Neuropeptide Y in brains of the Flinders Sensitive Line rat, a model of depression. Effects of electroconvulsive stimuli and d-amphetamine on peptide concentrations and locomotion

Neuropeptide Y (NPY), has been implicated in the pathophysiology of depression and the mechanisms of action of electroconvulsive treatment (ECT). In this series of experiments, we explored whether there are differences between Flinders Sensitive Line (FSL) rats, an animal model of depression, and controls, Flinders Resistant Line (FRL) in (1) baseline brain NPY-LI concentrations, (2) effects of ECS on locomotion and brain neuropeptides, (3) amphetamine effects on behavior, and (4) effects of ECS pretreatment on subsequent effects of amphetamine on behavior. Both strains were divided into two groups, receiving eight ECS or ShamECS. Twenty-four hours after the last session, animals were habituated in activity boxes for 45 min before given d-amphetamine (1.5 mg.kg(-1), subcutaneously) or vehicle. Locomotor activity was then recorded for an additional 45 min. Twenty-four hours later, rats were sacrificed by microwave irradiation, the brains dissected into frontal cortex, occipital cortex, hippocampus, hypothalamus and striatum, and the neuropeptides extracted and measured by radioimmunoassay. No differences between FSL and FRL rats in baseline locomotor activity were found. FSL compared to FRL animals showed a significantly larger locomotion increase following saline and a significantly smaller increase following amphetamine. ECS pretreatment significantly decreased the saline effects on locomotion in the FSL and the amphetamine effects in the FRL rats. 'Baseline' NPY-like immunoreactivity (LI) concentrations were lower in the hippocampus of the 'depressed' rats. ECS increased NPY-LI in frontal cortex, occipital cortex and hippocampus of both strains. The hippocampal NPY-LI increase was significantly larger in the FSL compared to FRL animals.     

A high-fat diet exacerbates depressive-like behavior in the Flinders Sensitive Line (FSL) rat, a genetic model of depression

Major depressive disorder (MDD) and diabetes mellitus type II (T2DM) are two of the major health challenges of our time. It has been shown that MDD and T2DM are highly co-morbid, and recent work has proposed a bi-directional connection between the diseases. The aim of the current study was to investigate the effect of a high-fat diet (HFD) on behavior and metabolism in a genetic rat model of depression, the Flinders Sensitive and Resistant Line (FSL/FRL) rats. Age and weight matched rats were fed a HFD or control diet for 10 weeks and subjected to behavioral testing and metabolic assessment. We found that HFD exacerbated the depressive-like behavior of the FSL rat in the Forced Swim Test (FST), a depression screening tool, although it did not affect the non-depressed FRL rat despite a higher caloric intake. Moreover, the depressive-like phenotype was associated with reduced anxiety and impairment in novel object recognition memory, while HFD consumption led to diminished object recognition memory as well. In both strains HFD increased insulin levels during an oral glucose tolerance test, although fasting blood glucose levels were only significantly increased by HFD in the FSL rat, suggesting a greater metabolic susceptibility in this rat strain. We conclude that compared with the FRL rat, the FSL rat is more susceptible to developing aberrant behaviors related to depression following metabolic stress induced by HFD. Further studies with a mechanistic focus could potentially lead to a better understanding of a possible pathophysiological link between T2DM and MDD.     

Effects of a new postnatal stress model on monoaminergic neurotransmitters in rat brains

Introduction: Stress and environmental perturbations influence postnatal brain development and may account for the high disability rates of preterm survivors following intensive care treatment. This study aims to investigate the impact of early environmental factors on the monoaminergic neurotransmitter system in the developing rat brain by using an innovative neonatal stress model. Materials and methods: After birth, in the experimental groups newborn rats were separated from their mothers and exposed to different stressful stimuli four times a day on day P0 to P6 for 10 min each. To mimic intensive care treatment, the stress protocol applied environmental factors like bright light, noise, and low temperature alternating with pain and handling stress at day- and night-time in a varying sequence. The non-stressed control mothers and litters were left completely undisturbed until sacrificing on day P7 or P20. Results: Brains of stressed animals revealed significantly higher levels of norepinephrine (NE) and dopamine (DA) as determined by HPLC-ED and electrochemical detection at day P7 as compared to controls. When returned to their mothers’ undisturbed care, juvenile rats at day P20 still showed higher (yet statistically not significant) concentrations of NE and DA in brain. The stressed animals gained less weight with significantly lower body weights at day P7 compared to controls. Their mothers developed various forms of stressed behaviour. Conclusions: A novel animal model for postnatal intensive care stress was established leading to changes in brain monoamine levels of newborn rats, while undisturbed maternal care seems to moderate the stress effects subsequently.     

Region-specific dysregulation of allopregnanolone brain content in the olfactory bulbectomized rat model of depression

Allopregnanolone (ALLO) is one of the most potent positive endogenous allosteric modulators of the type A γ-aminobutyric acid (GABA_A) receptors. While the robust anxiolytic profile of ALLO has been extensively characterized in rodents and its antidepressant-like effect was recently demonstrated in mice, there have been only few reports on alterations of brain ALLO levels in putative animal models of depression and anxiety. Removal of the olfactory bulbs of rats produces one of the most predictive animal models with which to screen for drugs with potential antidepressant activity following repeated treatment. We therefore investigated whether the olfactory bulbectomized (OB) rat model of depression may be associated with alterations of ALLO levels in whole brain tissue and in different brain regions. We determined ALLO levels in whole brain, amygdala, frontal cortex, hippocampus, and whole cerebral cortex of OB or sham-operated rats at 7, 14, or 28 days following bulbectomy or sham surgery. We observed a significant increase of whole brain ALLO content at 7 and 28 days post-surgery in the OB rats. At days 7 and 14 following olfactory bulb removal, ALLO levels were significantly decreased in amygdala and frontal cortex and significantly increased in whole cerebral cortex. In the hippocampus we observed only a tendency for decreased ALLO levels at day 14. Our data indicates a strong region-specific dysregulation of ALLO homeostasis in brains of OB rats which may contribute to the formation of the bulbectomy syndrome via a sustained reduction in physiological GABA-ergic tone in amygdala and frontal cortex.     

Chronic fluoxetine treatment has a larger effect on the density of a serotonin transporter in the Flinders Sensitive Line (FSL) rat model of depression than in normal rats

The 5‐hydroxytryptamine system is thought to play a crucial role in the pathophysiology of depression and represents the target for selective 5‐HT reuptake inhibitors (SSRIs). Flinders Sensitive Line (FSL) and Flinders Resistant Line (FRL) rats were bred from Sprague‐Dawley (SPD) rats to produce strains with increased (FSL) or decreased (FRL) sensitivity to the cholinesterase inhibitor. The FSL rats have been identified as a good model of depression. Many studies in normal rats showed that chronic treatments with SSRIs reduce the densities of SERT. The objective of the present investigation was to assess the influence of chronic fluoxetine treatment on SERT density (Bmax; fmol/mg) in the FSL rat model of depression, relative to that in the FRL rats and SPD rats. FSL, FRL and SPD rats were randomly assigned into groups receiving the vehicle or 10 mg/kg of fluoxetine i.p. for 14 days. Binding was assessed by incubating the brain sections in a buffer containing 20 pM of [¹²⁵I]‐RTI‐55 [[¹²⁵I] (?) ‐2β‐carbomethoxy‐3β‐(4‐iodophenyl)tropane and 200 nM of GBR12935 [1‐(2‐(diphenylmethoxy)ethyl)‐4‐(3‐phenylpropyl)piperazine]. The fluoxetine treatment reduced B_max in all three rat strains when the saline and respective fluoxetine groups were compared (e.g., the FSL‐SAL relative to FSL‐FLX groups). Chronic fluoxetine treatment reduces the densities of SERT in the FSL rats to a larger extent than in the normal SPD control rats. Synapse 64:231–240, 2010. © 2009 Wiley‐Liss, Inc.     

Plasma corticosterone, dexamethasone (DEX) suppression and DEX/CRH tests in a rat model of genetic vulnerability to depression

The hypothalamo-pituitary-adrenal (HPA) axis is hyperactive in major depressive disorder (MDD), and baseline cortisol levels are usually elevated in MDD patients, with alterations of the circadian hormone secretion pattern. The dexamethasone (DEX) suppression test (DST) has been extensively applied to diagnose a dysregulation of the HPA axis in MDD, but it has only a limited sensitivity to, and specificity for, depression. The DEX/CRH test, which combines the DST with a corticotropin-releasing hormone (CRH) challenge, has proved more reliable to show HPA axis dysfunction in MDD. We have applied these two tests to a putative model of vulnerability to depression in rodents, the Roman high-(RHA) and low-(RLA) Avoidance rat lines. As compared to RHA, RLA rats are behaviorally inhibited, they show an exaggerated response of the HPA axis to stress, and are more prone to develop depressive-like features when exposed to chronic stress. Our results show that (a) there were no significant differences in circadian plasma corticosterone (CORT) levels and/or secretion patterns between the two lines; (b) in the DST test, CORT was suppressed to the same extent in RHA and RLA rats; and c) in the DEX/CRH test, areas-under-the-curve (AUCs) and CORT delta (peak minus baseline) responses were significantly larger in RLA rats. One possible interpretation of these data is that an increased response to CRH could be a trait marker (or endophenotype) for depression, whereas alterations of circadian glucocorticoid secretion patterns and non-suppression of the daily glucocorticoid rise by dexamethasone could be state markers, i.e. features that are only present during depressive episodes.     

The antidepressant effects of running and escitalopram are associated with levels of hippocampal NPY and Y1 receptor but not cell proliferation in a rat model of depression

One hypothesis of depression is that it is caused by reduced neuronal plasticity including hippocampal neurogenesis. In this study, we compared the effects of three long‐term antidepressant treatments: escitalopram, voluntary running, and their combination on hippocampal cell proliferation, NPY and the NPY‐Y1 receptor mRNAs, targets assumed to be important for hippocampal plasticity and mood disorders. An animal model of depression, the Flinders Sensitive Line (FSL) rat, was used and female rats were chosen because the majority of the depressed population is females. We investigated if these treatments were correlated to immobility, swimming, and climbing behaviors, which are associated with an overall, serotonergic‐like and noradrenergic‐like antidepressant response, in the Porsolt swim test (PST). Interestingly, while escitalopram, running and their combination increased the number of hippocampal BrdU immunoreactive cells, the antidepressant‐like effect was only detected in the running group and the group with access both to running wheel and escitalopram. Hippocampal NPY mRNA and the NPY‐Y1 receptor mRNA were elevated by running and the combined treatment. Moreover, correlations were detected between NPY mRNA levels and climbing and cell proliferation and NPY‐Y1 receptor mRNA levels and swimming. Our results suggest that increased cell proliferation is not necessarily associated with an antidepressant effect. However, treatments that were associated with an antidepressant‐like effect did regulate hippocampal levels of mRNAs encoding NPY and/or the NPY‐Y1 receptor and support the notion that NPY can stimulate cell proliferation and induce an antidepressant‐like response. © 2009 Wiley‐Liss, Inc.     

Neuropeptide Y enhances the inhibitory effects of clonidine on3H-noradrenaline release in synaptosomes isolated from the medulla oblongata of the male rat

Summary In studies on superfused synaptosomes from the rat medulla oblongata, the inhibitory effects of the2-adrenergic agonist clonidine (0.1M) on potassium (15 mM K⁺) induced³H-noradrenaline (NA) release was potentiated by 20%, when neuropeptide Y was added to the system. The effect of NPY was detectable at low concentrations (1 nM) and was not dose-dependent. Neuropeptide Y alone produced no significant effects on³H-NA release. The results may indicate the existence of a presynaptic NPY receptor on the noradrenaline and/or adrenaline nerve terminals, which may enhance the presynaptic2-adrenoreceptor function to inhibit³H-NA release.     

Amitriptyline and scopolamine in an animal model of depression

Adult male Sprague-Dawley rats were subjected to acute (95 dB white noise) or chronic stress, or their combination. In comparison with unstressed controls, stressed rats were more active upon several measures of open field activity. A history of chronic stress eliminated the acute stress induced activation. Concurrent treatment of chronically stressed rats with amitriptyline or scopolamine, or with a combination of both drugs resulted in selective behavioral improvement (i.e., in motor activity, latency, defecation) for amitriptyline and combined treatment rats, with significant restoration of the normal behavioral response. Scopolamine however was only marginally effective. A higher dose of scopolamine proved effective, but only with a marked disruption of baseline activity. Examination of plasma corticosterone titers indicated that chronic stress induced an elevation of basal levels and that this was reversed by amitriptyline, scopolamine, and combined drug treatment. Thus while behavioral depression and elevated corticosteroids may covary they are not identically mediated.     

Changes of immunoreactive neuropeptide Y, somatostatin and corticotropin-releasing factor (CRF) in the brain of a novel epileptic mutant rat, Ihara's genetically epileptic rat (IGER)

Ihara's genetically epileptic rat (IGER) is a rat mutant with genetically scheduled spontaneous convulsions mimicking human limbic seizures. In the present study, the possible changes of three neuropeptides, neuropeptide Y (NPY), somatostatin (SRIF) and corticotropin-releasing factor (CRF), in the brains of IGER were investigated. Increased contents of immunoreactive (IR) NPY were found only in the hippocampus of 2-month IGERs before developing convulsive seizures, while similar increases of IR-NPY were discovered in the striatum and pyriform and entorhinal cortex as well as hippocampus in 8-month IGERs with repetitive seizures. There were no significant differences in the brain contents of IR-SRIF and IR-CRF between IGERs and the controls at both ages. These findings indicate an enhanced rate of NPY synthesis in this experimental model of epilepsy which may play a critical role in the development of epileptogenesis.     

Correlations among self-esteem, aggression, adverse childhood experiences and depression in inmates of a female juvenile correctional facility in Japan

Aim:  The purpose of this study was to examine the following hypothesis: (i) Self-esteem and aggressiveness, adverse childhood experiences (ACE), and a depressive tendency interact with each other. (ii) The subjects show a strong depressive tendency, and each of the other factors exerts a main effect on, and interacts with, the depressive tendency.
Method:  The subjects were 91 juveniles (all female) admitted to a female juvenile correctional facility between November 2005 and December 2006. They were aged 15–19 years, with a mean age of 17.0 (SD = 1.18) years. Self-esteem scale, aggression scale, ACE questionnaire, and depression scale were conducted.
Results:  Significant main effects (R² = 0.50, P  < 0.001) on the depression score were observed in self-esteem (β = −0.41, P  < 0.001) and aggression (β = 0.21, P  < 0.05). Self-esteem, aggression, ACE, and depression were found to be interrelated.
Conclusion:  Low self-esteem was also shown to exert marked effects on various factors. About half of the inmates of the facility were depressed, and the main effects of self-esteem, aggression, and the ACE score were shown to influence the depression score.     

Peripheral and central administration of neuropeptide Y in a rat middle cerebral artery occlusion stroke model reduces cerebral blood flow and increases infarct volume

Recent studies have shown increased immunoreactivity for neuropeptide Y (NPY) within the perilesional cortex following experimental middle cerebral artery occlusion (MCAO) or focal excitotoxic damage. Downregulation of the NPY Y1 receptor gene using an antisense oligodeoxynucleotide produced a doubling of the infarct volume, implying that NPY may mediate neuroprotection against focal ischemia. The effects of treatment with NPY on infarct volume and hemodynamic parameters were investigated in the present study. Adult male Sprague-Dawley rats were anesthetized with sodium pentobarbital to undergo right-sided endovascular MCAO for 2 h. A single dose of NPY was given via intracarotid injection (10 microg/kg) at the beginning of reperfusion, intracisternal injection (10 or 30 microg/kg) at 30 min of ischemia, or intracerebroventricular (i.c.v.) injection (10 or 70 microg/kg) at 30 min of ischemia. Control groups received the vehicle only via the same route. Body temperature was maintained constant, and hemodynamic parameters were monitored during anesthesia. Laser Doppler flowmetry was used to monitor the regional cerebral blood flow (rCBF) during ischemia and reperfusion in some rats. The rats were decapitated on day 3, and their brains were cut into 2-mm thick coronal slices before reaction with a 2% solution of 2,3,5-triphenyltetrazolium chloride to reveal the infarct. Compared to the respective control groups, NPY treatment via any method of administration increased the relative infarct volume. Suppression of rCBF was observed during reperfusion. These results indicate that peripheral or central administration of NPY impairs reperfusion following experimental MCAO and worsens the outcome of focal cerebral ischemia.     

Reduced GABAB receptor subunit expression and paired-pulse depression in a genetic model of absence seizures

Neocortical networks play a major role in the genesis of generalized spike-and-wave (SW) discharges associated with absence seizures in humans and in animal models, including genetically predisposed WAG/Rij rats. Here, we tested the hypothesis that alterations in GABA(B) receptors contribute to neocortical hyperexcitability in these animals. By using Real-Time PCR we found that mRNA levels for most GABA(B(1)) subunits are diminished in epileptic WAG/Rij neocortex as compared with age-matched non-epileptic controls (NEC), whereas GABA(B(2)) mRNA is unchanged. Next, we investigated the cellular distribution of GABA(B(1)) and GABA(B(2)) subunits by confocal microscopy and discovered that GABA(B(1)) subunits fail to localize in the distal dendrites of WAG/Rij neocortical pyramidal cells. Intracellular recordings from neocortical cells in an in vitro slice preparation demonstrated reduced paired-pulse depression of pharmacologically isolated excitatory and inhibitory responses in epileptic WAG/Rij rats as compared with NECs; moreover, paired-pulse depression in NEC slices was diminished by a GABA(B) receptor antagonist to a greater extent than in WAG/Rij rats further suggesting GABA(B) receptor dysfunction. In conclusion, our data identify changes in GABA(B) receptor subunit expression and distribution along with decreased paired-pulse depression in epileptic WAG/Rij rat neocortex. We propose that these alterations may contribute to neocortical hyperexcitability and thus to SW generation in absence epilepsy.     

Amphetamine and tranylcypromine in an animal model of depression: Pharmacological specificity of the reversal effect

Amphetamine and tranylcypromine are structurally related chemical isomers with pharmacologically distinctive activity profiles. Since they are equimolar and structurally similar they may be used to assess the pharmacological specificity of a proposed animal model of depression. Adult male Sprague-Dawley rats were exposed to a chronic stress regimen or remained undisturbed. They were then acutely stressed with white noise. The monoamine oxidase inhibitor tranylcypromine was effective in restoring otherwise reduced stress elicited open field activity in chronically stressed rats. Amphetamine did not resemble tranylcypromine or other antidepressants, and produced a variety of effects at least some of which indicated a potential increase rather than reduction in depression consequent to chronic administration.     

Neuropeptide Y increases the inhibitory effects of clonidine on potassium evoked3H-noradrenaline but not3H-5-hydroxytryptamine release from synaptosomes of the hypothalamus and the frontoparietal cortex of the male Sprague-Dawley rat

Summary The release of³H-noradrenaline (³H-NA) and of³H-5-hydroxytryptamine (³H-5-HT) evoked by high-K⁺ (15 mM) was studied in synaptosomes isolated from the hypothalamus and the frontoparietal cortex of the male Sprague-Dawley rat using a superfusion apparatus. Based on concentration-response curves obtained by analyzing the full-time course of the inhibitory effects of clonidine on³H-NA and on³H-5-HT release neuropeptide Y (NPY) (1 nM) was shown to significantly increase the ability of clonidine to inhibit³H-NA release in synaptosomes isolated from the hypothalamus and from the frontoparietal cortex. NPY (1 nM) alone had no effect on K⁺-evoked³H-NA release from these regions. In contrast, NPY (1 nM) did not modulate the inhibitory effects of clonidine on³H-5-HT release in the above mentioned regions. These results indicate that NPY can increase the sensitivity of the 2-autoreceptors belonging to hypothalamic NA and/or to adrenaline nerve terminals and to cortical NA nerve terminals, while the 2-heteroreceptors inhibiting³H-HT release in the same brain regions appear not to be regulated by high affinity NPY receptors. Thus, 2-autoreceptors and 2-heteroreceptors appear to be differentially controlled by high affinity NPY receptors at least with regard to regulation of³H-NA and³H-5-HT release, respectively.     

Effects of a selective Y2R antagonist, JNJ-31020028, on nicotine abstinence-related social anxiety-like behavior, neuropeptide Y and corticotropin releasing factor mRNA levels in the novelty-seeking phenotype

An outbred rat model of novelty-seeking phenotype has predictive value for the expression of locomotor sensitization to nicotine. When experimentally naïve rats are exposed to a novel environment, some display high rates of locomotor reactivity (HRs, scores ranking at top 1/3rd of the population), whereas some display low rates (LRs, scores ranking at bottom 1/3rd of the population). Basally, HRs display lower anxiety-like behavior compared to LRs along with higher neuropeptide Y (NPY) mRNA in the amygdala and the hippocampus. Following an intermittent behavioral sensitization to nicotine regimen and 1 wk of abstinence, HRs show increased social anxiety-like behavior in the social interaction test and robust expression of locomotor sensitization to a low dose nicotine challenge. These effects are accompanied by a deficit in NPY mRNA levels in the medial nucleus of the amygdala and the CA3 field of the hippocampus, and increases in Y2R mRNA levels in the CA3 field and corticotropin releasing factor (CRF) mRNA levels in the central nucleus of the amygdala. Systemic and daily injections of a Y2R antagonist, JNJ-31020028, during abstinence fully reverse nicotine-induced social anxiety-like behavior, the expression of locomotor sensitization to nicotine challenge, the deficit in the NPY mRNA levels in the amygdala and the hippocampus, as well as result an increase in Y2R mRNA levels in the hippocampus and the CRF mRNA levels in the amygdala in HRs. These findings implicate central Y2R in neuropeptidergic regulation of social anxiety in a behavioral sensitization to nicotine regimen in the LRHR rats.     

No association between the -399 C > T polymorphism of the neuropeptide Y gene and schizophrenia, unipolar depression or panic disorder in a Danish population

OBJECTIVE: A polymorphism in the promoter region of the NPY gene at position -399 C > T was recently reported to be associated with schizophrenia in a Japanese population and with treatment refractory unipolar depression in a Swedish population. The objective of this study was to investigate potential associations between the polymorphism and three psychiatric disorders in a Danish population. METHOD: We investigated the occurrence of the polymorphism in patients with schizophrenia (n = 291), unipolar depression (n = 256) and panic disorder (n = 142) compared with controls (n = 716). RESULTS: We detected the polymorphism -399 C > T at a frequency of 48% in controls. No significant differences were found between genotype or allele frequencies in controls vs. the patient groups. CONCLUSION: The lack of association between the -399 C > T polymorphism and schizophrenia, unipolar depression or panic disorder, respectively, suggests that the polymorphism is not involved in the etiology of these disorders in the Danish population.     

Anxiety- rather than depression-like behavior is associated with adult neurogenesis in a female mouse model of higher trait anxiety- and comorbid depression-like behavior

Adult neurogenesis has been implicated in affective disorders and the action of antidepressants (ADs) although the functional significance of this association is still unclear. The use of animal models closely mimicking human comorbid affective and anxiety disorders seen in the majority of patients should provide relevant novel information. Here, we used a unique genetic mouse model displaying higher trait anxiety (HAB) and comorbid depression-like behavior. We demonstrate that HABs have a lower rate of hippocampal neurogenesis and impaired functional integration of newly born neurons as compared with their normal anxiety/depression-like behavior (NAB) controls. In HABs, chronic treatment with the AD fluoxetine alleviated their higher depression-like behavior and protected them from relapse for 3 but not 7 weeks after discontinuation of the treatment without affecting neurogenesis. Similar to what has been observed in depressed patients, fluoxetine treatment induced anxiogenic-like effects during the early treatment phase in NABs along with a reduction in neurogenesis. On the other hand, treatment with AD drugs with a particularly strong anxiolytic component, namely the neurokinin-1-receptor-antagonist L-822 429 or tianeptine, increased the reduced rate of neurogenesis in HABs up to NAB levels. In addition, challenge-induced hypoactivation of dentate gyrus (DG) neurons in HABs was normalized by all three drugs. Overall, these data suggest that AD-like effects in a psychopathological mouse model are commonly associated with modulation of DG hypoactivity but not neurogenesis, suggesting normalization of hippocampal hypoactivity as a neurobiological marker indicating successful remission. Finally, rather than to higher depression-related behavior, neurogenesis seems to be linked to pathological anxiety.     

Effects of Rubus parvifolius L. on neuronal apoptosis and expression of apoptosis-related proteins in a rat model of focal cerebral ischemic-reperfusion injury

BACKGROUND:Studies have shown that the Ruhus parvifolius L.(RP)plant extract exhibits protective effects on cerebral ischemia.This effect is reflected in altered ischemic neuronal apoptosis and associated protein expression.OBJECTIVE:To explore the neuroprotective mechanism of RP after cerebral ischemia jniury.DESIGN,TIME AND SETTING:Randomized control experiment of cellular,molecular.and protein levels.The experiment was completed at Chongqing Medical University at the School of Pharmacy Laboratories and Basic Medical Institute from October 2005 to January 2006.MATERIALS:Twenty-four adult,male,Wistar rats,weighing(28±20)g.RP extract,which was a product of ethanol extraction.was provided by the LabOratory of Pharmaceutical Analysis,Chongqing Medical University.RP was dissolved in distilled water to a concentration of 10 mg/mL.All rats were randomly assigned into four groups:5 g/kg RP,10 g/kg RP,model,and sham-surgery,with 6 rats in each group.METHODS:In the 5 and 10 g/kg RP groups,as well as the model group,the middle cerebral artery was occluded(MCAO)for 60 minutes,resulting in focal cerebral ischemia,followed by reperfusion for 24 hours.Rats in the 5 and 10 g/kg RP groups received 5 and 10 g/kg RP,respectively.The RP treatment group received RP intragastrically(once a day)for 3 days.One hour after the last dose,rats were subjected to MCAO.The same surgical procedure was performed in the sham-surgery group,except the suture was introduced into the extemal carotid artery,but not advanced.Rats in the model group were subjected to MCAO.The sham-surgery and model groups received intragastrically administered normal saline once per day for 3 days.One hour after the last dose,the rats were subjected to surgery.MAIN OUTCOME MEASURES:TUNEL labeling and immunohistochemical methods were used to investigate changes in neuronal apoptosis and expression of the apoptosis-related proteins.Bax and Bcl-2.on the ischemic hemisphere,as well as the contralateral hemisphere,after administration of RP or normal saline.RESULTS:All 24 Wistar rats were included in the final analysis.without any loss.Compared with the sham-surgery group,the number of the TUNEL-positive cells in the model group,as well as the 5 and 10 g/kg RP treatment groups,were significantly increased in the ischemic hemisphere(P<0.05).Compared with the sham-surgery group,the number of Bcl-2-positive and Bax-positive cells increased significantly in the model group(P<0.01).The number of Bcl-2-positive cells increased,and the number of Bax-positive cells decreased in the model group,compared to the 5 and 10 g/kg RP treatment groups(P<0.05-0.01).CONCLUSION:RP can prevent neuronal apoptosis following cerebral ischemic-reperfusion of rats.The mechanism underlying the RP-induced neuroprotection in the cerebral ischemia iniury may be related to increased Bcl-2 expression and decreased Bax expression.     

Effects of Rubus parvifolius L. on neuronal apoptosis and expression of apoptosis-related proteins in a rat model of focal cerebral ischemic-reperfusion injury*★

BACKGROUND： Studies have shown that the Rubus parvifolius L. （RP） plant extract exhibits protective effects on cerebral ischemia. This effect is reflected in altered ischemic neuronal apoptosis and associated protein expression.
OBJECTIVE： To explore the neuroprotective mechanism of RP after cerebral ischemia injury.
DESIGN, TIME AND SETTING： Randomized control experiment of cellular, molecular, and protein levels. The experiment was completed at Chongqing Medical University at the School of Pharmacy Laboratories and Basic Medical Institute from October 2005 to January 2006.
MATERIALS： Twenty-four adult, male, Wistar rats, weighing （28 ± 20） g. RP extract, which was a product of ethanol extraction, was provided by the Laboratory of Pharmaceutical Analysis, Chongqing Medical University. RP was dissolved in distilled water to a concentration of 10 mg/mL. All rats were randomly assigned into four groups： 5 g/kg RP, 10 g/kg RP, model, and sham-surgery, with 6 rats in each group.
METHODS： In the 5 and 10 g/kg RP groups, as well as the model group, the middle cerebral artery was occluded （MCAO） for 60 minutes, resulting in focal cerebral ischemia, followed by reperfusion for 24 hours. Rats in the 5 and 10 g/kg RP groups received 5 and 10 g/kg RP, respectively. The RP treatment group received RP intragastrically （once a day） for 3 days. One hour after the last dose, rats were subjected to MCAO. The same surgical procedure was performed in the sham-surgery group, except the suture was introduced into the external carotid artery, but not advanced. Rats in the model group were subjected to MCAO. The sham-surgery and model groups received intragastrically administered normal saline once per day for 3 days. One hour after the last dose, the rats were subjected to surgery.
MAIN OUTCOME MEASURES： TUNEL labeling and immunohistochemical methods were used to investigate changes in neuronal apoptosis and expression of the apoptosis-related proteins, Bax and Bcl-2, on the ischemic hemisphere