Cellular Quantification of Tyrosine Hydroxylase in the Rat Brain by Immunoautoradiography

Abstract: We developed a rapid and sensitive radioimmunohistochemical method for the quantification of tyrosine hydroxylase (TH) at both the anatomical and cellular level. Coronal tissue sections from fresh-frozen rat brains were incubated in the presence of a TH monoclonal antibody. The reaction was revealed with a ³⁵S-labeled secondary antibody. TH content was quantified in catecholaminergic brain areas by measuring optical density on autoradiographic films or silver grain density on autoradiographic emulsion-coated sections. Regional TH concentrations determined in the locus ceruleus (LC), substantia nigra pars compacta (SNC), and ventral tegmental area (VTA) were significantly increased by 45% after reserpine treatment in the LC but unchanged in the SNC and VTA. Microscopic examination of TH radioimmunolabeling showed a heavy accumulation of silver grains over catecholaminergic cell bodies. In the LC, grain density per cell was heterogeneous and higher in the ventral than in the dorsal part of the structure. After reserpine treatment, TH levels were significantly increased (57%) in the neurons of the LC but not in those of the SNC or VTA. The data support the validity of this radioimmunohistochemical method as a tool for quantifying TH protein at the cellular level and they confirm that TH protein content is differentially regulated in noradrenergic and dopaminergic neurons in response to reserpine.     

Evidence for Glutamatergic Projections from the Cochlear Nucleus to the Superior Olive and the Ventral Nucleus of the Lateral Lemniscus

Abstract: This study attempts to determine if projections ascending from the guinea pig cochlear nucleus (CN) could be glutamatergic and/or aspartatergic. Multiple radio frequency lesions were made to ablate the right CN. The ablation was verified histologically. To identify the principal targets of CN efferents, silver impregnation methods were used to localize the preterminal degeneration of fibers in transverse sections of the brainstem 5 and 7 days after CN ablation. CN efferents projected heavily to the lateral superior olive (LSO) ipsilaterally, the medial superior olive (MSO) bilaterally, and contralaterally to the medial (MNTB) and ventral (VNTB) nuclei of the trapezoid body, the ventral (VNLL) and intermediate nuclei of the lateral lemniscus and the central nucleus of the inferior colliculus (ICc). There were smaller projections to the lateral nucleus of the trapezoid body ipsilaterally, the dorsal and dorsomedial periolivary nuclei bilaterally, and the dorsal nucleus of the lateral lemniscus contralaterally. There were sparse projections to the VNLL and ICc ipsilaterally and the CN contralaterally, and a very sparse projection to the contralateral LSO. To determine if CN efferents were glutamatergic and/or aspartatergic, the fresh brainstem was sectioned transversely and samples of the LSO, MSO, MNTB, VNLL, and ICc were taken to measure the electrically evoked release and the uptake of d -[³H]Asp and [¹⁴C]Gly or [¹⁴C]GABA 3–5 days after the CN ablation. The release studies suggest that only certain of the histologically identified projections ascending from the CN may be glutamatergic and/or aspartatergic. CN ablation depressed d -[³H]Asp release in the MSO bilaterally and in the contralateral MNTB and VNLL, suggesting that the CN efferents to these nuclei may use glutamate or aspartate as a transmitter. It was unclear whether a marginal depression of d -[³H]Asp release in the ipsilateral LSO reflected the presence of glutamatergic CN projections to this nucleus. d -[³H]Asp release in the ICc was unaffected, suggesting that CN efferents to this nucleus may not be glutamatergic. There were no deficits in d -[³H]Asp uptake. [¹⁴C]Gly release from the LSO and MSO was unchanged. [¹⁴C]Gly uptake was unchanged in the MSO and depressed only in the contralateral LSO, possibly reflecting subnormal uptake activity in endings contributed by contralateral MNTB cells that had lost their CN efferents. [¹⁴C]GABA uptake in the MNTB, VNLL, and ICc was unchanged. [¹⁴C]GABA release was unchanged in the VNLL and ICc. [¹⁴C]GABA release was depressed only in the contralateral MNTB, possibly reflecting the loss of a small complement of GABAergic CN efferents and the reaction of GABAergic projections from the contralateral VNTB to their loss of CN efferents.     

大鼠延髓腹外侧表面呼吸调节化学感受区的感受机制分析

用免疫组化（ＨＲＰ）、Ｈ＾＋表面透入、核团微量注射、微电泳及损毁等方法探讨了延髓腹侧表面中枢化学感受机制。结果表明它与其浅层核团：斜方体核、外周橄榄腹外侧核（ＬＶＰＯ）、斜方体后核、巨细胞旁外侧核和外侧网状核等有神经结构联系。表面Ｈ＾＋可能被上述核团的突起或胞体感受。非呼吸相关神经元（ＬＶＰＯ）与呼吸相关神经元,同样可能参与中枢化学感受而调节呼吸活动。     

Increase in proteinase activity in the cellular slime mould Polysphondylium pallidum induced by bacteria

Abstract Polysphondylium pallidum strain PPHU8 grown in association with bacteria contains aspartic and cysteine proteinases. When myxamoebae were grown in axenic medium the contribution of cysteine proteinases was much lower. The proteinase activity could be altered by addition of heat-killed bacteria to axenically growing cells. This was detected as an increase in the specific activity towards N -benzoyl-L-prolyl-L-phenylalanyl-L-arginine- p -nitroanilide, a cysteine proteinase substrate, and by the appearance of cysteine proteinase bands after electrophoretic analysis. The changes were inhibited by cycloheximide, azide and dinitrophenol. All the available evidence suggests that they are due to the de novo synthesis of cysteine proteinases.     

Morphine and Cocaine Exert Common Chronic Actions on Tyrosine Hydroxylase in Dopaminergic Brain Reward Regions

We studied levels of tyrosine hydroxylase immunoreactivity and phosphorylation state in the ventral tegmental area (VTA) and nucleus accumbens (NAc) in an effort to understand better the mechanisms by which these brain reward regions are influenced by opiates and cocaine. In the VTA, chronic, but not acute, administration of either morphine or cocaine increased levels of tyrosine hydroxylase immunoreactivity by 30-40%, with no change observed in the relative phosphorylation state of the enzyme. In the NAc, chronic, but not acute, morphine and cocaine treatments decreased the phosphorylation state of tyrosine hydroxylase, without a change in its total amount. In contrast, morphine and cocaine did not regulate tyrosine hydroxylase in the substantia nigra or caudate/putamen, brain regions generally not implicated in drug reward. Morphine and cocaine regulation of tyrosine hydroxylase could represent part of a common biochemical basis of morphine and cocaine addiction and craving.     

Neurochemical Studies of the Mesolimbic Dopaminergic Pathway: Somatodendritic Mechanisms and GABAergic Neurones in the Rat Ventral Tegmentum

Abstract: The rat ventral tegmentum (containing dendrites and somata of mesolimbic neurones) contained 1.3 μg/g of dopamine, which was reduced to 40% of the control level by reserpine. Slices of ventral tegmentum were able to accumulate and release (elevated potassium or protoveratrine A) exogenous [³H]dopamine. In parallel studies the uptake mechanism in ventral tegmentum was shown to be virtually identical to the nerve terminal uptake of [³H]dopamine by slices of nucleus accumbens. The release of [³H]dopamine was indistinguishable from that observed in substantia nigra, where there is substantial evidence for dendritic mechanisms. Basal adenylate cyclase activity was present, but dopamine-stimulated activity was not detected. A high GABA concentration (7.7 μmol/g) was present in ventral tegmentum, in conjunction with an uptake and a release mechanism for [³H]GABA. GABA and muscimol elicited a small, reproducible efflux of [³H]dopamine, but an interaction between dopamine and [³H]GABA efflux was not observed. The results are in accord with transmitter roles for dopamine and GABA in the somatoden-dritic area of mesolimbic dopaminergic neurons.     

The fine structure of ventral prostatic secretory epithelial cells in older rats

Summary The ventral prostatic secretory epithelial cells in older rats were studied by light and electron microscopy. The cells vary in height in different parts of the same organ, and ultrastructurally they show the presence of a developed secretory apparatus such as well-developed Golgi body and abundant rough endoplasmic reticulum. They also show signs of a depressed secretory activity, involving occasional emiocytosis of apical secretory vacuoles and a paucity of condensing vacuoles in the Golgi region and above it. Further, they are characterized by the frequent occurrence of supra and paranuclear pleomorphic lysosomes.     

梅毒螺旋体膜抗原基因在毕赤酵母中的表达和蛋白纯化

以梅毒螺旋体(Treponema pallidumsubsp.pallidum)Nichols菌株基因组DNA为模板,通过PCR扩增梅毒螺旋体47kDa、17kDa和15kDa 3个膜抗原基因,克隆进毕赤酵母表达载体pPICZ B,构建重组表达载体pTP47、pTP17、pTP15,转化酵母菌株GS115,甲醇诱导表达。表达菌体裂解后通过镍离子亲和层析获得3个抗原与6xHis tag的融合蛋白,重组蛋白的获得量分别为His-TP15:4.8mg/L;His-TP 17:6.6mg/L;His-TP47:25mg/L,经SDS-PAGE鉴定纯度都在96%以上,ELISA鉴定均具有很好的抗原性。从而首次在毕赤酵母中表达出梅毒螺旋体膜抗原,为梅毒血清学检测方法开辟了新的抗原制备途径。     

电化学发光法检测梅毒螺旋体特异性抗体的应用研究

目的 对罗氏电化学发光免疫法（ECLIA）检测梅毒螺旋体特异性抗体的临床价值进行评估。方法 收集梅毒疑似病例血清标本132份,分别用ECLIA、梅毒螺旋体明胶凝集试验（TPPA）和免疫印迹法（WB）进行检测,以WB为金标准,计算并比较ECLIA和TPPA的灵敏度和特异性,进而比较化学发光免疫法检测低S/CO值和高S/CO值的灵敏度和特异性差异。结果 针对132份血清标本,ECLIA敏感性为100.00%,特异性为83.33%,阳性预测值为96.43%,阴性预测值为100.00%,总符合率为96.97%。TPPA敏感性为93.52%,特异性为87.50%,阳性预测值为97.12%,阴性预测值为75.00%,总符合率为92.42%。ECLIA检测1≤S/CO＜3组与S/CO≥3组的敏感性均为100.00%,特异性分别为86.96%和95.24%,结论 ECLIA检测具有较高的敏感性,适合临床大样本筛查,对S/CO值低的标本应结合TPPA、WB及临床资料确诊。     

Hieraciumpallidum subsp. aetnense (Asteraceae), a new subspecies from Sicily (Italy), with notes on the taxonomy of H. pallidum Biv.

Hieracium pallidum subsp. aetnense Gottschl., Raimondo & Di Grist. is described and illustrated. H.pallidum is considered as an intermediate species between H. schmidtii and H. racemosum (“schmidtii>racemosum”) and is placed in H. sect. Grovesiana. By consequence, H.lanudae Gottschl. is transferred to H. pallidum as H. pallidum subsp. lanudae (Gottschl.) Gottschl., Raimondo & Di Grist. A key for the three recognised subspecies of H. pallidum is provided.