Functional aspects of serotonin transmission in the basal ganglia: a review and an in vivo approach using the push-pull cannula technique

Peripheral deafferentation of the rodent olfactory bulb results in loss of dopamine content, tyrosine hydroxylase activity and immunocytochemical staining for tyrosine hydroxylase in juxtaglomerular dopamine neurons. Reinnervation of the bulb by afferent neurons results in the return of all parameters to control levels suggesting that the dopamine neurons did not degenerate but that the expression of tyrosine hydroxylase enzyme was transneuronally regulated in a static population of juxtaglomerular cells. To evaluate this possibility, we determined the activity and immunocytochemical localization of the second enzyme in the dopamine biosynthetic pathway, DOPA decar?ylase. At a time when tyrosine hydroxylase activity was reduced to 25% of control values, DOPA decar?ylase activity in the lesioned bulb was maintained at about 65% of that in the unlesioned bulb. Immunocytochemical staining with antibodies to both enzymes, performed sequentially in the same sections, demonstrated that in the unlesioned bulb tyrosine hydroxylase and DOPA decar?ylase are co-localized in the same population of juxtaglomerular neurons. Similar results were obtained in adjacent sections each stained with one of the two antibodies. In contrast, in the deafferented bulb, about three times as many neurons were stained with DOPA decar?ylase as with tyrosine hydroxylase antibodies. The DOPA decar?ylase activity measurements and immunocytochemistry argue for the continued presence, in the lesioned olfactory bulb, of a population of tyrosine hydroxylase deficient dopamine neurons.The data suggest that olfactory receptor cell innervation transneuronally regulates the expression of tyrosine hydroxylase by mechanisms separate from those controlling the levels of DOPA decar?ylase.     

8OHdG levels in brain do not indicate oxidative DNA damage in Alzheimer''s disease

Accumulation of oxidative DNA damage has been proposed to underlie aging and neurodegenerative diseases such as Alzheimer's Disease (AD). The DNA adduct 8-hydroxy-2′-deoxyguanosine (8OHdG) is considered a good indicator of oxidative DNA damage. To investigate whether this type of DNA damage is involved in AD etiology, 8OHdG levels were determined in postmortem human brain tissue of controls and AD patients (in frontal, occipital, and temporal cortex and in hippocampal tissue). Parametric studies in rat revealed no influences of postmortem delay, repeated freezing/thawing or storage time. In human brain, approximately two 8OHdG molecules were present per 10⁵ 2′-deoxyguanosines. In AD patients and controls, 8OHdG-levels were not related to age, sex, or brain region. Also, no differences were found between controls and AD patients. It was concluded that 8OHdG in nuclear DNA, although present throughout the brain in fairly high amounts, does not accumulate with age, nor does it appear to be involved in AD. More detailed studies are required to extend this conclusion to other types of oxidative damage.     

格列齐特中亚硝酸盐的电化学测定法

目的：建立高效液相-电化学(ECD)检测法测定格列齐特中亚硝酸盐(NO2^-)的残留量。方法：阳离子色谱柱，醋酸盐缓冲液(pH为4．3)为流动相。结果：NO2^-线性范围0．04～0．80μg，r=0．9995，回收率为99．2％，RSD=1．7％(n=9)。结论：方法灵敏，简便，结果准确，     

高效液相色谱-电化学法测定四季青叶中4种化合物的含量

目的:建立同时测定四季青叶中原儿茶酸、咖啡酸、槲皮素和山奈酚4种成分含量的高效液相色谱-电化学(HPLC-ECD)法。方法:采用 Zorbax SB-C_(18)(150 mm×4.6 mm,5.0 μm)色谱柱;以(A)甲醇、(B)水-0.1%醋酸为流动相,梯度洗脱:0min 时 A-B(20:80),9 min 时 A-B(24:76),30 min 时 A-B(50:50),40 min 时 A-B(65:35);流速为0.8 mL·min~(-1),柱温为30℃,电化学检测器的工作电位为0.7 V,采用标准曲线法对4种化合物进行定量。结果:在选定的色谱条件下原儿茶酸、咖啡酸、槲皮素和山奈酚的检出限分别为3,6,2,5 ng,定量限分别为9.1,20.0,7.2,16.0 ng。4种化合物的平均加样回收率均在95.7%～102.6%之间,RSD 在1.6%～2.9%之间。结论:方法简便、准确,可用于同时检测四季青叶中4种化合物的含量。     

Effects of mu, kappa, and delta opioid receptor agonists and antagonists on rat hypothalamic noradrenergic neurotransmission

We have investigated the effects of specific mu-, kappa-and delta-opioid receptor agonists and antagonists on the hypothalamic noradrenergic neurotransmission and on luteinising hormone (LH) release in the ovariectomised and steroid-primed rat. The opioid agents were infused intracerebroventricularly under ketamine anaesthesia and blood samples collected at hourly intervals on the afternoon of the anticipated LH surge. At the end of the experiment, the rats were decapitated and the medial preoptic area, suprachiasmatic nucleus, median eminence and arcuate nucleus surgically isolated by micropunch. The concentrations of noradrenaline (NA) and its metabolite (3,4-dihydroxyphenylglycol; DHPG) in these samples was determined by high performance liquid chromatography with electrochemical detection. Plasma LH levels were measured by radioimmunassay. The three opioid agonists reduced concentrations of NA and DHPG in all four hypothalamic areas. These inhibitory effects of the opioid agonists were mostly prevented following coadministration with their respective antagonists. However, naloxone had no significant effect on DHPG levels in any of the hypothalamic regions examined. Plasma LH levels were found to be either low or undetectable in all groups. These results suggest that mu-, kappa- and delta-opioid receptors have inhibitory influence on the hypothalamic noradrenergic neurotransmission around the time of the LH surge. It is thought that the ketamine anaesthesia interfered with LH release.     

银杏叶片中银杏黄酮在Beagle犬体内的药动学研究

目的以槲皮素为代表,研究银杏叶片中银杏黄酮类成分在Beagle犬体内的药动学特征。方法 Beagle犬口服给药银杏叶片后,按设定时间静脉采血。血样经固相萃取、酸水解后,以HPLC-ECD测定不同时间点血浆中槲皮素的浓度。应用DAS数据处理软件对数据进行统计学处理。结果口服给药后,槲皮素在Beagle犬体内的药动学过程符合二房室模型,其主要药动学参数为t1/2z5.67 h,AUC0-t63.8 g.L 1.h,Cmax15.9 g.L 1。结论建立的方法简单、灵敏、准确,可用于研究以槲皮素为代表的银杏黄酮类成分的药动学特征,为银杏叶片的临床应用提供了依据。     

In vitro studies of endogenous noradrenaline and NPY overflow from the rat hypothalamus during maturation and ageing

Marked changes in brain monoamine content and NPY content occur during maturation and ageing. Earlier in vivo studies in our laboratory have reported blunted K⁺ stimulated noradrenaline release and reduced NPY overflow in aged animals using microdialysis and push pull techniques. In this study, in vitro superfusion techniques were established to measure endogenous noradrenaline, NPY, DOPAC and 5-HIAA overflow from the hypothalamus of 1, 5 and 16 month old Sprague-Dawley rats. A period of high K⁺ (56 mM) stimulation was carried out to elicit maximal release. Basal noradrenaline overflow was similar in all age groups of rats and during K⁺-induced depolarisation similar 3–4 fold increases were observed. On the other hand, basal and K⁺ stimulated NPY overflow were significantly greater in the adult rats compared to 1 month and 16 month old rats. Despite differences in absolute NPY overflow, the relative increase over resting was not significantly different across age groups. The molar quantities of hypothalamic NPY overflow at rest and under K⁺ stimulated conditions were three orders of magnitude lower than noradrenaline. Results of these studies suggest that both NPY and noradrenaline can be released from a similar hypothalamic pool. Basal and K⁺-evoked DOPAC and 5-HIAA overflow were similar between the 3 age groups. Thus the overflow of hypothalamic noradrenaline, DOPAC and 5-HIAA under in vitro conditions was not altered from 1 to 16 months. In contrast, 5 month old rats had significantly higher NPY overflow than the other age groups (P < 0.05), consistent with a reported decline in NPY content with advanced age. Hypothalamic noradrenaline overflow was not affected by ageing, suggesting that a selective loss of NPY in the arcuo-PVN projection, or other projections to the hypothalamus with ageing may contribute to the reduction in NPY overflow in aged rats. Received: 9 June 1997 / Accepted: 6 November 1997     

A rapid assay for neurotransmitter amino acids,aspartate, glutamate,glycine, taurine and γ-aminobutyric acid in the brain by high-performance liquid chromatography with electrochemical detection

Summary For simultaneous assay of the five neurotransmitter amino acids, Asp, Glu, Gly, Tau, and GABA in brain tissues, a very rapid and simple chromatographic method using high-performance liquid chromatography with electrochemical detection in combination with o-phthalaldehyde derivatization is described. Because the present method permits the determination of these five amino acids within less than five minutes in one chromatographic run, up to 100 samples a working day can be analyzed using an autosampler. Withinrun coefficients of variation for these five amino acids were less than 2% (n=20). The quantitative detection limit was 2.5 pmol for the 5 amino acids. The present method has been applied to the measurement of the five amino acid neurotransmitter levels in several discrete brain regions of mice treated with and without electroconvulsive shock.     

Formalin pain increases the concentration of serotonin and its 5-hydroxyindoleacetic acid metabolite in the CA1 region of hippocampus

Background and the purpose of the study

The hippocampal formation is involved in nociception. Prenatal serotonin depletion results in a significant decrease in the concentration of nociceptive sensitivity during the second phase of behavioral response in the formalin test.

Methods

A microdialysis probe was inserted via a guide cannula into the right CA1 region of the hippocampus. Extracellular serotonin (5HT) and its 5- hydroxyindoleacetic acid (5HIAA) metabolite overflow were collected every 10 min during the formalin test and measured by HPLC with electrochemichal detector.

Results

Compared to the sham group, formalin injection in the hind paw of the rat significantly increased 5HT after 10, 30, 40, and 50 min and increased 5HIAA after 10, 30, 40, 50, and 60 min collection time periods in hippocampal dialysate. (n=6 for each group at each sampling time). In the formalin treated rats serotonin and 5HIAA concentrations increased in the biphasic pattern in concert with the first and second phases of formalin pain.

Conclusion

The hippocampal formation might be involved in the processing of nociceptive information and serotonin-related mechanisms in the hippocampus may play a role in the biphasic behavioral responses to formalin noxious stimulation.     

BD总皂苷对营养性肥胖大鼠纹状体细胞外液中5-HT、5-HIAA含量的影响

目的：研究BD总皂苷对营养性肥胖大鼠纹状体细胞外液中单胺类神经递质含量的影响,探讨了BD总皂苷减肥调脂作用的可能机制。方法：灌胃6周后,定位至大鼠左侧纹状体,于第3天进行大鼠纹状体微透析试验,将微透析液分别注入高效液相-电化学系统,测定各组纹状体透析液中单胺类神经递质五羟色胺（5-HT）、五羟基吲哚乙酸（5-HIAA）的含量。结果：与肥胖模型组相比,BD总皂苷高剂量组、低剂量组和阳性对照组纹状体透析液中5-HT、5-HIAA的含量升高,具有统计学意义（P〈0.05或P〈0.01）,说明BD总皂苷减肥作用可能与5-HT的含量升高有关。结论：BD总皂苷具有减肥和调节血脂作用,其作用可能与通过抑制5-HT、的再摄取而减少食物摄入有关。