尼古丁处理对大鼠脑内多巴胺转运体和酪氨酸羟化酶的影响

目的 观察尼古丁处理大鼠脑内多巴胺转运体(DAT)和酪氨酸羟化酶(TH)的表达变化,探讨尼古丁处理对大鼠脑内多巴胺(DA)能神经体系的影响. 方法 选用雄性Wistar大鼠按每日 0.4 mg/kg 腹腔注射尼古丁 7d;利用免疫组织化学和免疫印迹法,检测尼古丁处理大鼠有关脑区DAT和TH的表达改变. 结果 与对照组相比:1. 免疫组织化学显示,尼古丁处理组大鼠伏核(NACC)和腹则被盖区(VTA)的DAT灰度值降低了12.43 %和12.85 %;TH的灰度值则降低了11.87 %和10.09 %.2. 免疫印迹法显示,尼古丁处理组大鼠尾壳核(CPu)-NACC、黑质(SN)-VTA的DAT与β-肌动蛋白(β-actin)条带相对吸光度比值增加了75.68 %和117.14 %;而TH的比值则分别增加了66.32 %和60.31 %. 结论 尼古丁处理增加大鼠脑内DAT和TH的表达,这可能与尼古丁的成瘾机制有关.     

大鼠纹状体边缘区多巴胺免疫阳性反应物的分布

目的　研究纹状体边缘区多巴胺和酪氨酸免疫阳性物质的分布。方法　用免疫细胞化学ABC法检测多巴胺和酪氨酸羟化酶免疫阳性物质在大鼠纹状体边缘区的分布。结果　多巴胺和酪氨酸羟化酶免疫阳性纤维在边缘区中密集分布形成一条明显的带状 ,带的宽度和位置与边缘区一致。结论　证明了纹状体边缘区中存在密度较高的多巴胺和酪氨酸羟化酶免疫阳性纤维 ,并讨论了这些免疫阳性纤维的来源以及与学习记忆的关系。     

Repeated stimulation of D1 dopamine receptors causes time-dependent alterations in the sensitivity of both D1 and D2 dopamine receptors within the rat striatum.

Recent evidence suggests that repeated stimulation of D1 dopamine receptors within the rat striatum leads to an enhancement of both D1 and D2 dopamine receptor-mediated responses. The present study used both behavioral observations and extracellular single unit recording techniques to investigate this phenomenon following repeated administration of selective D1 dopamine receptor agonists. Groups of rats received twice daily administration of either saline or the partial D1 dopamine receptor agonist SKF 38393 (8 mg/kg, s.c.) for three weeks. Rats were tolerant to the ability of SKF 38393 to enhance grooming behavior when tested immediately following the last of the 42 treatment injections. However, the ability of this last SKF 38393 injection to potentiate oral stereotyped behavior following administration of the D2 DA agonist quinpirole was still evident. Following a one-day withdrawal, grooming responses to SKF 38393 had returned to normal. At this time, administration of quinpirole, without concomitant SKF 38393, failed to significantly promote oral stereotypies, as is typical of normal rats. Following a one-week withdrawal period, SKF 38393-induced grooming behavior was significantly enhanced and quinpirole, administered without SKF 38393, produced pronounced oral stereotyped behavior in 10 of 12 rats tested. Following a one-month withdrawal, these sensitized responses were no longer evident. Single-cell recordings from rat lateral striatal neurons revealed similar time-dependent alterations in the effects of iontophoretically administered SKF 38393 and quinpirole. Current-response curves revealed that, without a withdrawal period, striatal neurons were subsensitive to the inhibitory effects of SKF 38393 but not quinpirole. The decreased inhibitory responses of striatal neurons to SKF 38393 returned to normal levels after a one-day withdrawal. Following a one-week withdrawal, the effects of both agonists were significantly greater than that in saline-treated controls. Normosensitivity was evident following a one-month withdrawal. Repeated administration of the full D1 DA agonist SKF 81297 (0.5 mg/kg, s.c., twice daily) also resulted in sensitized responses of striatal neurons following a one-week withdrawal, demonstrating that the sensitization to SKF 38393 was not due to its partial agonist character. The present findings provide both behavioral and electrophysiological evidence that repeated stimulation of D1 dopamine receptors results in a brief subsensitivity, followed by transient sensitization of the D1 receptors. The enhanced effects of D2 dopamine agonists might be due to an enhanced synergism (enabling) produced by endogenous dopamine stimulating supersensitive D1 receptors.(ABSTRACT TRUNCATED AT 400 WORDS)     

Methamphetamine-induced reduction in D1 and D2 dopamine receptors as evidenced by autoradiography: comparison with tyrosine hydroxylase activity

As determined by autoradiographic techniques, multiple high doses of methamphetamine elicited a reduction in dopamine receptor population (both D1 and D2) in several areas of the rat central nervous system. D1 receptors were labeled with the D1-selective antagonist, [3H]SCH 23390, and D2 receptors were labeled with the D2-selective neuroleptic, [3H]sulpiride. Scatchard analysis, obtained from saturation data in caudate-putamen, indicated that the receptor alterations were due to a decrease in the number of receptors (Bmax) without an apparent change in affinity (Kd). A time course demonstrated that five doses of methamphetamine were required to elicit significant changes in receptors in most brain areas examined. The onset of the receptor alterations in various brain regions correlated with the development of methamphetamine-induced depression of striatal tyrosine hydroxylase activity. In most brain areas, the dopamine receptors returned to normal within 7 days following methamphetamine.     

Functions of dopamine in the dorsal and ventral striatum

Manipulations of dopamine levels in the dorsal and ventral striatum are shown to affect the activation of behaviour in distinct, yet parallel ways, which depend upon the nature of the neocortical and limbic input to these structures. Whereas dopamine in the dorsal striatum contributes to the sensorimotor co-ordination of consummatory behaviour and the development of a ‘response set’ in motor preparatory processes for skilled responses, dopamine in the ventral striatum influences the impact of reward-related stimuli on appetitive aspects of behaviour. The circumstances under which the striatal dopamine projections are normally active to effect these functions are defined by studies which attempt to correlate firing in single units or neurochemical indices of dopamine activity with environmental conditions, internal states and behaviour.     

The distribution of dopamine D1 receptor and mu-opioid receptor 1 receptor immunoreactivities in the amygdala and interstitial nucleus of the posterior limb of the anterior commissure: relationships to tyrosine hydroxylase and opioid peptide terminal systems

Mismatches between dopamine innervation and dopamine D1 receptor (D1) distribution have previously been demonstrated in the intercalated cell masses of the rat amygdala. Here the distribution of enkephalin and beta-endorphin immunoreactive (IR) nerve terminals with respect to their mu-opioid receptors is examined in the intercalated cell masses, along with a further immunohistochemical analysis of the dopamine/D1 mismatches. A similar analysis is also made within the extended amygdala. A spatial mismatch in distribution patterns was found between the mu-opioid receptor-1 immunoreactivity and enkephalin IR in the main intercalated island of the amygdala. Discrete cell patches of dopamine D1 receptor and mu-opioid receptor-1 IR were also identified in a distinct region of the extended amygdala, the interstitial nucleus of the posterior limb of the anterior commissure, medial division (IPACM), which displayed sparse tyrosine hydroxylase or enkephalin/beta-endorphin IR nerve terminals. Furthermore, distinct regions of the main intercalated island that showed dopamine/D1 receptor matches (the rostral and rostrolateral parts) were associated with strong dopamine and cyclic AMP regulated phosphoprotein, 32 kDa-IR in several D1 IR neuronal cell bodies and dendrites, whereas this was not the case for the dopamine/D1 mismatch areas (the rostromedial and caudal parts) of the main intercalated island. The lack of correlation between the terminal/receptor distribution patterns suggests a role for volume transmission for mu-opioid receptor- and dopamine D1 receptor-mediated transmission in distinct regions of the amygdala and extended amygdala. This may have implications for amygdaloid function, where slow long lasting responses may develop as a result of volume transmission operating in opioid peptide and dopaminergic communication.     

Co-localization of tyrosine hydroxylase and GABA immunoreactivities in human cortical neurons

Samples of human cerebral cortex were stained immunocytochemically for tyrosine hydroxylase (TH) and gamma-aminobutyric acid (GABA). TH-positive neurons were in small number and predominated in the deep infragranular layers V-VI contrasting with numerous GABA-positive neurons scattered in all layers. Co-localization of TH- and GABA-like immunoreactivities in a single cell was studied by the double immunolabeling technique with the elution-restaining procedure. Only 50% of the TH-positive neurons also expressed GABA-like immunoreactivities. The two markers were detectable in the somata and not in the processes of the cells. The double-labeled cells were mainly fusiform and medium-sized and were observed in layer VI. These observations suggest that the TH-positive cells form a mixed neuronal population, only a part of which corresponds to the GABAergic class of intrinsic interneurons.     

Distribution of tyrosine hydroxylase and dopamine immunoreactivities in the brain of the South African clawed frog Xenopus laevis

Summary The distribution of dopamine (DA) and the biosynthetic enzyme tyrosine hydroxylase (TH) has been studied immunohistochemically in the brain of the adult South African clawed frog, Xenopus laevis. The goals of the present study are, firstly, to provide detailed information on the DA system of the brain of a species which is commonly used in laboratories as an experimental model and, secondly, to enhance our insight into primitive and derived characters of this catecholaminergic system in amphibians. Dopamine-immunoreactive cell bodies are present in the olfactory bulb, the preoptic area, the suprachiasmatic nucleus, the nucleus of the periventricular organ and its accompanying cells, the nucleus of the posterior tubercle, the posterior thalamic nucleus, the midbrain tegmentum, around the solitary tract, in the ependymal layer along the midline of the caudal rhombencephalon, and along the central canal of the spinal cord. In contrast to the DA antiserum, the TH antiserum fails to stain the liquor-contacting cells in the periventricular organ. On the contrary, the latter antiserum reveals additional immunoreactive cell bodies in the olfactory bulb, the isthmic region and the caudal brainstem. Both antisera yield an almost identical distribution of fibers. Distinct fiber plexuses are observed in the olfactory bulb, the basal forebrain, the hypothalamus and the intermediate lobe of the hypophysis. Features that Xenopus shares with other anurans are the larger number of DAi cells, which are generally smaller in size than those observed in urodeles, and the lack of DAi fibers in pallial structures. On the other hand, the paired midbrain DA cell group and the innervation of the tectum of Xenopus resemble those found in the newt rather than those in frogs. Despite the existence of these species differences, the brain of Xenopus offers an excellent model for studying general aspects of neurotransmitter interactions and the development of catecholamine systems in this class of vertebrates.     

Inactivation of tyrosine hydroxylase in rat striatum by 1-methyl-4-phenylpyridinium ion (MPP+)

We report that 1-methyl-4-phenylpyridinium ion (MPP+), the active metabolite of 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP), inactivated tyrosine hydroxylase (TH) when MPP+ was directly infused into the striatum. We examined both in vitro TH activity and TH content measured by an enzyme immunoassay in the rat striatum after MPP+ was administered by an in vivo brain microdialysis probe. MPP+ caused the inhibition of TH activity but did not influence TH content in the ipsilateral striatum. These results indicate that MPP+ may cause an acute inactivation of TH after continuous exposure at the high concentrations.     

Irradiation inactivation studies of the dopamine D1 receptor and dopamine-stimulated adenylate cyclase in rat striatum

In frozen rat striatal tissue, exposed to 10 MeV electrons from a linear accelerator, the sizes of the dopamine (DA) D1 receptor and the DA-sensitive adenylate cyclase complex were determined using target size analysis. The number of D1 receptors (labelled by [3H]SCH 23390) declined monoexponentially with increasing radiation intensity, yielding a molecular weight (mol.wt.) of 80 kDa. Also the activity of the catalytic unit (C) of the adenylate cyclase (as measured by forskolin stimulation), decreased monoexponentially, however with a mol.wt. of 145 kDa. Both basal, DA- and fluoride (F-)-stimulated activity declined in a concave-downward fashion with a limiting mol.wt. of 134, 138 and 228 kDa, respectively. It was estimated that the basal and DA-stimulated activity originated from an enzyme complex with a mol.wt. of 325 kDa, a value close to the combined size of RGs and C. These data suggest that F- stimulation of the adenylate cyclase, which occurs by a Gs activation, does not cause dissociation of Gs into the alpha s and beta gamma subunits. Further, the DA-regulated adenylate cyclase apparently exists as a complex consisting of RGs and C; the mechanism of hormonal activation is a dissociation of C from this complex.     

帕金森病大鼠未注射侧纹状体和黑质酪氨酸羟化酶的表达

目的:观察不同病程偏侧帕金森病(PD)大鼠未注射侧纹状体和黑质酪氨酸羟化酶(TH)的表达.方法:6-羟基多巴胺(6-OHDA)右侧前脑内侧束立体定位注射1周,阿朴吗啡旋转实验筛选PD模型大鼠,随机分为2、4周和6周模型组,另设正常对照组.行嘴侧纹状体节段和黑质节段连续冠状石蜡切片,采用焦油紫染色定位,以TH抗体免疫组织化学阳性显示多巴胺(DA)能神经元胞体和纤维.结果:正常对照组和2周模型组大鼠左侧纹状体有较强的TH表达,而4周和6周模型组左侧纹状体的TH表达强度较上述两组降低,4周和6周模型组之间无差异.4组大鼠左侧黑质TH表达阳性细胞数无差异.正常对照组、2周和4周模型组左侧黑质有较强的TH表达,3组的表达强度无差异,而6周模型组左侧黑质的TH表达强度较上述3组均降低.结论:6-OHDA单侧注射制备的PD模型大鼠,注射侧的长期病变致未注射侧纹状体和黑质TH的表达减弱.     

Ultrastructural evidence for co-localization of dopamine D2 and micro-opioid receptors in the rat dorsolateral striatum

Previous studies have shown significant changes in dopamine and opioid receptors in the basal ganglia following administration of cocaine. Cocaine administration results in a significant increase in the number of opioid receptors in dopamine-enriched brain regions. The aim of this study was to determine if dopamine D2 receptors (D2r) and micro-opioid receptors (microOr) are localized to the same neurons in the dorsolateral striatum. Immunoperoxidase and immunogold-silver labeling combined with electron microscopy was used to examine the ultrastructural localization of both receptors in the dorsolateral striatum. Approximately half of the microOr-labeled somatodendritic processes showed immunolabeling for the D2r. Similarly, about half of the D2r-labeled dendrites and cell bodies showed immunolabeling for the microOr. In conclusion, our results indicate that individual neurons in the rat dorsolateral striatum may be directly modulated by both dopaminergic and opioid ligands. These data also suggest that the molecular mechanism responsible for the up-regulation of microOrs in the caudate and putamen following cocaine exposure may depend, in part, on the co-existence of D2rs and micro-Ors in these cells.     

Mapping of the colocalization of calretinin and tyrosine hydroxylase in the rat substantia nigra and ventral tegmental area

The distribution of calretinin (CR), a calcium binding protein, was compared with that of tyrosine hydroxylase (TH), the rate-limiting enzyme in the synthesis of dopamine, throughout the rostrocaudal extent of the rat subsantia nigra (SN) and ventral tegmental area (VTA). After mapping the cells using double-labelling immunofluorescence, it was possible to distinguish three distinct cell types: cells immunoreactive for CR only, cells immunoreactive for TH only, and cells in which the two proteins were colocalized (CR+TH). Colocalized cells in rat brain sections comprised approximately 40–55% of the fluorescent labelled cells in the SN compacta, 30–40% in the VTA, and 55–80% in the SN lateralis. Colocalized cells in the SN reticulata were infrequent except in the more caudal sections where a majority of the TH-immunoreactive cells also contained CR. The percentage of CR cells that contained TH was approximately 80% in the SN compacta and averaged 65% in the VTA. Overall, the percentage of TH-immunoreactive cells which also contained CR was approximately 50% in the SN compacta and 45% in the VTA. These data reveal a significant degree of colocalization of CR in dopamine-producing cells of the SN and VTA and suggest the need for studies concerning the fate of these individual cell types following experimental manipulations.     

Regulation of genes involved in dopamine transporter modulation by acute cocaine in rat striatum

It is well established that acute administration of psychostimulants alters dopamine transport. However, the exact mechanism of this modulation is still unknown. In this study we examined the mRNA levels of several proteins involved in the various proposed processes following cocaine administration. The expression levels of several immediate early genes were also studied. This was performed in rat striatum using real-time quantitative PCR. As expected, a marked increase of the immediate early genes Fos, Egr1 and Egr3 was observed. Egr2 was also found up-regulated. Among the different genes studied only Synaptotagmin4 in the SNARE family and Synphilin1 in the synaptic vesicles binding family were modulated by acute cocaine treatment. Interestingly, acute amphetamine treatment did not increase either Synaptotagmin4 and Synphilin1 mRNA levels, although increases in early genes expression were noted.     

Heterogeneous distribution of dopamine D2 receptor mRNA in the rat striatum: a quantitative analysis with in situ hybridization histochemistry.

Dopamine D2 receptor mRNAs have recently been cloned and their gross distribution in the central nervous system described. Quantitative in situ hybridization histochemistry with a cRNA probe complementary to the mRNAs encoding approximately 70% of the third intracellular loop of the rat D2 receptor was performed on sections of rat brain to determine whether differences previously observed in the density of ligand binding sites in subregions of the striatum were related to differences in mRNA levels. Film autoradiographic analysis demonstrated 30% more hybridization signal in the lateral compared to the medial caudate-putamen, a distribution parallel to that of binding of ligands specific for the D2 receptor. Inspection at the cellular level using emulsion autoradiography also indicated a differential distribution of the D2 receptor mRNA. Fewer positively labelled cells, as well as fewer silver grains per cell, were seen in the medial compared to the lateral half of the striatum. This suggests that the gradient seen in autoradiographic studies of the distribution of D2 receptors is related both to regional differences in D2 mRNA levels and to the density of cells expressing the receptor. In addition, the distribution of cells expressing D2 receptor mRNA in the extrastriosomal matrix was compared to that in striosomes identified by the presence of a high density of 3H-naloxone binding sites. Labelled cells were mainly found in the matrix (3H-naloxone binding-poor) but were also seen in striosomes (3H-naloxone binding-rich). The results suggest that differences in levels of D2 binding sites in subregions of the striatum are related to differences in the level of expression of this receptor in intrinsic striatal neurons, suggesting differential regulation of dopamine D2 receptor gene expression in topographically distinct striatal neurons.     

Evidence for the existence of a population of arcuate neurons costoring choline acetyltransferase and tyrosine hydroxylase immunoreactivities in the male rat

By combined immunoperoxidase and immunofluorescence histochemistry we have analyzed the distribution of choline acetyltransferase (ChAT) and tyrosine hydroxylase (TH) immunoreactive (IR) perikarya within the same sections of the mediobasal hypothalamus of the male rat. Evidence was obtained for the existence of perikarya costoring TH and ChAT immunoreactivities in both the dorsomedial and ventrolateral part of the arcuate nucleus and in the adjacent periarcuate nucleus at all rostrocaudal levels. The results strongly implicate interactions between dopamine and acetylcholine as well as acetylcholine and growth hormone releasing factor in dorsomedially and ventrolaterally located TH/ChAT costoring tuberoinfundibular neurons, respectively.     

Co-expression of tyrosine hydroxylase messenger RNA 1 and 2 in human ventral mesencephalon revealed by digoxigenin- and biotin-labelled oligodeoxyribonucleotides

In situ hybridization experiments, using oligodeoxyribonucleotides specific for the two major expressed human tyrosine hydroxylase mRNAs, were performed on human brain sections at the level of the mesencephalon. The specificity of the probes was ascertained by Northern blot experiments carried out with independently in vitro synthesized human tyrosine hydroxylase mRNAs. For in situ hybridization experiments, oligodeoxyribonucleotides were labelled with nucleotides tagged with digoxigenin or biotin molecules. The hybridized oligonucleotides were detected by antibodies coupled with peroxidase and alkaline phosphatase enzymes, which yield, with appropriate substrates, brown and purple products, respectively. The simultaneous detection of the two mRNAs with digoxigeninated and biotinylated probes revealed that these two mRNAs are co-expressed in single cells. The purple product obtained with alkaline phosphatase exhibits a discrete distribution within the dopaminergic cells suggesting these mRNAs are associated with sub-cellular structures. Finally, a heterogeneity in the intensity of the labelling of reactive cells with both probes was visualized as well as the expression of the two mRNA species in neurites.     

Coactivation of D1 and D2 dopamine receptors is required for long-term synaptic depression in the striatum.

Long-term changes of synaptic transmission following brief trains of high-frequency stimulation of excitatory pathways in the brain have attracted attention as a possible correlate of memory. In the cerebellum, concurrent activation of parallel fibers and climbing fibers leads to a long-term depression (LTD) of synaptic transmission, which may be the cellular substrate of motor learning in this structure. We report here for the first time that high-frequency stimulation of corticostriatal glutamatergic fibers in the striatum, another brain structure strongly involved in motor control, also induces LTD of synaptic transmission. Induction of striatal LTD is blocked either by SCH 23390, a D1 dopamine (DA) receptor antagonist or by L-sulpiride, a D2 DA receptor antagonist. The lesion of the nigrostriatal DAergic pathway abolishes LTD. After DA depletion, LTD can be restored by the application of exogenous DA. LTD can also be restored by coadministration of D1 and D2 DA receptor agonists, but not by the application of a single class of DA agonists alone. Our data show that coactivation of D1 and D2 DA receptors is required for LTD in the striatum. D1/D2 receptor cooperation in the induction of LTD may play a crucial role in the behavioural function of DA and in the therapeutic effects of DA agonists in Parkinson's disease.     

Encoding of action history in the rat ventral striatum

In a dynamic environment, animals need to update information about the rewards expected from their alternative actions continually to make optimal choices for its survival. Because the reward resulting from a given action can be substantially delayed, the process of linking a reward to its causative action would be facilitated by memory signals related to the animal's previous actions. Although the ventral striatum has been proposed to play a key role in updating the information about the rewards expected from specific actions, it is not known whether the signals related to previous actions exist in the ventral striatum. In the present study, we recorded neuronal ensemble activity in the rat ventral striatum during a visual discrimination task and investigated whether neuronal activity in the ventral striatum encoded signals related to animal's previous actions. The results show that many neurons modulated their activity according to the animal's goal choice in the previous trial, indicating that memory signals for previous actions are available in the ventral striatum. In contrast, few neurons conveyed signals on impending goal choice of the animal, suggesting the absence of decision signals in the ventral striatum. Memory signals for previous actions might contribute to the process of updating the estimates of rewards expected from alternative actions in the ventral striatum.