Effects of Hypoxia on the Activity of the Dopaminergic Neuron System in the Rat Striatum as Studied by In Vivo Brain Microdialysis

The purpose of the present study is to clarify the effects of hypoxia on the activity of the dopaminergic neurons in the brain and its mechanism of action. For this purpose, the effects of hypoxia on the extracellular levels of 3,4-dihy-droxyphenylethylamine (dopamine) were examined in the rat Striatum using in vivo brain microdialysis in the presence or absence of pretreatment with either tetrodotoxin (a blocker of voltage-dependent sodium channels) or nomifensine (a blocker of dopamine reuptake). Exposure to various degrees of hypoxia (15, 10, and 8% O₂ in N₂) increased dopamine levels in striatal dialysates to 200, 400, and 1,100%, respectively, of the control value. On reoxygenation, dopamine levels in the dialysates rapidly returned to the control level. Reexposure to hypoxia increased the dopamine levels to the same extent as during the first exposure. After addition of tetrodotoxin (40 mUM) to the perfusion fluid or pretreatment with nomifensine (100 mg/kg, i.p.), exposure to hypoxia no longer increased the dopamine levels. These results suggest that although hypoxia induces an increase in the extracellular dopamine levels (hence, an apparent increase in the activity of the dopaminergic neurons), this increase is not the result of an increase in dopamine release itself, but rather the result of inhibition of the dopamine reuptake mechanism.     

Effect of Precursor Loading on the Synthesis Rate and Release of Dopamine and Serotonin in the Striatum: A Microdialysis Study in Conscious Rats

The effects of systemic administration of tyrosine and phenylalanine on the extracellular levels of tyrosine and dopamine were determined by microdialysis in the striatum of awake rats. In addition, the effects of these precursors on in vivo 3,4-dihydroxyphenylalanine (DOPA) formation were determined during continuous infusion of a decarboxylase inhibitor. Both precursors increased the dialysate levels of tyrosine sixfold, but only phenylalanine administration stimulated DOPA formation. However, neither precursor affected the release of dopamine. When the precursor administration was repeated in rats in which the release of dopamine was stimulated by haloperidol pretreatment, again no effect was seen on the release of dopamine. Systemic administration of tryptophan (100 mg/kg, i.p.) during continuous infusion of a decarboxylase inhibitor induced a threefold increase in the formation of 5-hydroxytryptophan and caused an increase in the release of serotonin during infusion of an uptake inhibitor to about 150% of controls. Finally, we investigated whether dietary precursors were able to influence neurotransmitter formation and release. Rats trained to consume their daily food in a period of 2 h were implanted with microdialysis probes. Scheduled eating induced a small increase in the extracellular levels of tyrosine (135% of controls), but the release of dopamine and the formation of 5-hydroxytryptophan during continuous infusion of a decarboxylase inhibitor were not affected.     

Melatonin Effects on Serotonin Synthesis and Metabolism in the Striatum,Nucleus Accumbens,and Dorsal and Median Raphe Nuclei of Rats

This work examined the influence of the pineal gland and its hormone melatonin on the metabolism of serotonin (5-HT) in discrete areas of the forebrain, such as the Striatum and the nucleus accumbens, and the midbrain raphe. The content of 5-HT and its major oxidative metabolite, the 5-hydroxyindoleacetic acid (5-HIAA), as well as the in-vivo tryptophan hydroxylation rate were examined after long-term pinealectomy (one month) and daily melatonin treatment (500 g/kg; twice daily for ten days) in pinealectomized rats. Pinealectomy did not alter 5-HT content in any of these brain areas, but it significantly increased the content of 5-HIAA in Striatum and the 5-HIAA/5-HT ratio in nucleus accumbens. The normal values of these parameters were recuperated after administration of exogenous melatonin, but it also increased the rate of tryptophan hydroxylation in both areas. In addition, melatonin treatment decreased the levels of 5-HIAA in dorsal raphe nucleus. These data suggest that the pineal gland, through the secretion of melatonin, modulates the local metabolism of 5-HT in forebrain areas by acting on the oxidative deamination. Moreover, melatonin injected in pinealectomized rats derives in a more extended effect than pinealectomy and induces a stimulation of 5-HT synthesis in the striatum, probably due to a pharmacological effect. These results point to the striatum as a target area for the interaction between pineal melatonin and the serotonergic function, and suggest a differential effect of the melatonin injected on areas containing serotonergic terminals and cell bodies, which may relevant for the mode of action of melatonin and its behavioral effects.     

Effects of Intranigral Substance P and Neurokinin A Injections on Extracellular Dopamine Levels Measured with Microdialysis in the Striatum and Frontoparietal Cortex of Rats

Extracellular levels of dopamine (DA) and its metabolite, 3,4-dihydroxyphenylacetic acid (DOPAC), in the striatum and frontoparietal (sensorimotor) cortex in halothane-anesthetized rats were analyzed simultaneously using in vivo microdialysis. Basal DA levels, measured from the microdialysis perfusate, were 6.4 +/- 0.8 nM (n = 15) in the striatum and 0.9 +/- 0.1 nM (n = 15) in the frontoparietal cortex. Subcutaneous injections of d-amphetamine (2 mg/kg) increased DA levels 10-fold in the striatum and fivefold in the cortex. Injections of substance P (0.07 nmol/0.2 microliters) into the substantia nigra pars reticulata (SNR) increased DA and DOPAC levels approximately 30% in the ipsilateral striatum and approximately 50% in the ipsilateral frontoparietal cortex. Injections of neurokinin A (0.09 nmol/0.2 microliter) into the SNR increased DA and DOPAC levels approximately 30% in the ipsilateral striatum but did not significantly affect DA levels in the ipsilateral frontoparietal cortex, although DOPAC levels were increased by approximately 50%. It is suggested that striatal and cortical DA release is regulated differently by nigral substance P and neurokinin A terminals.     

Determination of Acetylcholine Release in the Striatum of Anesthetized Rats Using In Vivo Microdialysis and a Radioimmunoassay

A vertical-type in vivo microdialysis probe and a sensitive, specific radioimmunoassay (RIA) were used to study the mechanism of acetylcholine (ACh) release in the striatum of anesthetized rats. Without the use of physostigmine, a cholinesterase inhibitor, our RIA could still detect the amount of ACh present in the perfusate (5.6 +/- 0.6 fmol/min, n = 16). Tetrodotoxin (1 microM) produced a significant decrease in the amount of ACh collected in the perfusate, suggesting that basal ACh determined under the present experimental conditions was related to cholinergic neural activity. Atropine (0.1-1 microM) applied topically via the dialysis probe did not affect the amount of ACh recovered in the perfusate in the absence of physostigmine. Addition of physostigmine (10 microM) to the perfusion fluid produced about a 100-fold increase in the amount of ACh collected. In the presence of physostigmine, topical administration of atropine and pirenzepine (0.01-1 microM) through a dialysis probe produced a further three- to fourfold increase in ACh output, whereas a slight increase was produced by AF-DX 116 at the highest concentration (1 microM). These results indicate that presynaptic modulation of ACh release in the striatum does not occur under basal conditions, and that presynaptic M1 muscarinic receptors are involved in the modulation of ACh release when the ACh concentration is raised under certain conditions.     

Dopaminergic Lesions of the Dorsolateral Striatum in Rats Increase Delay Discounting in an Impulsive Choice Task

Dysregulated dopamine transmission in striatal circuitry is associated with impulsivity. The current study evaluated the influence of dopaminergic inputs to the dorsolateral striatum on impulsive choice, one aspect of impulsive behavior. We implemented an operant task that measures impulsive choice in rats via delay discounting wherein intracranial self-stimulation (ICSS) was used as the positive reinforcer. To do so, rats were anesthetized to allow implanting of a stimulating electrode within the lateral hypothalamus of one hemisphere and bilateral dorsal striatal injections of the dopaminergic toxin, 6-OHDA (lesioned) or its vehicle (sham). Following recovery, rats were trained in a delay discounting task wherein they selected between a small ICSS current presented immediately after lever pressing, and a large ICSS current presented following a 0 to 15s delay upon pressing the alternate lever. Task acquisition and reinforcer discrimination were similar for lesioned and sham rats. All rats exhibited an initial preference for the large reinforcer, and as the delay was increased, preference for the large reinforcer was decreased indicating that the subjective value of the large reinforcer was discounted as a function of delay time. However, this discounting effect was significantly enhanced in lesioned rats for the longer delays. These data reveal a contribution of dopaminergic inputs to the dorsolateral striatum on impulsive choice behavior, and provide new insights into neural substrates underlying discounting behaviors.     

Concentrations of Different Sterols in the Striatum and Serum of 3-Nitropropionic Acid-Treated Wistar and Lewis Rats

In the present study, we examined the long-term effect of oxidative stress induced neurodegeneration on sterol concentrations in striatum and serum of Wistar and Lewis rats. Three weeks after treatment with 3-nitropropionic acid, no differences in 24S-hydroxycholesterol concentrations were observed in striatal homogenates and serum. Ratios of striatal campesterol to cholesterol were higher after 3-nitropropionic acid treatment as compared to controls of both rat strains suggesting an increased passage of this exogenous plant sterol across the blood-brain-barrier. Ratios of lathosterol to cholesterol in serum and striatum were lower in treated rats as compared with controls of both rat strains. Absolute concentrations of serum and striatal cholesterol precursors and plant sterols differed between the controls of both rat strains. It was concluded that the changes observed in sterol concentrations in the striatum and serum indicate that cholesterol homeostasis may be affected during neurodegenerative processes associated with blood-brain-barrier damage.     

Sustained Dopamine Release Induced by Secretoneurin in the Striatum of the Rat: A Microdialysis Study

Abstract: Secretoneurin (SN) is a neuropeptide derived from secretogranin II that is found in brain and endocrine tissues. The aim of the present study was to determine the influence of this novel peptide on dopamine (DA) release from rat striatum using the microdialysis technique. Rat SN (1–30 µmol/L added to the dialysis buffer) enhanced DA outflow of awake rats in a concentration-dependent way without marked effects on the outflow of 3,4-dihydroxyphenylacetic acid or homovanillic acid. The increase in extracellular DA content caused by the peptide was observed throughout the entire period of administration (up to 4 h). Human SN and its 15-amino-acid C-terminal sequence also increased DA outflow, but the effects were smaller than those of rat SN. Two other peptides derived from secretogranin II were without effect on DA efflux. These results establish that SN has a pronounced effect on DA release under in vivo conditions.     

The interstrain differences in the effects of D-amphetamine and raclopride on dorsal striatum dopaminergic system in KM and Wistar rats (microdialysis study)

The levels of dopamine (DA) was determined by intracerebral microdialysis in vivo in KM rats selected for high audiogenic epilepsy, and in Wistar rats selected for nonsusceptibility to loud sound. The basal level of dopamine was 25% higher in the KM rats (P < 0.05). A single amphetamine injection (1 mg/kg body weight, intraperitoneously) caused a significant increase in the DA basal level up to 250-260% in animals of both genotypes. However, in Wistar rats, the level of DA reached maximum as soon as 20 min after amphetamine administration, whereas in KM rats, this happened only after 120 min. After a single injection of the antagonist of D2 and D3 dopamine receptors raclopride (1.2 mg/kg of body weight, intraperitoneously), an increase in the level of DA was similar in amplitude in rats of both genotypes (up to about 210%); however, this occurred 20-30 and 100 min after raclopride administration to Wistar and KM rats, respectively. This evidence suggests that the genetic defect of KM rats, namely, the high level of audiogenic epilepsy, is caused by abnormalities of the neuromediator brain systems and presumably accompanied by the regulatory gene dysfunction.     

Inhibitory Effects of Lysophosphatidylcholine on the Dopaminergic System

Lysophosphatidylcholine (lyso-PTC) is formed by phospholipase A2 (PLA2) from phosphatidylcholine (PTC), that is produced through phosphatidylethanolamine (PTE) methylation. 1-Methyl-4-phenyl-pyridinium (MPP+), a Parkinson's disease (PD) inducing agent, and S-adenosylmethionine (SAM), a biological methyl donor, increase lyso-PTC formation and both induce PD-like changes in animal models. In the current study, we investigated the effect of lyso-PTC on the dopaminergic system to determine the modulating role of lyso-PTC in dopaminergic neurotransmission. The results of these experiments show that lyso-PTC has a remarkable inhibitory effect on dopamine D1 and D2 receptor binding activities in the striatal membrane prepared from Sprague-Dawley rats. Lyso-PTC decreased the Bmax values of both D1 and D2 receptor binding activities. The Kd values for D1 and D2 receptors were not changed, but lyso-PTC also inhibited dopamine transporter and decreased striatal dopamine turnover rate. MPP+ showed similar, but less potent effects. The current studies suggest that lyso-PTC significantly impair the dopaminergic system and might play a role in MPP+ and SAM induced PD-like changes through its inhibitory effects on dopaminergic neurotransmission.     

丙二醛(MDA)对小鼠游泳能力的影响

MDA是脂质过氧化的重要中间产物之一,常用于表征氧化紧张.采用昆明小鼠(KMMice)研究MDA对整体动物的应激反应,考察一次性给药后小鼠的体重变化和游泳能力,结果发现:给药后10min,注射生理盐水(0.85%)能增加小鼠的体重,注射MDA(200mmol/L,0.01mL/g体重)能提高小鼠的游泳能力,二者均达到显著水平(P<0.05).这一结果对于了解氧应激疲劳和解释相关的体力恢复生化机制,具有一定的临床指导意义.     

Characterization of Basal and Morphine-Induced Uridine Release in the Striatum: An In Vivo Microdialysis Study in Mice

Uridine, a pyrimidine nucleoside, has been proposed to be a potential signaling molecule in the central nervous system. The understanding of uridine release in the brain is therefore of fundamental importance. The present study was performed to determine the characteristics of basal and morphine-induced uridine release in the striatum of freely moving mice by using the microdialysis technique. To ascertain whether extracellular uridine was derived from neuronal release, the following criteria were applied: sensitivity to (a) K⁺ depolarization, (b) Na⁺ channel blockade and (c) removal of extracellular Ca²⁺. Uridine levels were not greatly affected by infusion of tetrodotoxin (TTX) and were unaffected by either Ca²⁺-free medium or in the presence of EGTA (a calcium chelator), suggesting that basal extracellular uridine levels were maintained mainly by non-vesicular release mechanisms. In addition, both systemic and local application of morphine increased striatal uridine release. The morphine-induced release was reversed by naloxone pretreatment, but was unaffected by TTX or EGTA infusion. Moreover, co-administration of morphine and nitrobenzylthioinosine (NBTI, an inhibitor of nucleotide transporter) produced increases of uridine levels similar to that produced by NBTI or morphine alone, suggesting a nucleotide transporter mechanism involved. Taken together, these findings suggest that morphine produces a μ-opioid receptor-mediated uridine release via nucleoside transporters in a TTX- and calcium-independent manner.     

Effect of Systemic Administration of N-Methyl-d-Aspartic Acid on Extracellular Taurine Level Measured by Microdialysis in the Hippocampal CA1 Field and Striatum of Rats

Abstract: The extracellular concentrations of amino acids in the hippocampal CA1 field and striatum of conscious freely moving rats were monitored simultaneously by in vivo brain microdialysis using HPLC with electrochemical detection. Under basal conditions, aspartate, glutamate, glutamine, glycine, taurine, and alanine were detected, but γ-aminobutyric acid was undetectable in both regions. In-traperitoneal injection of N -methyl- d -aspartic acid (NMDA; 10 mg/kg) caused a significant increase (three-to fivefold) in the taurine concentration in the dialysate obtained from both the hippocampal CA1 and striatum, whereas other amino acids (aspartate, glutamate, and alanine) did not show significant changes. Local application of NMDA (300 γ) to both regions via the dialysis probes also caused a similar increase (three-to fivefold) in both regions. Under infusion of hypertonic Ringer's solution containing 150 m M sucrose, the effect of NMDA on the level of taurine in both the regional dialysates was not affected. The effect of NMDA was totally reduced by intraperitoneal administration of MK-801 (0.3–1.0 mg/kg), a noncompetitive antagonist of NMDA receptors. Continuous infusion of dl -2-amino-5-phosphonovaleric acid (1.0 mM), a competitive antagonist of NMDA receptors, via the dialysis probes completely inhibited the effect of NMDA. These findings suggest that systemic administration of NMDA is effective as well as local administration into the brain and that NMDA receptors might be involved in the regulation of the extracellular taurine level in the brain without dependence on cell swelling.     

Prenatal Amphetamine Exposure Effects on Dopaminergic Receptors and Transporter in Postnatal Rats

We investigated the influence of prenatal amphetamine exposure (PAE) on dopamine (DA) receptors, and dopamine transporter (DAT) in various striatal and limbic subregions and locomotor activity induced by novel environmental conditions and amphetamine at two postnatal ages, 35 days old (prepubertal) and 60 days old (postpubertal). Experiments were carried out on pregnant female Sprague–Dawley rats, which were daily injected with either d-amphetamine sulfate (1 mg/kg) or saline solution (0.9%) for 11 days, from gestation day 11–21. In PAE rats compared to control we found the following: at pre-pubertal age, an enhancement of DA D1 in the dorsolateral area of the caudate-putamen (CPu), CPu-ventral and shell of the nucleus accumbens (NAcc) with a decrement of the DA D3 receptors in NAcc, olfactory tubercle (OT), and the islands of Calleja (IoC); whereas at postpubertal age, an increase in the levels of DAT in the NAcc and fundus of the CPu, and OT along with a decrease in the expression of DA D2 receptors only in the NAcc shell were found in PAE rats compared to control. In addition, amphetamine induces a marked decrease in locomotor activity at postpubertal age in rats with PAE. These results suggest a differential effect of amphetamines on the DAT mechanism of the nervous system during embryonic development of animals with implications in behavior and drug addictions at adulthood age.     

Comparison of Nitrogen Narcosis and Helium Pressure Effects on Striatal Amino Acids: A Microdialysis Study in Rats

Exposure to nitrogen–oxygen mixture at high pressure induces narcosis, which can be considered as a first step toward general anaesthesia. Narcotic potencies of inert gases are attributed to their lipid solubility. Nitrogen narcosis induces cognitive and motor disturbances that occur from 0.3 MPa in man and from 1 MPa in rats. Neurochemical studies performed in rats up to 3 MPa have shown that nitrogen pressure decreases striatal dopamine release like argon, another inert gas, or nitrous oxide, an anaesthetic gas. Striatal dopamine release is under glutamatergic and other amino acid neurotransmission regulations. The aim of this work was to study the effects of nitrogen at 3 MPa on striatal amino acid levels and to compare to those of 3 MPa of helium which is not narcotic at this pressure, by using a new technique of microdialysis samples extraction under hyperbaric conditions, in freely moving rats. Amino acids were analysed by HPLC coupled to fluorimetric detection in order to appreciate glutamate, aspartate, glutamine and asparagine levels. Nitrogen–oxygen mixture exposure at 3 MPa decreased glutamate, glutamine and asparagine concentrations. In contrast, with helium–oxygen mixture, glutamate and aspartate levels were increased during the compression phase but not during the stay at maximal pressure. Comparison between nitrogen and helium highlighted the narcotic effects of nitrogen at pressure. As a matter of fact, nitrogen induces a reduction in glutamate and in other amino acids that could partly explain the decrease in striatal dopamine level as well as the motor and cognitive disturbances reported in nitrogen narcosis.     

大鼠视网膜变性相关组织形态及功能研究

目的　探讨视网膜变性RCS (RoyalCollegeSurgeon)大鼠的视网膜形态及功能特点。 方法　应用HE染色、免疫组化染色和眼电生理技术 ,对比研究正常和变性两组大鼠视网膜特点。结果　 1 RCS大鼠在 3月龄时 ,视网膜外核层和感光细胞内外节完全消失 ;突触素免疫组化染色显示外丛状层不着色 ;视紫红质免疫组化染色显示原视网膜外层部位有阳性反应 ;胶质纤维酸性蛋白染色显示原视网膜外层部位有强阳性反应。 2 RCS大鼠的闪光视网膜电图 (flashelectronicretinogram ,FERG)a、b波振幅较正常Wistar大鼠明显降低 (P <0 0 1)。结论　在形态和功能上 ,3月龄RCS大鼠视网膜与人类晚期视网膜色素变性极为相似 ,因此可用于视网膜联合移植研究。     

Extracellular Dopamine and Serotonin in the Rat Striatum During Transient Ischaemia of Different Severities: A Microdialysis Study

Abstract: Generalised neurotransmitter overflow into the extracellular space on cerebral ischaemia has been widely reported and implicated in events leading to subsequent neu-ronal death. As little is known about the effect of depth of ischaemia on these changes, we have subjected anaesthetised rats to a sequence of four challenges [high K⁺ stimulus, moderate (penumbral) ischaemia, severe ischaemia, cardiac arrest] and have concurrently monitored both electrophysio-logical parameters and changes in extracellular dopamine, serotonin, and their metabolites in the striatum. Oi'particu-lar relevance to human stroke therapy was penumbral ischaemia, where ionic homeostasis was maintained even though electrical function was lost. All challenges increased extracellular monoamines, although levels were significantly greater when ischaemia was severe enough to produce sustained anoxic depolarisation. Baseline levels were rapidly restored during recovery phases. Acidic monoamine metabolites decreased significantly during each insult, returning to basal levels during reperfusion after moderate ischaemia, and to significantly higher levels after severe ischaemia. Results indicate that sustained anoxic depolarisation may be a critical factor in determining outcome after ischaemia, being associated with significantly greater release of monoamines, and impairment of electrical function recovery.     

Effects of (−)-Sulpiride on Dopamine Release in Striatum of Developing Rats: Degree of Depolarization Influences Responsiveness

Abstract: The purpose of this study was to determine the effects of localized delivery of the D₂ antagonist (−)-sulpiride (via microdialysis) on spontaneous and evoked dopamine release in the neostriatum of urethane-anesthetized rats 5, 10, 15, 21, and 70 days of age. Sulpiride increased spontaneous dopamine release approximately threefold relative to baseline measures, and this effect decreased with maturation. The relationship between sulpiride- and potassium-evoked release was complex; sulpiride increased evoked dopamine outflow at 5, 10, and 15 days of age. At 21 and 70 days of age, however, the effects of sulpiride were inversely related to the degree of stimulation with potassium. Furthermore, the D₂ agonist quinpirole (100 µ M ) reversed the effects of sulpiride (10 µ M ), suggesting receptor mediation. These experiments demonstrate that the maturational decline in the efficacy and potency of D₂ antagonism appears to be related to the degree of stimulation at the nerve terminal.     

Transmitter-Like Basal and K+-Evoked Release of 3,4-Dihydroxyphenylalanine from the Striatum in Conscious Rats Studied by Microdialysis

Using microdialysis and HPLC, characteristics of the release of endogenous 3,4-dihydroxyphenylalanine (DOPA) from striatum in conscious rats were studied in comparison with those of 3,4-dihydroxyphenylethylamine (dopamine; DA). Purified L-aromatic amino acid decarboxylase (AADC) converted a putative peak of DOPA to DA. The retention time of DOPA differed from that of DA and major metabolites of DA and norepinephrine. The DOPA peak of dialysates comigrated with that of authentic DOPA when the pH of the HPLC buffer was modified. The ratio of the basal release of DOPA:DA was 1:2. 3-Hydroxybenzylhydrazine (NSD-1015; 100 mg/kg, i.p.), an AADC inhibitor, markedly increased the basal release of DOPA but produced no effect on DA. The basal release of DOPA was markedly decreased by alpha-methyl-p-tyrosine (200 mg/kg, i.p.), substantially tetrodotoxin (1 microM) sensitive, and Ca2+ (removal plus 12.5 mM Mg2+ addition) dependent. Fifty millimolar K+ released DOPA and this release was also Ca2+ dependent. These characteristics of the basal and evoked release of DOPA were similar to those of DA. The ratio of the evoked release of DOPA:DA was 1:3. These results indicate that DOPA is released under physiological conditions and by K(+)-induced depolarization in a manner similar to that for transmitter DA from striatum in freely moving rats.