Compensation for behavioral disorders caused by 6-hydroxydopamine administration via the transplantation of embryonic tissue from the locus coeruleus in rats

Neonatal injection of 6-OHDA produces defects of sensory attention and emotional reactions in the rats. Possibility of compensation of these behavioural defects by the grafts of embryonal locus coeruleus (LC) tissue into the neocortex was investigated in male rats of the Wistar strain. Histological analysis revealed spindle-shaped and oval cells typical of LC in the neocortex of all animals with LC grafts. Characteristic green fluorescence of these noradrenergic cells was demonstrated by Falck's method. Normalization of the orienting reaction to sensory (tactile and visual) stimuli, as well as of frustration effects was observed in the animals with LC grafts, but not in the control groups with saline injections or hippocampal grafts. The data show the possibility of stable compensation of the behavioural defects resulting from neonatal injection of 6-OHDA by the grafts of embryonal tissue containing noradrenergic neurones.     

Open-field behavior of rats following bilateral coagulation of the locus coeruleus

A modified open-field test was used in the experiments on rats. It was shown that animals 6 days after bilateral coagulation of locus coeruleus displayed weight changes and some behavioural parameters distinct from those of sham-operated and control animals. The most pronounced differences observed in animals after locus coeruleus coagulation, as compared to the control, were inversion and adaptive responses to a sudden brief stimulus, locomotor dysfunction and absence of response to handling.     

Neonatal 6-hydroxydopamine prevents adaptation to chemical disruption of the pituitary-adrenal system in the rat

Three days after the subcutaneous implant of a dexamethasone pellet, which depletes both corticosterone and ACTH, normal rats showed impaired acquisition of a two-way avoidance task. Rats who had received systemic 6-hydroxydopamine at birth to lesion the forebrain noradrenergic terminals from the locus coeruleus did not show this impairment. After a single injection of metyrapone, which inhibits corticosterone synthesis and increases ACTH release, both intact and norepinephrine (NE)-depleted rats showed impaired avoidance acquisition. After the seventh injection, however, acquisition in normal rats was no longer impaired by the drug while the NE-depleted rats were still deficient. These results indicate that the simple combination of forebrain NE loss with reduced corticosterone levels does not necessarily retard avoidance acquisition. Rather, they suggest that the NE efferents from the locus coeruleus are essential for the brain's adaptation to at least some behavioral consequences of changes in the circulating level of ACTH.     

中枢去甲肾上腺素能系统对大鼠颈动脉窦反射的影响

孤离颈动脉窦,向侧脑室(LCV)注射6-羟多巴胺(6-OHDA,200μg)和电解损毁蓝斑(LC),建立窦内压(ISP)-平均动脉压(MAP)关系曲线,与对照组比较,研究中枢去甲肾上腺素能系统对ISP调节MAP的影响。通过Logistic曲线方程符合所有ISP-MAP曲线,曲线的特征是由两个参数即曲线斜率的反演点坐标和MAP变动范围决定的,MAP变动范围同斜率因子的变化率有关,并受ISP的控制。结果如下:(1)注射6-OHDA后,ISP-MAP曲线斜率变化率显著低于对照组,窦内压控制的平均动脉压变动范围明显缩小,而曲线斜率反演点的横坐标(即ISP的位置)无改变。(2)与对照组相比,电解损毁LC后,引起ISP-MAP曲线相似于注射6-OHDA后的变化,但不如后者显著。结果提示:中枢去甲肾上腺素能神经系统可易化颈动脉窦反射。     

Afferent pathway from locus coeruleus to the nucleus solitarius

After the destruction of the nucleus tractas solitarii, just caudally to the writing pen by means of a stereotaxic instrument, the system of afferent fibres to the nucleus in question was investigated by the methods of Nauta and Fink--Heimer. The fibre terminals were revealed near locus coeruleus. Investigation of the locus coeruleus by Golgi method demonstrated that it usually has neurons of reticular type and transitional ones which resemble by their form the neurons specific for sensory formations. It is possible to conclude that locus coeruleus posesses connections of visceral origin which may play a part in the afferent influence of locus coeruleus on the brain cortex.     

6-hydroxydopamine lesions in anuran amphibians: a new model system for Parkinson's disease?

We investigated the effects of dopamine depletion on acoustically guided behavior of anurans by conducting phonotaxis experiments with female gray treefrogs (Hyla versicolor) before and 90 min after bilateral injections of 3, 6, or 12 microg 6-hydroxydopamine (6-OHDA) into the telencephalic ventricles. In experiments with one loudspeaker playing back a standard artificial mating call, we analyzed the effects of 6-OHDA on phonotactic response time. In choice tests we measured the degree of distraction from the standard call (20 pulses/s) by three different variants with altered pulse-rate (30/s, 40/s, 60/s). Five days after experiments, brains were immunostained for tyrosine hydroxylase. Labeled neurons were counted in the suprachiasmatic nucleus, posterior tuberculum, interpeduncular nucleus, and locus coeruleus, and correlation between neuronal numbers and behavioral scores was tested. Response times increased together with 6-OHDA concentrations, which was mainly due to longer immobile periods before the animals started movement. In choice tests the most irrelevant stimulus (60/s) distracted 6-OHDA injected females from the standard stimulus, while sham injected controls were undistracted. The number of catecholaminergic neurons decreased with increasing 6-OHDA concentration in the suprachiasmatic nucleus, posterior tuberculum, and interpeduncular nucleus. The normalized number of immunoreactive neurons in the posterior tuberculum was positively correlated with phonotaxis scores in the one-speaker test, demonstrating that motor deficits are a function of tubercular cell loss. We conclude that bilateral 6-OHDA lesions in anuran amphibians cause motor (difficulty to start movements) as well as cognitive symptoms (higher distraction by irrelevant stimuli) that have also been described for human Parkinson patients.     

Neuronal and Extraneuronal Effects of Intracisternally Administered 6-Hydroxydopamine on the Developing Rat Brain

Abstract: Intracisternal administration of 100 μg 6-OHDA to newborn rats causes permanent defects, not only of the monoaminergic neuron system, but also of extraneuronal tissue elements. The long noradrenergic fibre tracts are irreversibly destroyed, while the short projections recover and regenerate after a transient period of injury. In the major noradrenergic cell group, the locus coeruleus, most of the cells in the caudal and middle parts degenerate, while a small dorsorostral group survives and forms the source of the regenerating fibres. Dopaminergic and serotonergic fibre tracts are also affected. The 6-OHDA treatment also damages granule and dial cells of the cerebellar cortex as well as the mesenchymal cells of the pial coverings of the cerebellum, leading to primitive foliation, absence of fissuration, and defective migration of granule cells and resulting in a marked reduction of cerebellar size, area, and granule cell number.     

Monoamine Metabolism in the Locus Coeruleus Measured Concurrently with Behavior During Opiate Withdrawal: An In Vivo Microdialysis Study in Freely Moving Rats

Abstract: Using microdialysis, changes in monoamine metabolism were monitored in the locus coeruleus of freely moving rats during opiate withdrawal concomitantly with behavioral symptoms. Rats were infused with morphine (2 mg/kg/h, s.c.) or saline for 5 days and challenged with naltrexone (100 mg/kg, s.c.) on day 6. Following naltrexone challenge, the classic behavioral symptoms of morphine withdrawal were observed in rats treated with morphine but not in saline-infused rats. In morphine-dependent rats, naltrexone induced a marked increase (280%) in dialysate concentrations of 3,4-dihydroxyphenylacetic acid, an index of the functional activity of the noradrenergic neurons in the locus coeruleus. The local concentrations of the serotonin metabolite 5-hydroxyindoleacetic acid were also increased (70%) during morphine withdrawal. Taken together, these results (a) confirm in unanesthetized rats the hypothesis of an activation by opiate withdrawal of noradrenergic neurons in the locus coeruleus and (b) suggest an increase in serotonergic transmission in the same nucleus during morphine withdrawal.     

The dynamic behavioral changes in rats after destruction of the locus coeruleus]

Destruction of locus coeruleus (LC) was conducted in adult rats (male rats of the Wistar line, 250-300 g) electrolytically or by means of 6-hydroxydopamine (6-OHDA). Behaviour of the intact animals sham-operated and of rats with destructed LC was evaluated before and every week after the operation during 5 weeks. Statistically significant alterations in the behaviour were revealed only in the initially low-active rats. These animals were characterized firstly, by low motor and exploratory activity in all tests (open field and automated devices), disturbance of the reactivity to novelty--only in one week after the operation with subsequent compensation of shifts; secondly, by low level of investigatory activity in the open field and absence of habituation to the repetitive tests--from the 21-st day after the operation.     

Immobilization stress-induced lesions of structures of the midbrain reticular formation

Emotional stress in rats resulted in blood-brain barrier increased permeability and brain parenchymal vessel disruptions. Stress induced microvascular damages were mainly observed in midbrain reticular formation. In this article the components of midbrain reticular formation were studied 1, 2, 4 and 6 weeks after immobilization stress. The destructive changes in some neurons, glia cells and myelin fibers were shown up to 6 weeks after immobilization. The signs of the recovery were also observed. It was supposed that the brain parenchymal vessel damages under emotional stress were due to the stress induced locus coeruleus dysfunctions.     

Lack of effect of bilateral locus coeruleus lesion and antidepressant treatment on gamma-aminobutyric acidB receptors in the rat frontal cortex

The aim of the present study was to investigate whether a disturbance of the central noradrenergic (NA) system could cause changes in gamma-aminobutyric acid_B (GABA_B) receptors in the rat frontal cortex. Manipulation of the NA projection to the frontal cortex was achieved by bilateral lesion of the locus coeruleus with 6-hydroxydopamine (6-OHDA) or chronic treatment with the NA reuptake blocker and antidepressant drug, desipramine. Precautions were taken to ensure that the GABA_B receptor assay was performed optimally and was not confounded by the presence of endogenously generated GABA. The results show conclusively that manipulation of the NA projection did not result in any significant change in the number (Bmax) or affinity (Kd) of GABA_B receptors in the frontal cortex. These results do not support the hypothesis that hypoactivity of the central NA system can lead to changes in cortical GABA_B receptors and that antidepressant drugs act by increasing GABA_B receptor binding in the frontal cortex.     

Autoradiographic Localization of Angiotensin II Receptor Binding Sites on Noradrenergic Neurons of the Locus Coeruleus of the Rat

The locus coeruleus (LC) of the rat was lesioned by microinjection of selective neurotoxins into the brainstem. 6-Hydroxydopamine (6-OHDA), 3 micrograms/microliter, given unilaterally at two sites 0.6 mm apart on the rostro-caudal axis of the LC, was used to lesion catecholamine-containing neuronal elements. Ibotenic acid, 2.5 micrograms/0.5 microliters, administered similarly was used to lesion nerve cell bodies. Two weeks after administration of the neurotoxin, lesion efficacy was determined based on the norepinephrine content of the cerebral cortex ipsi- and contralateral to the lesion. 6-OHDA lesions of the LC caused a 46% reduction in ipsilateral cortical norepinephrine and a 60% reduction in specific 125I-[Sar1, Ile8]-angiotensin II (125I-SIAII) binding in the LC. Ibotenic acid lesions of the LC caused a 73% reduction in ipsilateral cortical norepinephrine and a 81% reduction in specific 125I-SIAII binding in the LC. These results indicate that AII receptor binding sites in the LC are localized on noradrenergic nerve cell bodies or their dendritic and axonal ramifications within the LC.     

The behavioral changes and the EEG of the rat brain induced by hyperactivation of the amygdaloid body under noradrenergic deficiency]

The brain NA deficit was produced by bilateral injection of 6-OHDA into the locus coeruleus. Three weeks later amygdala hyperactivation was initiated by local penicillin injection (1% solution, 0.5 mcl). Saline in the same volume was used in control groups in both cases. It was shown that the decrease in NA level facilitated the development of epileptiform activity in rat brain and appearance of immobility-related high-voltage spindles during waking. The amygdala hyperactivation after NA deprivation resulted in a decrease in exploratory activity and disruption of the reaction to novelty. The delta component of the EEG power spectrum increased. The alterations appeared 1-2 weeks after the experimental procedure and became more pronounced towards the end of the third week.     

大鼠PTSD后蓝斑核 β-catenin的表达下降伴随细胞凋亡

目的 探讨创伤后应激障碍( PTSD) 大鼠蓝斑神经元β-catenin(β-连环蛋白)的表达变化.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1 d、4 d、7 d、14 d 组和对照组,应用免疫组化、免疫印迹方法检测PTSD 大鼠蓝斑神经元β-catenin的表达变化;透射电镜观察PTSD大鼠蓝斑神经元的超微结构变化.结果 经SPS 刺激后大鼠蓝斑神经元细胞内β-catenin于1d开始逐渐减少,14d表达最少;蓝斑神经元出现细胞凋亡改变.结论 蓝斑神经元细胞凋亡可能是导致PTSD 患者蓝斑功能失调的重要原因之一.     

Peripheral and central mechanisms of the pressor response elicited by stimulation of the locus coeruleus in the rat

Electrical stimulation of the pontine nucleus locus coeruleus (LC) caused an increase of the arterial blood pressure in anesthetized rats, and elevated plasma noradrenaline (NA) and adrenaline (A) levels. The stimulation-induced pressor response was characteristically biphasic and consisted of a sharp rise in arterial pressure at the onset of the stimulation, followed by a second elevation at the end of the stimulus. Bilateral adrenalectomy or adrenal demedullation completely blocked the secondary phase of the pressor response elicited by stimulation, but did not affect the primary phase. The latter was specifically eliminated by the destruction of the peripheral sympathetic vasomotor axons with intravenous 6-hydroxydopamine (6-OHDA). The active sites eliciting the secondary adrenomedullary pressor component appeared to be restricted to the nucleus LC, whereas the primary sympathetic vasomotor response could be elicited from sites in and around the nucleus. After brain transection at the midbrain level, stimulation of LC failed to evoke the adrenomedullary pressor response, while the sympathetic vasomotor component was not affected. Similarly, destruction of brain NA neurons by intraventricular administration of 6-OHDA did not change the sympathetic vasomotor response, but virtually abolished the adrenal response. The results demonstrate that the pressor response to stimulation of LC in the rat is due to both increased sympathetic vasomotor activity and CA released from the adrenal medulla. The study also provides evidence suggesting that the noradrenergic LC cell group play an important role in the activation of the adrenal medulla, but is not essential for the activation of the sympathetic vasoconstrictor fiber system.     

Morphologic characteristics of locus coeruleus neurons after selective chemical damage to the catecholaminergic system of the brain

The initial part of the noradrenergic cerebral system--the locus caeruleus neurons--has been studied light and electron microscopically at various time after injection of 6- hydrodopamine (6-OHDA), a substance possessing a selective neurotoxic effect on the catecholaminergic mediatory system. Intracisternal injection of 6-OHDA (300 mcg) produces a number of reactive rearrangements in the neurons and large dendrites. Nevertheless, the death of the neural cells is not observed even by the 36th day.     

CYCLIC AMP IN THE RAT CEREBRAL CORTEX AFTER ACTIVATION OF NORADRENALINE NEURONS OF THE LOCUS COERULEUS

The levels of cyclic AMP in the rat brain were studied in vivo following destruction or stimulation of the noradrenergic pathway originating in the locus coeruleus. After chronic lesion of the locus coeruleus no alterations in cyclic AMP content were found. Electrical stimulation of the locus coeruleus produced an elevation of cyclic AMP in the cerebral cortex of chloral hydrate anaesthetized rats of 30%. Maximal increases were found after 15–60 s stimulation at a frequency of 30–100 Hz. This maximal response was slightly inhibited by phenoxybenzamine, an α-adrenergic blocking agent, and by the β-blocker propranolol. When the α and β blockers were administered together a highly significant decrease in cyclic AMP response was observed. Pretreatment of the rats with reserpinc +α methyl-p-tyrosine prevented the cyclic AMP response. In addition to the effect in the cerebral cortex, cyclic AMP-levels were also enhanced in the hippocampus, in the striatum and in the hypothalamus. These results suggest that the locus coeruleus regulates a small fraction of cerebral cyclic AMP levels, by both α- and β-adrenergic receptors.     

Locus coeruleus fibre growth in oculo induced by trigeminotomy

Locus coeruleus from fetal donors was homologously grafted to the anterior eye chambers of adult rats whose eyes were sympathetically denervated. After intraocular maturation, outgrowth of noradrenaline-containing fibres from the locus coeruleus neurons on the host iris was studied by Falck--Hillarp fluorescence histochemistry.In control animals locus coeruleus grafts produce a halo of noradrenaline-containing nerve fibres around the graft, covering approximately one third of the surface of the host iris. Sensory denervation of host eyes carrying maturated locus coeruleus grafts was produced by intracranial lesions of the trigeminal nerve. Such lesions induced a rapid growth response in the grafted locus coeruleus neurons, leading within three weeks to complete innervation of the host iris. It was concluded that removal of non-sympathetic, non-parasympathetic nerve fibres on the host iris elicits a strong fibre-growth response in the grafted locus coeruleus.     

A selective 5-HT1 agonist,RU 24969, increases locus coeruleus catechol metabolic and neuronal activity

Locus coeruleus activity was monitored by either in vivo electrochemistry, post-mortem HPLC, or single unit activity, after systemic administration of RU 24969, a potent serotonin-1 agonist. Whatever the methodology, activation of the locus coeruleus appeared after RU 24969 injection. Catechol oxidation current, assessed by in vivo differential pulse voltammetry and single unit activity in the locus coeruleus showed simultaneous increases after RU 24969. The increase in catechol oxidation current after RU 24969 was dose dependent (ED₅₀ = 1.4 mg kg⁻¹ of i.p. RU 24969). This increased activity was also observed on microdissected locus coeruleus as shown by the increased levels of dopamine and 3,4-dihydroxyphenylacetic acid measured with high performance liquid chromatography. Furthermore, RU 24969 treatment decreased 5-hydroxyindolacetic acid/serotonin ratio in the same microdissected locus coeruleus. This increased locus coeruleus catechol metabolic activity was suppressed by making lesions in the serotonergic systems with 5,7-dihydroxytryptamine. By contrast, neither 8-OHDPAT nor methysergide produced significant changes in the catechol oxidation current recorded in the locus coeruleus.     

Characterization of a developmentally transient adrenergic depolarizing response in cultured locus coeruleus neurons

The effects of iontophoretically applied noradrenaline have been tested on intracellularly recorded locus coeruleus neurons grown in explant cultures from neonatal mice. In addition to hyperpolarizing responses mediated by alpha 2-adrenergic receptors, as observed in locus coeruleus neurons in vivo and in brain slices from adult animals, alpha 1-mediated depolarizations were observed to succeed the initial hyperpolarizations in some cultures. It was shown that the depolarizing responses were only present in younger cultures, i.e., less than 26 days in vitro. In cultures less than 20 days old, all cells displayed the biphasic hyperpolarizing-depolarizing responses. Both components of the response appear to be direct, since they were present when synaptic transmission was blocked by including tetrodotoxin or by altering divalent cations in the perfusate. The depolarizing responses were frequently reduced in solutions with altered divalent cation content, and this might reflect a calcium dependency of this response. The hyperpolarizing and depolarizing components of the responses to noradrenaline were progressively blocked by increasing concentrations of the selective antagonists yohimbine and prazosin, respectively, in the dose ranges of 100 mM - 1 microM (yohimbine) and 20-200 nM (prazosin). Recent results from electrophysiological studies of locus coeruleus neurons in brain slices suggest that similar changes occur in the animal as well as in culture. It is possible that the transient depolarizing responses reflect a developmentally important enhanced responsiveness of locus coeruleus neurons during the early postnatal period.