吗啡戒断后焦虑症状大鼠伏核和杏仁核突触结构可塑性

Objective The possible correlations between morphological contracture and plastic variability of synaptic structure in nucleus accumbens and amygdala neurons were surveyed in anxious symptom rats suffered from morphine withdrawal. Methods The escalating doses of morphine and the elevated plus-maze were applied to validate anxiety-like behavior in rats. The electron microscope was applied to detect the parameters involving the synaptic stereology and structural plasticity of synaptic interface structure of the nucleus accumbens and amygdala neurons in the control group, the morphine-withdrawal group and the cured group ( n = 6 ), associated with the stereological ways. Results ( 1 ) Compared with the control group and the cured group, reductions of frequency and time of open-arm were observed in the morphine-withdrawal group ( P ＜ 0. 01 or P ＜ 0. 05 ). ( 2 ) Higher numerical density ( Nv ) [( 1. 012 ±0. 036 )/μm3] of synapses of nucleus accumbens was detected in the anxious rats ( P ＜ 0. 01 ) than in the controls [( 0. 701 ±0. 138 )/μm3] and the cured rats [( 0. 751 ±0. 254 )/μm3] . No significant difference between the surface density ( Sv ) and the mean profile area ( S ) of synapse of the nucleus accumbens was discovered. Compared with the control group [( 0. 247 ± 0. 117 )/μm3] and the cured one [( 0. 246 ±0. 116 )/μm3] , higher values of Nv [( 0. 427 ±0. 178 )/μm3] in amygdala were detected in anxious rats ( P＜0.01 or P＜0.05 ). Similarly, higher score of Sv [( 0.047 ±0.018 )μm2/μm3] in amygdala was observed in the anxious rats ( P ＜ 0. 01 ) than those of the control group [( 0. 030 ±0. 012 )μm2/μm3] and cured group [( 0. 030 ±0. 015 )μm2/μm3] . However, anxious rats [( 0. 124 ±0. 066 )μm2] appear to be lower S of synapse in amygdale ( P ＜ 0. 05 ) than those of the control group [( 0. 157 ±0. 119 )μm2] and the cured group [( 0. 159 ±0. 114 )μm2] . ( 3 )No significant difference among postsynaptic density, length of synaptic thickening, widths in synaptic interface structure on junctions and curvature of synaptic cleft region was detected in the nucleus accumbens and amygdala neurons. Conclusion In the present study, the results suggest that anxious rats suffered from morphine withdrawal could possibly be related to the plastic variability of synaptic morphological structure in nucleus accumbens and amygdale.     

吗啡戒断性焦虑大鼠海马突触界面结构及突触素表达变化

目的 探讨吗啡依赖戒断焦虑行为与海马CA1、CA3区突触界面结构和突触素表达变化之间的相关性.方法 剂量递增法建立大鼠吗啡依赖模型,高架十字迷宫检测焦虑行为,透射电镜技术结合图像分析系统、免疫组织化学比较对照组、模型组和治疗组(各6只)大鼠海马CA1、CA3区突触界面结构和突触素(P38)的表达.结果 (1)行为学:模型组开放臂的次数和时间均少于对照组和治疗组[最小有意义差异t检验(下同),P＜0.01或P＜0.05).(2)突触界面结构:模型组CA1区突触后致密物厚度[(10.7±0.9)nm]、突触活性区长度[(45±4)am]、突触间隙宽度[(3.80±0.30)nm]和突触界面曲率(1.37±0.12)均高于对照组和治疗组(P＜0.01或P＜0.05);模型组CA3区突触后致密物厚度[(12.7±1.1)nm]、突触活性区长度[(53±8)nm]、突触间隙宽度[(3.81 ±0.59)nm]、突触界面曲率(1.39±0.30)亦均高于对照组和治疗组(P＜0.01或P＜0.05).(3)突触素表达:模型组CA1、CA3区突触素吸光度(A)值分别为(0.42±0.06)和(0.43±0.05),显著高于对照组(0.2±0.02,0.25±0.03)和治疗组(0.27±0.04,0.26±0.03).结论吗啡戒断焦虑行为与海马CA1、CA3区突触形态结构可塑性及突触素表达水平有一定的相关性.     

慢性吗啡处理大鼠伏隔核、海马CA1区神经元突触界面结构改变

为探讨慢性吗啡处理导致的神经元突触可塑性改变，采用透射电镜技术测量慢性吗啡处理大鼠伏隔核及海马CA1区神经元突触界面结构参数，并与对照组进行比较。     

电针对老年性痴呆大鼠海马神经元突触形态可塑性的影响研究

目的探讨电针治疗老年性痴呆的突触可塑性机制,为老年性痴呆的针灸作用机理研究提供新的思路。方法 SD大鼠随机分为对照组、模型组、假手术组和电针组;以穹隆-海马伞切断进行“老年性痴呆”造模,电针“百会”、“涌泉”、“太溪”、“血海”后,电镜观察海马CA3区突触形态可塑性指标突触数密度(Nv)、突触面密度(Sv)及平均面积(S)的变化。结果模型组大鼠的Nv(2．16± 0．17)、Sv(0．09±0．02)较对照组(6．16±0．96,0．27±0．05)明显减少(P<0．01);电针组大鼠的Nv(5．08±1．02)、Sv (0．23±0．04)较模型组明显增加(P<0．01)。模型组(0．20±0．03)大鼠S较对照组(0．04±0．01)明显增加fP<0．01); 电针组大鼠的S(0．06±0．01)较模型组明显减少(P<0．01)。结论电针“百会”、“涌泉”、“太溪”、“血海”具有一定程度的促进突触可塑性发挥的作用。     

电刺激伏核对吗啡成瘾心理依赖大鼠行为学的影响

目的 通过建立脑深部刺激(deep brain stimulation,DBS)SD大鼠双侧伏核的动物模型,探讨DBS伏核对吗啡心理依赖大鼠行为学的影响.方法 40只大鼠同期依大鼠立体定向图谱行双侧伏核刺激电极的植入术,20只为吗啡假刺激组,20只为DBS组.术前(用药后第15天)、术后(刺激后第2～6天)对戒断症状进行评分,术前(用药后第15天)和术后(刺激后第2、4、6天)行条件性位置偏爱实验,研究大鼠的行为学变化40只大鼠成功地同期双侧伏核植入DBS刺激电极, 条件位置偏爱实验结果为吗啡成瘾大鼠在伴药箱平均停留时间长于非伴药箱,与生理盐水组比较具有统计学意义(P<0.01);DBS组与术前及吗啡假刺激组相比在伴药箱平均停留时间均明显缩短,具统计学意义(P<0.01),提示高频电刺激伏核能有效减轻大鼠吗啡心理依赖症状.术前戒断症状评分结果表明吗啡成瘾大鼠模型的成功建立(P<0.01);术后DBS大鼠的戒断症状与术前相比具统计学意义,证实DBS减轻了大鼠的躯体依赖症状.结论 高频刺激吗啡成瘾大鼠双侧伏核,能够有效地减轻大鼠吗啡心理依赖症状,为DBS伏核治疗吗啡等成瘾药物的心理依赖提供了实验依据.     

吗啡成瘾对大鼠伏隔核神经元自发放电的影响

目的 研究吗啡成瘾对大鼠伏隔核电生理的影响及静脉吗啡注射对吗啡成瘾大鼠伏隔核神经元自发放电的影响探讨伏隔核在吗啡成瘾过程中的作用.方法 通过连续14日递增腹腔吗啡注射,建立急性大鼠吗啡成瘾模型,通过玻璃微电极记录吗啡依赖大鼠伏隔核单细胞细胞外放电,观察吗啡成瘾及静脉注射吗啡对大鼠伏隔核神经元放电的影响.结果 与生理盐水组相比,吗啡依赖组大鼠伏隔核神经元单位自发放电的频率分布组间差异显著(P<0.05),放电形式无明显差异.吗啡依赖大鼠伏隔核神经元放电频率静脉注射吗啡前为14.40±4.92Hz,静脉注射吗啡后降为4.10±2.65Hz.结论 吗啡成瘾对大鼠伏隔核神经元自发放电有影响,吗啡可以显著抑制吗啡成瘾大鼠的伏隔核神经元放电,伏隔核在吗啡成瘾过程具有重要作用.     

脑深部电刺激对吗啡心理依赖大鼠伏核多巴胺受体的影响

目的探讨脑深部电刺激(DBS)对大鼠双侧伏核多巴胺D1A受体(D1AR)和D2受体(D2R)表达的影响以及多巴胺受体(DAR)在DBS治疗吗啡心理依赖中的作用。方法将60只SD大鼠随机分为假刺激组(ShS组)、电刺激组(DBS组)和生理盐水对照组(NS组),20只/组。用免疫组化法和RT-PCR法检测各组大鼠伏核多巴胺D1AR和D2R表达的变化。结果ShS组伏核D1AR阳性细胞数较NS组、DBS组明显增多(P0.01),而DBS组与NS组比较,差异无统计学意义(P0.05);三组间D1ARmRNA比较,差异无统计学意义(P0.05)。DBS组大鼠D2R阳性细胞数较NS组明显下降(P0.01),但较ShS组明显上升(P0.01);ShS组大鼠伏核D2RmRNA较NS组及DBS组显著上升(P0.01),而DBS组与NS组比较,差异无统计学意义(P0.05)。结论DBS对吗啡心理依赖大鼠伏核D1AR和D2R的表达起反向调节作用。     

皮层电刺激联合康复治疗对大鼠脑缺血皮层运动区突触超微结构及结构参数的影响

目的 探讨皮层电刺激联合康复锻炼对大鼠脑缺血皮层运动区突触超微结构的影响。方法 选取成年健康雄性清洁级SD大鼠30只,在大鼠训练模具中进行单个食粒抓取训练,训练7 d后记录大鼠的优势爪,训练共持续14 d,14 d后连续3 d测定每只大鼠抓取食粒成功率。纳入标准要求连续3 d抓取食粒成功率至少高于30%,共有18只大鼠入组。参照大鼠脑立体定位图谱定位脑运动区,将内皮素注射到大脑中动脉的起始点附近,阻断大脑中动脉的供血区域,制造大鼠脑梗死模型,并在相应运动皮层硬膜外放置刺激电极。脑梗死模型建立后,大鼠随机分为刺激组和对照组,每组9只,进行14 d的康复治疗：刺激组大鼠在电刺激的同时进行单个食粒抓取功能锻炼;对照组大鼠只进行单个食粒抓取功能锻炼。康复治疗14 d后对大鼠进行灌注取脑,取大鼠梗死灶周围皮层,电镜下观察突触超微结构改变,应用Image J图像分析软件,对选定的突触超微结构参数,即突触数量及突触间隙宽度等进行分析。结果 ①术后大鼠共死亡3只,死亡原因考虑为术后感染。②术前两组大鼠抓取成功率差异无统计学意义（P >0.05）,康复治疗后刺激组大鼠抓取成功率高于对照组［（31.8±8.3）% vs （18.1±4.4）%,P =0.021］。③采用Image J软件进行电镜图像分析,结果显示刺激组大鼠脑缺血皮层运动区突触数量较对照组增多［（0.0520±0.0383）个/μm3 vs （0.0387±0.0315）个/μm3,P =0.022］。两组间突触间隙宽度差异无统计学意义（P ﹥0.05）。结论 皮层电刺激联合康复治疗较单纯康复治疗能够促进大鼠脑梗死灶周围皮层运动区突触数量增加,增强了突触超微结构的可塑性。     

吗啡依赖及戒断对大鼠强啡肽原mRNA表达的影响

目的　研究吗啡依赖形成及戒断对大鼠强啡肽原mRNA表达的影响。方法　将 18只Sprague Dawley雄性大鼠随机分为吗啡依赖组、吗啡戒断组和空白对照组 ,每组 6只。采用皮下注射吗啡建立大鼠吗啡依赖模型 ,初剂量为 10mg/kg ,以后每天增加 5mg/kg ,3次 /d ,连续 6天。应用放射性3 2 磷标记的三磷酸脱氧胞苷掺入标记探针法进行Northern印迹杂交技术检测三组大鼠强啡肽原mRNA的表达水平。结果　 (1)连续 6天给予吗啡 ,吗啡依赖组大鼠下丘脑 (2 3 88± 1 6 2 )、纹状体(19 87± 2 0 3)及脊髓 (13 36± 1 4 6 )的强啡肽原mRNA相对含量分别低于对照组 (分别为 34 36±1 4 6、31 2 4± 2 83和 2 7 6 0± 2 89;均P <0 0 1) ,海马 (2 9 6 7± 3 2 3)强啡肽原mRNA相对含量高于对照组 (2 5 87± 1 74 ,P <0 0 5 ) ;(2 )给予纳洛酮处理 6 0min后 ,下丘脑 (10 2 2± 1 2 2 )、纹状体(5 90± 0 84 )、脊髓 (2 99± 0 4 8)强啡肽原mRNA的相对含量低于吗啡依赖组 (均P <0 0 1) ,而垂体强啡肽原mRNA(2 6 72± 1 79)却高于吗啡依赖组 (11 6 0± 2 2 4 ,P <0 0 1)。结论　慢性给予吗啡可影响大鼠强啡肽原mRNA的表达 ,从而参与吗啡依赖形成和戒断反应。     

Gene expression analysis of heat shock proteins in the nucleus accumbens of rats with different morphine seeking behaviours

Heat-shock proteins play functional roles on brain regulatory processes which are deeply involved in drug addiction, such as synaptic plasticity. However, few studies have been focused on gene expression of heat-shock proteins (Hsp) as potential diagnostic tools for addiction risk. This work tries to provide new knowledge on this field by using two rat models of differential vulnerability to morphine addiction in order to study differential gene expression of a selected group of Hsp genes in the nucleus accumbens (NAc). Hsp70-1A, 84, 86 and 105 genes were similarly regulated by an acute injection of morphine in two subpopulations of Sprague Dawley (SD) rats showing different rates of extinction of morphine conditioned preference. However, Lewis and Fischer rats, two strains that differ in many aspects of drug seeking behaviours, exhibited marked differences in their expression patterns of Hsp84, 86 and 105. These results suggest that differential Hsp gene expression could be related to addiction vulnerability and recommend further work to validate these proteins as potential markers for drug addiction risk.     

Effects of dopamine agonists on excitatory inputs to nucleus accumbens neurons from the amygdala: modulatory actions of cholecystokinin

The interaction of the cholecystokinin octapeptide (CCK-8) with dopamine (DA) and dopamine agonists on neurons in the nucleus accumbens was investigated using single unit recording and iontophoretic techniques in urethane-anaesthetized rats. Neurons in the nucleus accumbens were activated by single pulse stimulation of amygdala. Using seven-barrel microelectrodes, the effects of iontophoretic application of CCK-8, DA, dopamine D1 and/or D2 receptor agonists (SKF 38393 and LY 171555 respectively) were compared. The iontophoretic application of DA, LY 171555 and LY 171555 + SKF 38393 attenuated by 50-60% the excitatory responses of accumbens neurons to electrical stimulation of basolateral amygdala whereas SKF 38393 attenuated the response by less than 30%. The iontophoretic application of CCK reduced these attenuating effects of DA, LY 171555 and SKF 38393 + LY 171555. With CCK there was a rather small reduction of the attenuating effect of SKF 38393. These observations provide additional electrophysiological evidence of the interaction of CCK and dopamine and suggest that the interaction is associated mainly with dopamine D2 mechanisms.     

Effect of morphine applied by intrapallidal microdialysis on the release of dopamine in the nucleus accumbens

The effect of morphine, administered intrapallidally, on extracellular concentrations of DA, DOPAC, and HVA in the nucleus accumbens and striatum was studied in the behaving rat using the in vivo microdialysis technique. Unilateral application of morphine hydrochloride was perfomed through microdialysis probes into the rat ventral pallidum (10 μ1 of 0 2.6 4.0, 13.0, and 26.0 mM) or globus pallidus (10 μ1 of 0 and 26.0 mM). The levels of DA, DOPAC, and HVA were measured using the HPLC with EC detection in dialysates collected from the nucleus accumbens, anteromedial, and anterolateral striatum. Samples were taken every 45 min over 3 h before and over 5 h after morphine or vehicle administration. Administration of morphine into the ventral pallidum resulted in increased DOPAC and HVA concentrations in the nucleus accumbens. Pretreatment with naloxone (1 mg/kg, SC) abolished this effect of morphine. Administration of morphine into the globus pallidus resulted in increased DA, DOPAC, and HVA concentrations in the nucleus accumbens and DA in the anteromedial striatum. The levels of DA and metabolites in anterolateral striatum remained rather unchanged following morphine administered into the ventral pallidum or the globus pallidus. The changes in DA neurotransmission into the nucleus accumbens induced by morphine application into the ventral pallidum and globus pallidus are reminiscent of a phasic and tonic release of DA respectively. The results show that intrapallidal morphine increases DA neurotransmission in nucleus accumbens and suggest that the effect of morphine is mediated by ventral pallidum/mesolimbic and globus pallidus/thalamocortical pathways, depending on the site of injection.     

伏核在药物成瘾中的作用

伏核是脑奖赏中枢的重要组成部分，参与成瘾药物的强化、耐受、成瘾过程及药物戒断综合征的表达。对伏核功能解剖、受体激动与信号转导、基因转录与分子表达、神经元可塑性与行为变化等方面的深入研究，将帮助我们揭示药物成瘾的中枢机制，进而为临床戒毒治疗提供理论依据。     

Substance P release from rat nucleus accumbens and striatum: an in vivo study using antibody microprobes

Antibody-coated microprobes have provided evidence for the release of neuropeptides in localized regions of the cat spinal cord. We have applied this method to study the release of substance P (SP) from different regions of the rat brain. Anti-SP microprobes were inserted (to a depth of 8 mm) through cortex, striatum, and nucleus accumbens of halothane anaesthetised rats and remained in situ for 10 min. Microprobes (4 control and 10 post-treatment, per rat) were then incubated with¹²⁵I-SP and an autoradiographic image produced. In the region of the nucleus accumbens immunoreactive (ir) SP was detected during the first 30 min after intraperitoneal injection of d-amphetamine (4 mg/kg, P < 0.05) but not following saline (P > 0.05). During this time, no release of ir SP was seen over areas of the probes that corresponded to the striatum. At later time intervals (1–4 h) after amphetamine, binding of ir SP was detected along the whole length of the microprobes. Release of SP is thought to be due to the action of dopamine on postsynaptic cells containing this peptide. The later spread of the peptide requires further study.     

Electrophysiological evidence of modulatory interaction between dopamine and cholecystokinin in the nucleus accumbens

Dopamine has been shown to modulate responses of accumbens neurones to excitatory inputs from the amygdala. The demonstration that cholecystokinin (CCK) co-exists and appears to be co-released with dopamine in the accumbens suggests that the modulatory action of dopamine in the accumbens may in turn be modified by CCK. This possibility was investigated in the present study. Single unit recordings were obtained in the medial and caudal accumbens of urethane-anaesthetized rats. These neurones were strongly excited by amygdala stimulation, and concurrent stimulation of the ventral tegmental area (VTA) at 10 Hz attenuated the responses, presumably due to dopamine release. Iontophoretic application of proglumide (PRG) at 30 nA enhanced the attenuating effect of VTA stimulation on the excitatory response to amygdala stimulation. Exogenous dopamine produced a similar attenuation in response and the attenuation was in turn suppressed by concurrent iontophoresis of sulphated CCK fragments applied at a current titrated not to produce significant effect on the spontaneous activity of the neurone nor its response to amygdala stimulation. These results demonstrate that exogenous and endogenous CCK can modify the postsynaptic action of dopamine in the nucleus accumbens in addition to modulating its release shown in other studies, and further suggests that CCK is likely an endogenous functional antagonist of dopamine, serving a comodulatory role in regulating synaptic transmission in the ventral striatum.     

Differential basal proenkephalin gene expression in dorsal striatum and nucleus accumbens, and vulnerability to morphine self-administration in Fischer 344 and Lewis rats

We have previously shown that the acquisition rate of intravenous morphine self-administration under a fixed ratio one (FR1) schedule of reinforcement was greater in Lewis (LEW) than Fischer 344 (F344) rats. The purpose of the present experiment was to examine the relative motivational properties of morphine (1 mg/kg) or food under progressive ratio (PR) schedules of reinforcement in LEW and F344 rats. In addition, by using in situ hybridization histochemistry we have measured in both strains of rats the basal level of proenkephalin (PENK) gene expression in dorsal striatum and nucleus accumbens (NAcc). The results show that LEW rats responded to significantly higher breaking points (BPs) than F344 rats for intravenous morphine self-administration. In contrast, no differences were found in BPs for food pellets. Basal PENK mRNA levels were significantly higher in the dorsal striatum and nucleus accumbens of F344 than in LEW rats. Taken together, these results reveal a strain difference in the reinforcing efficacy of morphine and in the basal PENK gene expression in brain regions involved in the reinforcing actions of opiates. These data also suggest that the strain differences in opiate self-administration behavior found in this and other studies may be related, at least in part, to differences in basal opioid activity between LEW and F344 rats.