超过14 d时神经细胞活性明显下降;2~3周鼠龄的脊髓组织片培养细胞,在培养14 d后仍有较高的神经细胞的活性并保持较好的细胞形态结构;1~4各周龄组大鼠200μm和300μm左右厚度的脊髓片培养神经细胞活性最稳定(85%~95%)。结论进行离体脊髓片培养,选择7~14 d(即1~2周龄)大鼠作为取材对象较为理想,选择200μm~300μm切片厚度的脊髓片进行培养较为合适。     

血液流变学因素对缺血心肌冠脉流量的影响

选用长白猪８只于麻醉下开胸，于在体猪心上探讨心肌缺血时冠脉仍保留有部分扩张储备，而未被充分利用的机理。实验结果表明：降低左冠状动脉前降支（ＬＡＤ）内压至４．６７ｋＰａ时，由ＬＡＤ分别输入腺苷、山莨菪碱、生理盐水（２ｍｌ／ｍｉｎ），皆能诱发出明显的ＬＡＤ流量增加（Ｐ＜０．０５）。在维持ＬＡＤ内压不变，并持续分别输入上述三种溶液的同时，从ＬＡＤ远端取血测血液流变学指标，结果显示，输入生理盐水和山莨菪碱使红细胞压积降低，血浆粘度和全血粘度下降；心肌缺血时，冠脉内输注药物所诱发出的“冠脉扩张储备”部分是由于此种给药方式所引起的血液稀释及血液粘度下降的结果。它提示在冠脉狭窄时，血液粘度对狭窄远端的心肌血供可能是一个相当重要的因素，降低血液粘度可能并不亚于扩血管药物的作用。