Time course of changes in [3H]prazosin binding after catecholaminergic denervation by 6-hydroxydopamine in rat cerebral cortex. Inhibitory effects of ascorbic acid on the alpha 1-adrenoceptor

The binding of [3H]prazosin to alpha 1-adrenoceptors was studied in membranes isolated from the rat cerebral cortex, at different times after the intraventricular injection of 6-hydroxydopamine. In controls, the binding was saturable with the Bmax = 113 pmol/g protein and KD = 0.93 nM, and revealed a single class of sites. After the action of the neurotoxin there was an increase in binding that reached +68% after 7 days and +35% after 14 days. This effect corresponded to an increase in Bmax (190 pmol/g protein) and a decrease in affinity (Kd = 2.1 nM), at 7 days after 6-hydroxydopamine. Carrying on experiments without ascorbic acid in the diluent, the well known inhibitory effect in vitro, that this drug has on the binding of several receptor ligands, was confirmed in vivo. The results obtained suggest that the noradrenergic denervation produced by the neurotoxin is capable of inducing adaptive changes, in alpha 1-post-synaptic adrenoceptors as measured by the binding of [3H]prazosin.     

The effects of PK 11195, a ligand for benzodiazepine binding sites, in animal tests of anxiety and stress

PK 11195 (1-(2-chlorophenyl)-N-methyl-(1-methylpropyl)-3-isoquinolinecarboxami de, a potent ligand for peripheral benzodiazepine binding sites, was unable to reverse the anxiogenic effects of Ro 5-4864 (chlorodiazepam) in the social interaction test or in the punished drinking test. However, at 90 mg/kg PK 11195 also reduced social interaction, indicating an anxiogenic effect. Both PK 11195 (30-120 mg/kg) and Ro 5-4864 (20-60 mg/kg) significantly increased the plasma corticosterone concentrations of rats left in their home cages after injection, and in those placed in novel apparatus. Because both drugs had effects in the same direction and because the doses were far higher than those needed to saturate the peripheral receptor, it is unlikely that these behavioural actions are mediated by peripheral benzodiazepine binding sites. It is suggested that the effect of PK 11195 could even be mediated by the classical CNS benzodiazepine binding sites.     

Antidepressant-like effect of Ro5-4864, a peripheral-type benzodiazepine receptor ligand,in forced swimming test

This study examined the effects of peripheral-type benzodiazepine receptors in the forced swimming test. PK 11195 (1-(2-chloro-phenyl)-N-methyl-N-(1-methylpropyl)-1-isoquinoline carboxamide) and Ro5-4864 (7-chloro-5-(4-chlorophenyl)-1,3-dihydro-1-methyl-2H-1,4-benzodiazepine-2-one) were i.p. injected in mice, according to an acute (1 or 24 h) and a repeated (14 days) schedule. Pretreatment with the agonist, Ro5-4864, significantly reduced immobility time 1 h after treatment but not 24 h after it, whereas the antagonist, PK11195, did not interfere with the test parameters. Nevertheless, PK11195 pretreatment inhibited the Ro5-4864 antidepressant-like effect. Animals repeatedly treated with Ro5-4864 had a similar profile of action with no sign of motor impairment or locomotor activation as evaluated in the rota-rod and open-field tests, respectively. Aminoglutethimide pretreatment, which blocks the early step of steroid synthesis, inhibited the antidepressant-like effect of Ro5-4864. The present findings suggest an antidepressant-like profile for the benzodiazepine, Ro5-4864, that seems to involve steroid synthesis as underlying mechanism.     

Supersensitivity of the noradrenergic system in the spinal cord following intracisternal injection of 6-hydroxydopamine

The intracisternal administration of 6-hydroxydopamine (6-OHDA; 100 micrograms on two consecutive days) resulted in a 90-95% depletion of norepinephrine (NE) in the spinal cord 3-60 days following administration of the drug. At 90 days following injection of 6-OHDA, the depletion was only 60% suggesting the regrowth of NE-containing fibers. Consistent with the depletion of NE, 3-60 days after the injection of 6-OHDA a 50-80% decrease in the uptake NE occurred with only a 25% decrease 90 days after administration of 6-OHDA. Associated with decreases in NE and its uptake was a 100-200% increase in the accumulation of NE-stimulated cyclic AMP in slices of spinal cord tissue from 6-OHDA-treated rats. The increased response was evident at 3-60 days after injection of 6-OHDA decreasing on day 90 to 40% above control. No shift in the EC50 for NE was evident and the specificity of agonists for this response was limited to agents with alpha and/or beta-adrenergic receptor activation properties. Radioligand receptor binding techniques, utilizing [3H]prazosin for alpha1 receptors and [3H]dihydroalprenolol for beta receptors demonstrated that increases in the Bmax of response of alpha1 (30-50%) and beta (15-20%) receptors in spinal cord may mediate the enhanced response of NE-stimulated cyclic AMP following the administration of 6-OHDA. This is indicated by similar time-dependent patterns of onset and termination of increased receptor binding and the accumulation of NE-stimulated cyclic AMP.(ABSTRACT TRUNCATED AT 250 WORDS)     

GABAA receptor subtype selectivity underlying anxiolytic effect of 6-hydroxyflavone

6-Hydroxyflavone (6HF), a naturally occurring flavonoid, was previously reported to bind to type A γ-aminobutyric acid (GABA_A) receptors benzodiazepine (BZ) site with moderate binding affinity. In the present study, we showed that 6HF partially potentiated GABA-induced currents in native GABA_A receptors expressed in cortical neurons via BZ site, as the enhancement was blocked by the antagonist flumazenil. Furthermore, in patch clamp studies, 6HF displayed significant preference for α₂- and α₃-containing subtypes, which were thought to mediate anxiolytic effect, compared to α₁- and α₅-containing subtypes expressed in HEK 293T cells. In mice, 6HF exhibited anxiolytic-like effect in the elevated plus-maze test, unaccompanied at anxiolytic doses by the sedative, cognitive impairing, myorelaxant, motor incoordination and anticonvulsant effects commonly associated with classical BZs when tested in the hole-board, step-through passive avoidance, horizontal wire, rotarod, and pentylenetetrazol (PTZ)-induced seizure tests, respectively. The findings therefore identified 6HF as a promising drug candidate for the treatment of anxiety-like disorders.     

The effect of long-term penfluridol treatment on the sensitivity of the dopamine receptors in the nucleus accumbens and in the corpus striatum

The effect of local application of dopamine to the nucleus accumbens or corpus striatum on locomotor activity was studied in rats 4 days after withdrawal from a 6 weeks term of penfluridol medication. The bilateral application of dopamine into the nucleus accumbens of penfluridol-treated rats produced a very marked increase in coordinated locomotor activity which was 3–5 times higher than that of rats not treated with penfluridol. This effect of dopamine in both penfluridol-treated and control rats was antagonized by intraperitoneally administered haloperidol. The bilateral application of dopamine into the corpus striatum of penfluridol-treated animals produced a marked stereotyped behavioural syndrome in all rats studied, whereas no signs of stereotyped behaviour were observed in any of the rats not treated with penfluridol. The results indicate that long-term treatment of rats with the dopamine receptor blocking agent penfluridol produces an increase in the sensitivity of the dopamine receptors in the nucleus accumbens and corpus striatum and that the nucleus accumbens may play a role in locomotor activity.     

Blockade of non-opioid analgesia in intruder mice by selective neuronal and non-neuronal benzodiazepine recognition site ligands

In male mice, the biologically significant experience of social defeat is associated with an acute non-opioid form of analgesia. Recent studies have shown that this reaction is sensitive to certain benzodiazepine receptor ligands but is unaffected by others. The present experiments were designed to assess the possibility that activity at “non-neuronal” benzodiazepine binding sites might account for this unusual pharmacological profile. Our results show that defeat analgesia was blocked by clonazepam (0.06–3 mg/kg), Ro05-4864 (2.5–20 mg/kg), Ro05-5115 (20 mg/kg), PK11195 (5–20 mg/kg) and PK14067 (10–20 mg/kg). Furthermore, when given in combination, subthreshold doses of PK11195 (2.5 mg/kg) and clonazepam (0.03 mg/kg) totally prevented defeat analgesia. All of these effects were observed in the absence of intrinsic activity on basal nociception. Together with earlier findings, current data imply that inhibition of defeat analgesia by ligands for neuronal and/or non-neuronal benzodiazepine recognition sites is most probablyunrelated to their activity at these sites. Alternative explanations for the overall patterns of results are considered.     

褪黑素对不同月龄小鼠免疫反应的影响及其机制

目的:研究褪黑素对不同月龄小鼠免疫反应的影响及其作用机制。方法:淋巴细胞增殖采用MTT法;IL-2活性检测 采用激活小鼠脾淋巴细胞增殖法;cAMP测定用竞争蛋白结合法;甲硫氨酸脑啡肽的测定采用放免法。结果:6月龄和11月龄BALB/c小鼠胸腺淋巴细胞增殖能力和IL-2的产生均降低,褪黑素(5mg/kg或30mg/kg,ig×7d)对上述指标有不同程度的改善作用,在体外,11月龄小鼠胸腺淋巴细胞增殖反应明显降低,而其产生的cAMP则明显升高,褪黑素(0.1nmol/L和1μmol/L)对此有反向调节作用,弗司可林(10μmol/L)(腺苷酸环化酶选择性激活剂)能够增强2月龄和11月龄BALB/c小鼠胸腺淋巴细胞内的cAMP水平,褪黑素可部分拮抗此作用,褪黑素的这一作用可被百日咳毒素(1mg/L)完全取消,同时发现褪黑素(1μmol/L和0.1nmol/L)能够明显提高2月龄和11月龄BALB/c小鼠胸腺淋巴细胞的甲硫氨酸脑啡肽含量,这一作用能被硝苯地平(1μmol/L)所阻断,提示褪黑素促进淋巴细胞甲硫氨酸脑啡肽释放可能由Ca~(2+)通道介导。结论:褪黑素对不同月龄小鼠具有免疫调节作用;G蛋白偶联的AC-cAMP信号转导通路和淋巴细胞甲硫氨酸脑啡肽的释放可能是褪黑素发挥免疫调节作用的重要机制之一。     

Bilateral lesions of the striatum induced with 6-hydroxydopamine abolish apomorphine-induced yawning in rats

The subcutaneous injection of 0.1 mg/kg apomorphine induced a syndrome consisting of yawning, chewing and sexual arousal in male rats. Bilateral lesions of the striatum induced with 6-hydroxydopamine abolished the drug-induced yawning, chewing and sexual arousal and produced a 58% depletion of the concentrations of dopamine in the striatum. These data suggest that apomorphine-induced yawning is mediated by presynaptic dopamine receptors (which may be autoreceptors). Furthermore, it appears that dopaminergic innervation of the striatum may play an important role in the production of yawning elicited by small doses of dopamine agonists.     

Role of neurosteroids in the anticonvulsant activity of midazolam

BACKGROUND AND PURPOSE

Midazolam is a short-acting benzodiazepine that is widely used as an i.v. sedative and anticonvulsant. Besides interacting with the benzodiazepine site associated with GABA_A receptors, some benzodiazepines act as agonists of translocator protein (18 kDa) (TSPO) to enhance the synthesis of steroids, including neurosteroids with positive modulatory actions on GABA_A receptors. We sought to determine if neurosteroidogenesis induced by midazolam contributes to its anticonvulsant action.

EXPERIMENTAL APPROACH

Mice were pretreated with neurosteroid synthesis inhibitors and potentiators followed by midazolam or clonazepam, a weak TSPO ligand. Anticonvulsant activity was assessed with the i.v. pentylenetetrazol (PTZ) threshold test.

KEY RESULTS

Midazolam (500–5000 µg·kg⁻¹, i.p.) caused a dose-dependent increase in seizure threshold. Pretreatment with the neurosteroid synthesis inhibitors finasteride, a 5α-reductase inhibitor, and a functional TSPO antagonist PK 11195, reduced the anticonvulsant action of midazolam. The anticonvulsant action of midazolam was enhanced by the neurosteroidogenic drug metyrapone, an 11β-hydroxylase inhibitor. In contrast, the anticonvulsant action of clonazepam (100 µg·kg⁻¹) was reduced by finasteride but not by PK 11195, indicating a possible contribution of neurosteroids unrelated to TSPO.

CONCLUSION AND IMPLICATIONS

Enhanced endogenous neurosteroid synthesis, possibly mediated by an interaction with TSPO, contributed to the anticonvulsant action of midazolam. Enhanced neurosteroidogenesis may also be a factor in the actions of other benzodiazepines, even those that only weakly interact with TSPO.     

Ginsenoside Rg1 protects dopaminergic neurons in a rat model of Parkinson's disease through the IGF-I receptor signalling pathway

Background and purpose:

We have shown that ginsenoside Rg1 is a novel class of potent phytoestrogen and activates insulin-like growth factor-I receptor (IGF-IR) signalling pathway in human breast cancer MCF-7 cells. The present study tested the hypothesis that the neuroprotective actions of Rg1 involved activation of the IGF-IR signalling pathway in a rat model of Parkinson''s disease, induced by 6-hydroxydopamine (6-OHDA).

Experimental approach:

Ovariectomized rats were infused unilaterally with 6-OHDA into the medial forebrain bundle to lesion the nigrostriatal dopamine pathway and treated with Rg1 (1.5 h after 6-OHDA injections) in the absence or presence of the IGF-IR antagonist JB-1 (1 h before Rg1 injections). The rotational behaviour induced by apomorphine and the dopamine content in the striatum were studied. Protein and gene expression of tyrosine hydroxylase, dopamine transporter and Bcl-2 in the substantia nigra were also determined.

Key results:

Rg1 treatment ameliorated the rotational behaviour induced by apomorphine in our model of nigrostriatal injury. This effect was partly blocked by JB-1. 6-OHDA significantly decreased the dopamine content of the striatum and treatment with Rg1 reversed this decrease. Treatment with Rg1 of 6-OHDA-lesioned rats reduced neurotoxicity, as measured by tyrosine hydroxylase, dopamine transporter and Bcl-2 protein and gene level in the substantia nigra. These effects were abolished by JB-1.

Conclusions and implications:

These data provide the first evidence that Rg1 has neuroprotective effects on dopaminergic neurons in the 6-OHDA model of nigrostriatal injury and its actions might involve activation of the IGF-IR signalling pathway.     

Using Ro 15-1788 to investigate the benzodiazepine receptor in vivo: Studies on the anticonvulsant and sedative effect of melatonin and the convulsant effect of the benzodiazepine Ro 05-3663

Both the anticonvulsant and sedative effects of diazepam (5 mg/kg) were reversed by subsequent administration of the suggested specific benzodiazepine antagonist Ro 15-1788. In contrast neither the seizure threshold raising or sedative effect of melatonin (200 mg/kg) was reversed by Ro 15-1788. Ro 15-1788 had no effect on the convulsant action of the benzodiazepine Ro 05-3663. These data therefore argue against the suggestion that melatonin produces its sedative and anticonvulsant effects in vivo by interacting with the benzodiazepine receptor, and also strengthens the suggestion that Ro 05-3663 does not act at this site. The use of Ro 15-1788 in demonstrating whether a drug acts in vivo at the benzodiazepine site to produce a pharmacological response is discussed.     

The effect of drugs on the choline and acetylcholine content of the rat striatum following two methods of sacrifice

Choline and acetylcholine concentrations were determined in the striatum of rats sacrificed either by decapitation or by microwave irradiation. In this structure, as shown by isotopic experiments using intravenously administered [³H]-choline, decapitation caused a 50% loss of acetylcholine which was quantitatively recovered as choline. After microwave irradiation, choline levels were identical in the striatum and in the frontal cortex. The levels of choline found in the two brain structures (23 dnmol/g wet weight) were almost identical to the blood choline concentration (19.4 nmol/mn plasma). Various cholinergic and dopaminergic drugs such as oxotremorine, atropine, haloperidol, reserpine, a-methyl-p-tyrosine and L-DOPA altered the striatal acetylcholine content measured after microwave irradiation.     

促红细胞生成素对慢性心力衰竭小鼠急性失代偿的保护机制

目的:探讨促红细胞生成素(erythropoietin,EPO)对脂多糖(lipopolysaccharides,LPS)诱导的急性失代偿性心力衰竭(acute decompensated heart hailure,ADHF)的保护机制。方法:50只C57Bl/6小鼠随机分为空白(Nor)组、阿霉素(doxorubicin,Dox)组、阿霉素感染(Dox+LPS,D&L)组、阿霉素治疗(Dox+EPO,D&E)组和阿霉素感染治疗组(Dox+LPS+EPO,DLE),每组10只。前5周,除Nor组外,其他组腹腔注射Dox(每次5 mg.kg-1,1次.周-1,共5周),建立CHF小鼠模型。第7周时,D&L和DLE组腹腔注射单剂LPS 5 mg.kg-1,D&E和DLE组腹腔注射单剂EPO5 000 iu.kg-1。观察1周后,第8周行心脏超声检查、血清中白介素-6(interleukin-6,IL-6)和脑钠肽(brain natriuretic peptide,BNP)含量测定、HWI、心肌组织的免疫组织化学分析P-STAT3,Bcl-2和Bax蛋白的表达。结果:与D&L组相比,DLE组小鼠给予EPO处理后...     

Activation of peripheral mitochondrial benzodiazepine receptors in the hippocampus stimulates allopregnanolone synthesis and produces anxiolytic-like effects in the rat

RATIONALE AND OBJECTIVES: Stimulation of the mitochondrial benzodiazepine receptor (MBR) in the brain activates the synthesis of neurosteroids that can act as positive modulators of the GABA(A) receptor complex. Allopregnanolone is a potent anxiolytic, anticonvulsant, sedative and hypnotic GABAergic neurosteroid. The anxiolytic-like effects of FGIN 1-27, an MBR agonist, were determined after microinjection into the dorsal hippocampus. METHODS: Behavior in the elevated plus-maze was assessed in adult male rats after bilateral injections of 0, 1.25, 2.5, or 5 microg FGIN 1-27. The behavioral effects of FGIN 1-27 were also determined in animals receiving intrahippocampal co-administration of 20 ng picrotoxin, 5 microg flumazenil, or 200 ng PK 11195. The effects of FGIN 1-27 on behavior in the elevated plus-maze and shock-probe burying test were measured in animals pretreated systemically with 10 mg/kg 4-MA, a 5alpha-reductase inhibitor. Hippocampal and blood plasma levels of allopregnanolone were measured in separate groups of animals pretreated with 4-MA and receiving an intrahippocampal injection of FGIN 1-27. RESULTS: Intrahippocampal injections of FGIN 1-27 produced anxiolytic-like effects in the plus-maze and in the shock-probe burying test. Hippocampal and blood levels of allopregnanolone were also increased by FGIN 1-27. The anxiolytic-like effects of FGIN 1-27 were attenuated by PK 11195 and were blocked by picrotoxin and 4-MA pretreatment, but remained unaffected by flumazenil pretreatment. The neurosteroidogenic effect of FGIN 1-27 was also eliminated by 4-MA. CONCLUSION: Activation of the MBR in the hippocampus leads to the synthesis of allopregnanolone, an anxiolytic neurosteroid that potentiates GABA(A) receptor function.     

Behavioral effects of norepinephrine and dibutyryl 3',5'AMP in centrally sympathectomized rats

Male Wistar rats were injected intraventricularly with two doses of 250 μg of 6-hydroxydopamine (6-OHDA) at a 48 hr interval. Seven days after the second injection, 50 μg of norepinephrine (NE) or 100 μg of dibutyryl cyclic AMP (DCAMP) were injected intraventricularly. There were no differences in gross behavior but an increase in irritability was observed in rats treated with 6-OHDA compared with controls. NE increased locomotor activity and irritability of animals. Chemical sympathectomy intensified locomotor excitation and irratibility caused by NE. DCAMO caused an increase of locomotor activity, irritability and convulsions 30 min after injection. These behavioral phenomena were intensified in animals treated with 6-OHDA. It is suggested that 6-OHDA sensitizes the central nervous system to action of NE and DCAMP.     

Electrophysiological analysis of exogenous and endogenous adenosine actions in the rat and human hippocampus in vitro

Adenosine acts in at least three different ways at A₁ receptors in order to reduce excitability and signal transfer. In the hippocampus, it causes in a steady state outward (potassium) current which is blocked by barium ions and not sensitive to voltage; (ii) an increase in the calcium and cyclic AMP-dependent current 1AHP which is responsible for a long lasting afterhyperpolarization and the accommodation of action potential firing; (iii) a presynaptic reduction of excitatory but not inhibitory transmitter release. The mechanism of this effect is unclear. It is, in contrast to the former, not sensitive to pertussis toxin. When the adenosine elicited potassium currents at the postsynaptic site are blocked, slow inward currents, normally carried by calcium, are unaffected by adenosine. A2 receptor mediated actions have not been found in the hippocampus. While the precise analysis of these effects has been done largely on rats they were all found in human hippocampal slices as well. © 1993 Wiley-Liss, Inc.     

Suriclone and diazepam in the treatment of neurotic anxiety

Suriclone is a new anxiolytic drug belonging to the family of cyclopyrrolones. Although chemically entirely different from the benzodiazepines, it acts as a benzodiazepine agonist with very high affinity for the benzodiazepine receptors. In the present cross-over study, 33 out-patients with a diagnosis of neurotic anxiety were treated with suriclone (mean dose 2 mg/day) and diazepam (25 mg/day) in two 6-week periods. Both drugs had a significant anxiolytic effect, but diazepam appeared to have a better effect within the first 2 weeks of treatment, while no significant difference was seen after treatment for 6 weeks. Suriclone and diazepam had a different side effect profile: suriclone produced mainly dizziness, while diazepam caused sedation. This may reflect the fact that suriclone and benzodiazepines bind to distinct sites or different allosteric conformations of the benzodiazepine receptors.     

The effect of long-term ethanol treatment on the sensitivity of the dopamine receptors in the nucleus accumbens

The effect of local application of dopamine into the nucleus accumbens on locomotor activity was studied in rats during and after withdrawal of long-term ethanol treatment. The bilateral application of dopamine into the nucleus accumbens of both the ethanol and withdrawal rats produced a pronounced increase in coordinated locomotor activity, which was 8–10 times higher than that of untreated water control rats. This effect of dopamine was antagonized by intraperitoneally administered haloperidol indicating a specific effect on dopamine receptors. It is concluded that prolonged ethanol administration may produce an increased sensitivity of the dopamine receptors in the nucleus accumbens and further support the contention that central catecholamine mechanisms are involved in the mediation of the withdrawal syndrome observed after longterm treatment with ethanol.     

The effects of intraperitoneal injection of 6-hydroxydopamine on the turnover and the levels of the brain catecholamines and the levels of plasma corticosterone in rats

1 . The effects of intraperitoneal injection of 6-hydroxydopamine (6-OHDA) on the levels and the turnover of brain catecholamines and the levels of plasma corticosterone were studied in rats. 2. Two weeks after intraperitoneal injection of 6-OHDA (150 mg/kg) a virtually complete disappearance of cardiac noradrenaline was observed. 3. An increment and an accelerated turnover of noradrenaline in the hypothalamus was observed 2 weeks after peripheral administration of 6-OHDA (150 mg/kg). 4. There was no change in the levels and the turnover of noradrenaline in the cortex of the rats so treated. 5. There was no change in the levels and the turnover of dopamine in either the hypothalamus or the cortex of the 6-OHDA-treated rats. 6. An increment and an accelerated turnover of hypothalamic noradrenaline were not associated with any change in plasma corticosterone.