人胎蓝斑神经元免疫组织化学研究

为了探讨人蓝斑神经元的胚胎发育特征 ,为蓝斑 -脊髓移植选择适宜胎龄提供形态学根据 ,本研究用免疫组织化学技术系统地观察了人胎蓝斑酪氨酸羟化酶样免疫反应阳性神经元的发育。结果证明 :( 1)蓝斑酪氨酸羟化酶样神经元在胎龄 4个月时已经出现在蓝斑的腹侧部 ;( 2 )蓝斑酪氨酸羟化酶样神经元随胎龄增长逐渐增多 ,以 5个月时增加显著 ;( 3)酪氨酸羟化酶样神经元的密度在胚胎早期升高 ,晚期呈下降趋势 ;( 4)酪氨酸羟化酶样神经元主要分布在蓝斑的背侧部 ,少量散在于腹侧部 ;( 5)酪氨酸羟化酶样神经元开始出现时呈圆形或卵圆形 ,5～ 6个月时呈锥形和梭形 ,7～ 8个月时则以梭形、多角形为主。其胞体逐渐增大 ,胞浆逐渐增多 ,核浆之比由大变小 ,胞突从粗短变为细长平滑。本研究结果提示 ,人胎蓝斑移植以 4个月胎龄者作移植供体较为适宜     

Distribution of norepinephrine transporters in the non-human primate brain

Noradrenergic terminals in the central nervous system are widespread; as such this system plays a role in varying functions such as stress responses, sympathetic regulation, attention, and memory processing, and its dysregulation has been linked to several pathologies. In particular, the norepinephrine transporter is a target in the brain of many therapeutic and abused drugs. We used the selective ligand [(3)H]nisoxetine, therefore, to describe autoradiographically the normal regional distribution of the norepinephrine transporter in the non-human primate central nervous system, thereby providing a baseline to which alterations due to pathological conditions can be compared. The norepinephrine transporter in the monkey brain was distributed heterogeneously, with highest levels occurring in the locus coeruleus complex and raphe nuclei, and moderate binding density in the hypothalamus, midline thalamic nuclei, bed nucleus of the stria terminalis, central nucleus of the amygdala, and brainstem nuclei such as the dorsal motor nucleus of the vagus and nucleus of the solitary tract. Low levels of binding to the norepinephrine transporter were measured in basolateral amygdala and cortical, hippocampal, and striatal regions. The distribution of the norepinephrine transporter in the non-human primate brain was comparable overall to that described in other species, however disparities exist between the rodent and the monkey in brain regions that play a role in such critical processes as memory and learning. The differences in such areas point to the possibility of important functional differences in noradrenergic information processing across species, and suggest the use of caution in applying findings made in the rodent to the human condition.     

Loss of pigmented dopamine-beta-hydroxylase positive cells from locus coeruleus in senile dementia of Alzheimer's type

Serial sections of human brainstem were used to determine the total number of pigmented cells in locus coeruleus and, by immunohistochemical staining using an antiserum directed against human dopamine-beta-hydroxylase (DBH), the number of DBH-positive cells. In 12 brains from elderly control and dementia subjects there wer not significant differences in the total cell populations determined in the same brain by the two techniques. In 6 patients with senile dementia of Alzheimer's type there was a variable loss (average about 60% reduction) in locus coeruleus cells when compared to controls of similar age. The loss of noradrenergic neurones from locus coeruleus was accompanied by an average reduction of similar magnitude in noradrenaline concentration in temporal cortex, with no change or an increase in dopamine content. There was also a significant reduction in the cholinergic marker choline acetyltransferase in cortex samples from the dementia cases.     

Estrogen receptor-alpha immunoreactive neurons in the brainstem and spinal cord of the female rhesus monkey: species-specific characteristics

The distribution pattern of estrogen receptors in the rodent CNS has been reported extensively, but mapping of estrogen receptors in primates is incomplete. In this study we describe the distribution of estrogen receptor alpha immunoreactive (ER-alpha IR) neurons in the brainstem and spinal cord of the rhesus monkey. In the midbrain, ER-alpha IR neurons were located in the periaqueductal gray, especially the caudal ventrolateral part, the adjacent tegmentum, peripeduncular nucleus, and pretectal nucleus. A few ER-alpha IR neurons were found in the lateral parabrachial nucleus, lateral pontine tegmentum, and pontine gray medial to the locus coeruleus. At caudal medullary levels, ER-alpha IR neurons were present in the commissural nucleus of the solitary complex and the caudal spinal trigeminal nucleus. The remaining regions of the brainstem were devoid of ER-alpha IR neurons. Spinal ER-alpha IR neurons were found in laminae I-V, and area X, and were most numerous in lower lumbar and sacral segments. The lateral collateral pathway and dorsal commissural nuclei of the sacral cord and the thoracic intermediolateral cell column also contained ER-alpha IR neurons. Estrogen treatment did not result in any differences in the distribution pattern of ER-alpha IR neurons. The results indicate that ER-alpha IR neurons in the primate brainstem and spinal cord are concentrated mainly in regions involved in sensory and autonomic processing. Compared with rodent species, the regional distribution of ER-alpha IR neurons is less widespread, and ER-alpha IR neurons in regions such as the spinal dorsal horn and caudal spinal trigeminal nucleus appear to be less abundant. These distinctions suggest a modest role of ER-alpha in estrogen-mediated actions on primate brainstem and spinal systems. These differences may contribute to variations in behavioral effects of estrogen between primate and rodent species.     

人胎蓝斑神经元的电镜观察

为了探讨人蓝斑神经元在胚胎发育过程中的超微结构和突触形成特征 ,为蓝斑 -脊髓移植选择适宜胎龄提供形态学资料。用透射电镜观察了 4～ 8个月人胎蓝斑神经元在胚胎发育过程中的变化。结果证明 :胎龄 4个月人胎蓝斑神经元显示不成熟细胞特征 ,胎龄 6个月为发育成熟过程中的细胞特征 ,胎龄 8个月为成熟细胞特征。提示进行人胎蓝斑 -脊髓移植时以 4个月胎龄蓝斑作移植供体较为适宜     

Role of locus coeruleus in foot shock-evoked Fos expression in rat brain

The robust activation of locus coeruleus neurons in response to a variety of stressors, in conjunction with the widespread outputs of the locus coeruleus, suggest that the locus coeruleus may be important in mediating responses to stress. Previous studies in rats have demonstrated that exposure to foot shock elicits Fos expression, a marker of neuronal activation, in the locus coeruleus and other brain sites. In order to evaluate the involvement of the locus coeruleus in foot shock-induced activation of other brain sites, shock-induced Fos expression was examined in the locus coeruleus and other brain areas known to be activated by foot shock, following direct inhibition of the locus coeruleus by local infusion of muscimol, a GABA agonist, prior to foot shock. Control rats received infusions of artificial cerebrospinal fluid into the locus coeruleus or muscimol into areas outside of locus coeruleus. Rats infused with artificial cerebrospinal fluid and then exposed to foot shock had significant increases in Fos expression in several brain areas, including locus coeruleus, nucleus O, several subdivisions of the hypothalamus, subnuclei of amygdala, bed nucleus of the stria terminalis and cingulate cortex. Inhibition of the locus coeruleus prior to foot shock significantly inhibited Fos expression in the locus coeruleus, nucleus O, some subdivisions of the hypothalamus including the magnocellular and medial parvicellular paraventricular hypothalamic nucleus, subnuclei of amygdala, and cingulate cortex. In contrast, inhibition of the locus coeruleus did not affect shock-induced Fos expression in other areas, including certain subdivisions of the hypothalamus and bed nucleus of the stria terminalis. We suggest that foot shock may activate multiple pathways, with activation of certain discrete nuclei requiring input from the locus coeruleus and activation of others occurring independently of locus coeruleus input.     

Characterization of neurochemically specific projections from the locus coeruleus with respect to somatosensory-related barrels

Tactile information from the rodent mystacial vibrissae is relayed through the ascending trigeminal somatosensory system. At each level of this pathway, the whiskers are represented by a unique pattern of dense cell aggregates, which in layer IV of cortex are known as "barrels." Afferent inputs from the dorsal thalamus have been demonstrated repeatedly to correspond rather precisely with this modular organization. However, axonal innervation patterns from other brain regions such as the noradrenergic locus coeruleus are less clear. A previous report has suggested that norepinephrine-containing fibers are concentrated in the center/hollow of the barrel, while other studies have emphasized a more random distribution of monoaminergic projections. To address this issue more directly, individual tissue sections were histochemically processed for cytochrome oxidase in combination with dopamine-beta-hydroxylase, the synthesizing enzyme for norepinephrine, or the neuropeptide galanin. These two neuroactive agents were of particular interest because they colocalize in a majority of locus coeruleus neurons and terminals. Our data indicate that discrete concentrations or local arrays of dopamine-beta-hydroxylase- or galanin-immunoreactive fibers are not apparent within the cores of individual barrels. As such, the data suggest that cortical inputs from the locus coeruleus are not patterned according to cytoarchitectural landmarks or the neurochemical identity of coeruleocortical efferents. While transmitter-specific actions of norepinephrine and/or galanin may not be derived from the laminar/spatial connections of locus coeruleus axons, the possibility remains that the release of these substances may mediate distinctive events through the localization of different receptor subclasses, or the contact of their terminals onto cells with certain morphological characteristics or ultrastructural components.     

Age-dependent changes in axonal branching of single locus coeruleus neurons projecting to two different terminal fields

Age-dependent changes in the axonal branching patterns of single locus coeruleus neurons, which innervate both the frontal cortex and hippocampus dentate gyrus, have been studied in male F344 rats. We used an electrophysiological approach involving antidromic activation to differentiate single from multi-threshold locus coeruleus neurons in each terminal field with age (7-27 mo of age). Most of these neurons have a single threshold in the young rats, whereas in the older brains, the neurons have multi-threshold responses. This implies an increased amount of axonal branching in the older brains. The time course of the increase differs in the two terminal fields, suggesting that the degree of plasticity or age-dependent increase in branching can differ across terminal fields.     

Comparative analysis of the primate X-inactivation center region and reconstruction of the ancestral primate XIST locus

Here we provide a detailed comparative analysis across the candidate X-Inactivation Center (XIC) region and the XIST locus in the genomes of six primates and three mammalian outgroup species. Since lemurs and other strepsirrhine primates represent the sister lineage to all other primates, this analysis focuses on lemurs to reconstruct the ancestral primate sequences and to gain insight into the evolution of this region and the genes within it. This comparative evolutionary genomics approach reveals significant expansion in genomic size across the XIC region in higher primates, with minimal size alterations across the XIST locus itself. Reconstructed primate ancestral XIC sequences show that the most dramatic changes during the past 80 million years occurred between the ancestral primate and the lineage leading to Old World monkeys. In contrast, the XIST locus compared between human and the primate ancestor does not indicate any dramatic changes to exons or XIST-specific repeats; rather, evolution of this locus reflects small incremental changes in overall sequence identity and short repeat insertions. While this comparative analysis reinforces that the region around XIST has been subject to significant genomic change, even among primates, our data suggest that evolution of the XIST sequences themselves represents only small lineage-specific changes across the past 80 million years.     

The locus coeruleus: its possible role in memory consolidation.

Three experiments were done looking at the effects on memory of discrete electrolytic lesions in the locus coeruleus of mice. In Experiment 1 mice received electrolytic lesions of the locus coeruleus immediately following training on the 1 trial inhibitory avoidance step-through task. Retention of this response, measured 48 hr later was normal, suggesting that locus coeruleus lesions per se do not interfere with the performance of this rather simple response. In Experiments 2 and 3 mice were treated exactly as in Experiment 1 except that a transcorneal ECS was administered 40 hr or 7 days after initial training and locus coeruleus lesions. Mice with locus coeruleus lesions were amnesic following a 40 hr delayed ECS treatment when tested either 8 hr or 24 hr after ECS. Mice with locus coeruleus lesions were not amnesic following a 7 day delayed ECS. These data are interpreted to suggest that the locus coeruleus is normally involved in the temporal delineation of the susceptibility period of newly formed memory. A malfunction of the locus coeruleus can result in a profound extension of the normal susceptibility period of newly formed memory to ECS produced amnesia.     

An associativity requirement for locus coeruleus-induced long-term potentiation in the dentate gyrus of the urethane-anesthetized rat

Norepinephrine has been hypothesized to provide a learning and memory signal. Norepinephrine long-term potentiation of perforant path input to the dentate gyrus of the hippocampus provides a model for norepinephrine initiated memory processes. However, in vitro, the pairing of perforant path stimulation and norepinephrine is not required for the occurrence of norepinephrine-dependent long-term potentiation. Since bath application of norepinephrine induces long-term changes in 2nd messenger signalling and differs in a number of ways from physiological norepinephrine release, the present study is an in vivo test of the associative requirement for the pairing of perforant path input with norepinephrine to induce long-term potentiation. Phasic activation of the locus coeruleus is provided by glutamate infusion into the locus coeruleus to initiate transient norepinephrine release in the hippocampus of urethane-anesthetized Sprague-Dawley rats. Perforant path stimulation (0.067 Hz) was given throughout the experiment in the paired condition. In the unpaired condition perforant path stimulation was interrupted 10 min prior to locus coeruleus activation and resumed 10 min after locus coeruleus activation. Locus coeruleus-induced long-term potentiation of both EPSP slope and population spike only occurred in the pairing condition. This result argues that, in vivo, temporal proximity of locus coeruleus-associated norepinephrine release and perforant path stimulation are required to induce long-term plasticity. The associativity requirement for locus coeruleus activation and perforant path stimulation in vivo is consistent with the hypothesis that norepinephrine can initiate circuit changes supporting learning and memory.     

Baseline Spike Activity of Neurons in the Locus Coeruleus of the Rat after Lesions to a Number of Medulla Oblongata Nuclei

Bilateral lesioning of the presublingual nucleus (nucleus praepositus hypoglossi) of the medulla oblongata induced significant changes in the nature of baseline spine activity in the locus coeruleus. After lesioning, the mean frequency of spike activity of locus coeruleus neurons decreased more than two-fold. The numbers of neurons with train-grouped activity and polymodal neurons increased significantly. Exclusion of the solitary tract nucleus led to an increase in the number of neurons with regular activity and some decrease in the mean discharge frequency in locus coeruleus neurons. These data support the suggestion that the presublingual nucleus plays a significant role in the transmission of afferent influences on the activity of locus coeruleus neurons.     

No disease in the brain of a 115-year-old woman

Are there limits to the duration of high quality of life? Are there limits to healthy life for a human brain? We have had the opportunity to evaluate the performance of a 112-113-year-old woman and perform full pathological examination of her body immediately after death at the age of 115. The psychological tests revealed that her general performance was above average of healthy adults of 60-75 years. The pathological observations revealed almost no atherosclerotic changes throughout the body. In the brain almost no beta-amyloid plaques or vascular changes were found and only slight accumulation of hyperphosphorylated tau protein with a Braak-stage 2. Counts of the number of locus coeruleus neurons corresponded with the number of neurons found in the brains of healthy people of 60-80 years old. Our observations indicate that the limits of human cognitive function extends far beyond the range that is currently enjoyed by most individuals and that brain disease, even in supercentanarians, is not inevitable.     

Haloperidol-induced increase in neuropeptide Y immunoreactivity in the locus coeruleus of the rat brain.

The effect of haloperidol, a dopamine (preferably D2) receptor blocking agent on neuropeptide Y immunoreactivity was studied immunohistochemically in neurons of the locus coeruleus and striatum of rat brain. It was found that haloperidol given four times (5 and 2.5 mg/kg, i.p.) induced, after 24 h, a significant increase in the level of neuropeptide Y immunoreactivity in the locus coeruleus but not in the striatum. No changes in neuropeptide Y immunoreactivity in studied structures were observed after alpha-adrenergic receptor blocking agent phenoxybenzamine or serotonin-synthesis inhibitor D,L-p-chlorophenylalanine. The results suggest that the content of neuropeptide Y-immunoreactive material in nerve cell bodies of the locus coeruleus is inhibitorally controlled by monoaminergic (may be dopaminergic D2) receptors.     

GABA in locus coeruleus regulates spontaneous rapid eye movement sleep by acting on GABAA receptors in freely moving rats

The aminergic neurons in the locus coeruleus are known to cease firing during rapid eye movement sleep. Since electrical stimulation of locus coeruleus reduced, while carbachol stimulation increased rapid eye movement sleep and γ-aminobutyric acid (GABA) neurons as well as terminals are present in the locus coeruleus, we hypothesized that GABA may be involved for cessation of locus coeruleus neuronal firing during rapid eye movement sleep. Under surgical anaesthesia male Wistar rats (250–300 g) with bilateral guide cannulae targeting locus coeruleus were prepared for chronic sleep-wakefulness recording. Electroencephalogram (EEG), electrooculogram (EOG), electromyogram (EMG) were recorded in normal, after 250 nl saline and after picrotoxin (250 ng in 250 nl) injection bilaterally into the locus coeruleus. The results showed that mean duration per episode of rapid eye movement sleep was significantly reduced, although its frequency of generation/h was not significantly affected. This study suggests that GABA in locus coeruleus is involved in tonic regulation of rapid eye movement sleep and the action is mediated through GABA_A receptor.     

Evidence that dorsal locus coeruleus neurons can maintain their spinal cord projection following neonatal transection of the dorsal adrenergic bundle in rats

Summary In adult rats, locus coeruleus neurons which extend axons to the spinal cord are found only at mid-rostrocaudal levels of the nucleus, where they are essentially confined to its ventral, wedge-shaped half (Satoh et al. 1980; Westlund et al. 1983; Loughlin et al. 1986). However, during early postnatal development, coeruleospinal cells are found throughout the locus coeruleus (Cabana and Martin 1984; Chen and Stanfield 1987). This developmental restriction of the distribution of coeruleospinal neurons is due to axonal elimination rather than to cell death, since neurons retrogradely labeled through their spinal axons perinatally are still present in the dorsal portion of the locus coeruleus at survival periods beyond the age at which these cells lose their spinal projection (Chen and Stanfield 1987). I now report that if axons ascending from the locus coeruleus are cut by transecting the dorsal adrenergic bundle on the day of birth, a more widespread distribution of coeruleospinal neurons is retained beyond the perinatal period. These results not only indicate that the absence of the normally maintained collateral of a locus coeruleus neuron is sufficient to prevent the elimination of a collateral which would otherwise be lost, but also may imply that during normal postnatal development the presence of the maintained collateral is somehow causally involved in the elimination of the transient collateral.     

Decreased activity of locus coeruleus neurons in the rat after DSP-4 treatment

The activity of noradrenergic neurons of the rat locus coeruleus was investigated at 10 and 50 days after the administration of DSP-4 (N-(2-chloroethyl)-N-ethyl-2-bromobenzyl-amine), a selective noradrenergic neurotoxin. The mean neuronal firing rate in control animals was 2.4 Hz. In contrast, DSP-4 animals had lower rates of 1.2 Hz at 10 days and 1.7 Hz at 50 days. Histological examinations revealed no morphological changes of locus coeruleus cell bodies at either the 10- or 50-day time points. These results suggest that DSP-4 can impair neuronal activity of the locus coeruleus without altering the structural appearance of locus coeruleus perikarya.     

Relationship between locus coeruleus discharge rates and rates of norepinephrine release within neocortex as assessed by in vivo microdialysis.

The relationship between discharge rates of locus coeruleus noradrenergic neurons and rates of norepinephrine release was examined in the anesthetized rat. Neuronal discharge rates of locus coeruleus neurons were altered and quantified using a combined recording-infusion probe. Peri-locus coeruleus infusions of either the cholinergic agonist, bethanechol, or the alpha2-agonist, clonidine, were used to enhance or suppress neuronal discharge activity, respectively. Alterations in concentrations of extracellular norepinephrine within the prefrontal cortex were determined using in vivo microdialysis and high-pressure liquid chromatography with electrochemical detection. A linear relationship between locus coeruleus activity and norepinephrine dialysate concentration was observed between complete suppression of locus coeruleus discharge activity and approximately 300-400% of basal discharge levels (1.58+/-0.29 Hz). Above these levels, increases in locus coeruleus discharge rates were not accompanied by similar increases in dialysate norepinephrine concentrations. In general, neither activation nor suppression of locus coeruleus neuronal discharge rates appeared to alter the relationship between discharge activity and norepinephrine efflux during subsequent epochs. The one exception to this was observed during recovery from relatively high-magnitude locus coeruleus activation. In two out of three cases in which locus coeruleus discharge rates were increased greater than 450%, a recovery of norepinephrine concentrations to basal levels occurred more quickly than the recovery of locus coeruleus neuronal discharge rates to basal levels. Although limited, these latter observations suggest that dysregulation of norepinephrine release may occur following sustained activation of locus coeruleus at the highest rates examined, which may mimic those associated with intense arousal or stress.     

缝隙连接蛋白在PTSD大鼠蓝斑的表达变化

目的观察创伤后应激障碍(PTSD)样行为异常大鼠蓝斑神经元缝隙连接蛋白43(Connexin43,Cx43)表达变化,探讨PTSD的发病机制。方法采用国际认定的SPS方法刺激建立大鼠PTSD模型,取成年健康雄性Wistar大鼠100只,随机分为连续单一刺激(single prolonged stress,SPS)模型1d、4d、7d、14d组和对照组,应用免疫组化、免疫印记和逆转录-聚合酶链式反应(RT-PCR)方法检测PTSD大鼠蓝斑神经元缝隙连接蛋白43的表达变化。结果经SPS刺激后大鼠蓝斑神经元细胞内Cx43蛋白和mRNA于1d开始逐渐升高,4d时表达最多,之后逐渐下降。结论创伤后应激障碍模型大鼠蓝斑Cx43的表达变化,可能与PTSD大鼠蓝斑功能异常相关。     

Locus coeruleus neurons projecting to the forebrain and the spinal cord in the cat

The intranuclear organization of the cat locus coeruleus neurons was investigated anatomo-physiologically. The locus coeruleus neurons project to the forebrain through the dorsal noradrenergic bundle and to the spinal cord. Horseradish peroxidase, a retrograde tracer, was pressure-injected into either the dorsal noradrenergic bundle or the ventrolateral funiculus of the high cervical cord (C1-C2). The cats (n = 12) were killed after a 2- or 3-day survival period. The frontal sections (100 micron) throughout the locus coeruleus were observed by light microscope after carrying out the diaminobenzidine reaction. The labeled locus coeruleus neurons were located predominantly in the rostral locus coeruleus proper and locus coeruleus alpha when horseradish peroxidase was injected into the dorsal noradrenergic bundle, whereas they were predominantly located in the caudal locus coeruleus alpha and subcoeruleus when horseradish peroxidase was injected into the spinal cord. In the electrophysiological experiments, cats (n = 30) were anesthetized with alpha-chloralose and two stimulating electrodes were placed stereotaxically in the dorsal noradrenergic bundle and the ipsilateral ventrolateral funiculus of the high cervical cord (C1-C2), respectively. Monophasic square-wave pulses (2.5 Hz, 100 microsecond duration, 800 microA) were delivered. A recording glass electrode, filled with 2 M NaCl saturated with Fast Green, was placed in the locus coeruleus. Neurons with different conduction velocities, which were evoked by the antidromic stimulation of the dorsal noradrenergic bundle and spinal cord, were verified in the locus coeruleus and the adjacent areas. The slow conductive neurons with a conduction velocity of less than 1 m/s had a slow firing rate (1.6 +/- 0.9/s). They were located predominantly in the rostral locus coeruleus proper and locus coeruleus alpha by the dorsal noradrenergic bundle stimulation. From the anatomical and electrophysiological experimental results, it was concluded that the conduction velocities of the horseradish peroxidase-labeled neurons observed in locus coeruleus proper and locus coeruleus alpha were mostly slow and less than 1 m/s. Most of the slow conductive neurons were considered to be noradrenergic. Neurons evoked antidromically by both the dorsal noradrenergic bundle and spinal cord stimulation were not observed.