GBR12909 stimulates hypothalamo-pituitary-adrenal activity by inhibition of uptake at hypothalamic dopamine neurons

We have previously demonstrated that the inhibition of neurotransmitter uptake at dopamine (DA) terminals stimulates the hypothalamo-pituitary-adrenal axis. In the present study we investigated the role of central DA neuronal systems in the regulation of this axis. Administration of the DA uptake inhibitor GBR12909 (3–30 μg) into the third ventricle dose-dependently elevated serum adrenocorticotropin hormone (ACTH) levels in rats. GBR12909 (10 μg) elevated serum ACTH levels following administration into the paraventricular nucleus of the hypothalamus but not into the lateral ventricle. The administration of 6-OHDA into the third ventricle significantly decreased DA content in the hypothalamus and striatum and significantly attenuated the ACTH response to GBR12909 (10 μg, third ventricle or 10 mg/kg, i.p.). Conversely, 6-OHDA lesions of the medial forebrain bundle, which depleted 99% of DA in the caudate but did not decrease DA content in the hypothalamus, and did not attenuate the ACTH response to i.p. GBR12909. Measurement of GBR12909-induced inhibition of [³H]DA uptake into synaptosomal preparations indicates the presence of GBR12909-sensitive DA transporters in the region of the hypothalamus surrounding the thrid ventricle. The present findings suggest that an endogenous DA neuronal system terminating in the hypothalamus mediates the effects of stimulants on hypothalamo-pituitary-adrenal function and might play a role in the ongoing regulation of hypothalamo-pituitary-adrenal activity.     

Selective lesion of central dopamine or noradrenaline neuron systems in the neonatal rat: motor behavior and monoamine alterations at adult stage

Different parameters of motor behavior (locomotion, rearing and total activity counts) were studied in the adult rat following neonatal intracisternal 6-hydroxydopamine (6-OHDA, 50 micrograms) treatment combined with noradrenaline (NA) uptake blocker (desipramine) or dopamine (DA) uptake blockers (amfolenic acid or GBR 12909) to obtain selective DA or NA lesions respectively. At 61-65 days of age, selective DA-lesioned animals showed an initial decrease in spontaneous motor behavior at test days 1 and/or 2, while at test days 4 and 5 hyperactivity was observed. However, following amfolenic acid or GBR 12909 pretreatment leading to a selective NA lesion, no difference in spontaneous motor behavior was seen on any of the 5 test days. Determination of regional brain levels of NA and DA confirmed the type of lesion predicted from the various pretreatments with selective uptake blockers. These data suggest that changes in motor behavior in the adult rats, following neonatal 6-OHDA treatment, are specifically related to a DA-denervation, whereas an NA lesion does not seem to influence the spontaneous motor behavior. However, following the selective DA lesion, significant increases of serotonin levels in striatum and cerebellum were observed, while following selective NA lesions an increase of cerebellar NA levels was found concomitant with drastic reductions of NA levels in frontal cortex and spinal cord.     

Possible involvement of serotonergic neurons in the reduction of locomotor hyperactivity caused by amphetamine in neonatal rats depleted of brain dopamine

This experiment attempted to determine the mechanism by which amphetamine reduces locomotor hyperactivity in neonatal rats given brain dopamine (DA)-depleting 6-hydroxydopamine (6-OHDA) injections. Brain DA neurons were destroyed selectively in neonatal rats by intraventricular (i.v.t.) injections of 6-OHDA following desmethylimipramine (DMI) pretreatment. Control rats received DMI and i.v.t. injections of the 6-OHDA vehicle solution. Rats given the 6-OHDA treatment displayed 7-fold increases in locomotor activity compared to controls during days 16–55 of life. Throughout this period, amphetamine (1 mg/kg) reduced locomotor hyperactivity in 6-OHDA-treated rats but increased locomotor activity in control rats. The reduction of hyperactivity caused by amphetamine (0.5–4 mg/kg) was dose-related and was not accompanied by stereotyped behavior. Like amphetamine, methylphenidate (4 mg/kg) reduced locomotor hyperactivity in rats given 6-OHDA. The DA antagonist, spiroperidol (50–200 μg/kg) failed to attenuate the hyperactivity-reducing effect of amphetamine in 6-OHDA-treated rats at doses which abolished the stimulant effect of amphetamine in control rats. However, the serotonin antagonist methysergide (0.5–4 mg/kg) produced dose-dependent antagonism of the effect of amphetamine in 6-OHDA-treated rats. Pretreatment with propranolol (5 mg/kg), phentolamine (5 mg/kg), atropine (0.5 mg/kg) or naloxone (10 mg/kg) failed to alter the reduction in locomotor hyperactivity caused by amphetamine. The serotonin releasing agent, fenfluramine (3 mg/kg), and the serotonin agonist, quipazine (0.5–4 mg/kg), both reduced locomotor hyperactivity in 6-OHDA-treated rats while not altering locomotion in control rats. These results confirm previous observations that amphetamine reduces locomotor hyperactivity caused by neonatal 6-OHDA administration and suggest that this effect is mediated by increased serotonergic neurotransmission.     

The amphetamine-chlordiazepoxide mixture, a pharmacological screen for mood stabilizers, does not enhance amphetamine-induced disruption of prepulse inhibition

In rodents, administration of a mixture of the psychostimulant d-amphetamine and the benzodiazepine chlordiazepoxide results in supra-additive hyperlocomotion, a phenomenon used to identify mood stabilizers. In an attempt to determine whether the d-amphetamine/chlordiazepoxide assay could extend to other behaviors that are affected in mania, we evaluated the effects of the mixture on prepulse inhibition. In addition, we combined chlordiazepoxide with the selective dopamine reuptake inhibitor GBR 12909 or the noradrenergic stimulant (−) ephedrine, and tested these alternative mixtures in locomotor activity and prepulse inhibition tests. Chlordiazepoxide (3 mg/kg) robustly potentiated amphetamine-induced hyperactivity, but did not change the amphetamine-induced disruption of prepulse inhibition. This indicates that the d-amphetamine-chlordiazepoxide-induced hyperlocomotion does not extend to other dopamine-driven behaviors. GBR 12909 (16 mg/kg) and (−) ephedrine (50 mg/kg) both enhanced locomotor activity and disrupted PPI, but combined treatment of either of these compounds with chlordiazepoxide had no significant additive effect on locomotor activity or prepulse inhibition. These findings suggest that the effect of the d-amphetamine/chlordiazepoxide mixture cannot be accounted for by the dopamine enhancing properties of amphetamine alone. Last, valproic acid (120-240 mg/kg) did not reduce the GBR-induced hyperactivity. Therefore, further pharmacological evaluation of GBR 12909-induced hyperactivity is warranted to determine its pharmacological potential to model mania-like behavior. Based on the current results, it is concluded that the utility of the pharmacological d-amphetamine/chlordiazepoxide assay as a tool to study brain mechanisms relevant to mania is limited.     

Intracortical 6-hydroxydopamine induces an asymmetrical behavioral response in the rat

Doses of 6-hydroxydopamine (6-OHDA) from 1 to 6 μg injected into the right frontal cortex of rats led to spontaneous hyperactivity beginning 1 week after injection and lasting throughout the 30-day postoperative period. Identical left hemispheric injections did not elicit hyperactivity until a dose of 6 μg was reached. Biochemical assays revealed that 6-OHDA injected into either hemisphere caused approximately the same amount of depletion of norepinephrine. These results suggest that the neural asymmetry which leads to hyperactivity may be post-synaptic to the cortical noradrenergic terminals.     

Lesion of the Cerebellar Noradrenergic Innervation Enhances the Harmaline-Induced Tremor in Rats

Abnormal synchronous activation of the glutamatergic olivo-cerebellar pathway has been suggested to be crucial for the harmaline-induced tremor. The cerebellum receives two catecholaminergic pathways: the dopaminergic pathway arising from the ventral tegmental area/substantia nigra pars compacta, and the noradrenergic one from the locus coeruleus. The aim of the present study was to examine a contribution of the cerebellar catecholaminergic innervations to the harmaline-induced tremor in rats. Rats were injected bilaterally into the cerebellar vermis with 6-hydroxydopamine (6-OHDA; 8 μg/0.5 μl) either alone or this treatment was preceded (30 min earlier) by desipramine (15 mg/kg ip). Harmaline was administered to animals in doses of 7.5 or 15 mg/kg ip. Tremor of forelimbs was measured as a number of episodes during a 90-min observation. Rats were killed by decapitation 30 or 120 min after harmaline treatment. The levels of dopamine, noradrenaline, serotonin, and their metabolites were measured by HPLC in the cerebellum, substantia nigra, caudate–putamen, and frontal cortex. 6-OHDA injected alone enhanced the harmaline-induced tremor. Furthermore, it decreased the noradrenaline level by ca. 40–80% in the cerebellum and increased the levels of serotonin and 5-HIAA in the caudate–putamen and frontal cortex in untreated and/or harmaline-treated animals. When 6-OHDA treatment was preceded by desipramine, it decreased dopaminergic transmission in some regions of the cerebellum while inducing its compensatory activation in others. The latter lesion did not markedly influence the tremor induced by harmaline. The present study indicates that noradrenergic innervation of the cerebellum interacts with cerebral serotonergic systems and plays an inhibitory role in the harmaline-induced tremor.     

Low brain glutathione and ascorbic acid associated with dopamine uptake inhibition during rat's development induce long-term cognitive deficit: relevance to schizophrenia

Schizophrenia is associated with a cerebral glutathione deficit, which may leave the brain susceptible to oxidants. To study the consequences of a glutathione deficit, we treated developing rats with L-buthionine-(S,R)-sulfoximine (BSO), an inhibitor of glutathione synthesis, and later investigated their behaviour until adulthood. Since rodents may in some occasions compensate for a glutathione deficit by ascorbic acid (AA), we used Osteogenic Disorder Shionogi (ODS) mutant rats, which like humans, cannot synthetize ascorbic acid. Moreover, as hyperactivity of the dopaminergic system may be associated with schizophrenia, some rats were treated with the dopamine uptake inhibitor GBR 12909. Whereas ODS rats treated with either BSO or GBR 12909 alone had normal behaviour, rats treated with both BSO and GBR 12909 failed to discriminate between familiar and novel objects although other behaviours proved to be normal. In contrast, nonmutant rats were not affected by treatment with BSO and GBR 12909. Our results suggest that low brain glutathione and ascorbic acid levels associated with a perturbation of the dopaminergic system actively participate in the development of some cognitive deficits affecting schizophrenic patients.     

Exploratory activity and conditioned avoidance acquisition after early postnatal 6-Hydroxydopamine administration

Summary Newborn rats were treated with 3 injections of 6-hydroxydopamine (6-OHDA) either subcutaneously (s. c.) or intracerebrally (i. cer.).General motor activity, exploratory activities (crossing and rearing) and acquisition of conditioned avoidance response (CAR) in shuttle-box were tested in the course of the 9th postnatal week.50g/g s. c. 6-OHDA injected daily during the first 3 postnatal days led to a moderate decrease in rearing but was ineffective on other behavioural parameters. A dose of 100g i. cer. (20, 30, 50g on days 2, 4, 6 resp.) resulted in a considerable reduction in both rearing and acquisition of CAR, while 200g i. cer. 6-OHDA (40, 60, 100g on days 2, 4, 6) induced a marked impairment in rearing and acquisition of CAR, and a less pronounced decrease in crossing activity. No significant change was found in general motor activity.Increasing doses of 6-OHDA led to an 80–90% depletion in brain catecholamines and induced a content-related behavioural suppression in rearing and conditioned avoidance responding in particular.It is concluded that intracerebral administration of 6-OHDA into neonatal rats almost completely prevents the development of central catecholaminergic neuronal systems, and concomitantly the capability of acquiring conditioned avoidance behaviour, while reducing exploratory behaviour.     

Postnatal iron-induced motor behaviour alterations following chronic neuroleptic administration in mice

Summary. C57/BL6 mice were administered either 7.5 mg Fe²⁺/kg or vehicle (saline) postnatally on days 10–12 after birth. From 61 days of age onwards for 21 days, groups of mice were administered either clozapine (1 or 5 mg/kg, s.c.) or haloperidol (1 mg/kg, s.c.) or vehicle (Tween-80). Twenty-four hours after the final injection of either neuroleptic compound or vehicle, spontaneous motor activity was measured over a 60-min interval. Following this, each animal was removed, injected apomorphine (1 mg/kg, s.c.) and replaced in the same test chamber. It was found that postnatal administration of Fe²⁺ at the 7.5 mg/kg dose level reduced activity during the initial 20-min periods (0–20 and 20–40 min) and then induced hyperactivity during the final 20-min period over all three parameters of activity. Subchronic treatment with the higher, 5 mg/kg, dose of clozapine abolished or attenuated the hypoactivity in by postnatal Fe²⁺ during the 1^st two 20-min periods over all three parameters of activity. Subchronic treatment with the higher, 5 mg/kg, dose of clozapine abolished or attenuated the hyperactivity in by postnatal Fe²⁺ during the 3^rd and final 20-min period. Subchronic administration of haloperidol, without postnatal iron, increased the level of both locomotion (1^st 20 min) and rearing (2^nd 20 min) activity. Postnatal administration of Fe²⁺ at the 7.5 mg/kg dose increased the levels of both locomotion and rearing, but not total activity, following administration of apomorphine (1 mg/kg). Subchronic administration of clozapine, at both the 1 and 5 mg/kg doses, reduced the increased locomotor activity caused by postnatal Fe²⁺, whereas clozapine, 5 mg/kg, elevated further the postnatal Fe²⁺-induced increased in rearing. Subchronic administration of clozapine, at both the 1 and 5 mg/kg doses, and haloperidol, 1 mg/kg, increased the level of locomotor following administration of apomorphine (1 mg/kg) in mice treated postnatally with vehicle, whereas only clozapine increased the level of rearing. Correlational analyses indicated that both apomorphine-induced locomotion and rearing were highly correlated with the total iron content in the basal ganglia, thereby offering direct evidence of the linear relationship between iron content in the basal ganglia and the behavioural expression of DA D₂-receptor supersensitivity in mice.     

Cross-sensitization between cocaine and GBR 12909, a dopamine uptake inhibitor.

In the present experiment, rats which had received repeated injections of 20 mg/kg GBR 12909, a selective dopamine uptake inhibitor, were subjected to challenge doses of GBR 12909 and cocaine which were near or below the threshold for locomotor activation in drug-naive animals. The behavior measured was general motor activity, monitored in photocell testing cages. Results verify earlier findings which showed that rats treated chronically with GBR 12909 display a potentiated locomotor response to low doses of the drug (behavioral sensitization). In addition, it was found that GBR 12909-sensitized rats show a potentiated locomotor response to 5 mg/kg cocaine (cross-sensitization). These results are discussed with regard to various mechanisms which may underlie the sensitization phenomena observed, including possible neurochemical changes at the presynaptic level.     

Proposed animal model of attention deficit hyperactivity disorder

Dopamine (DA) neurons are implicated in the hyperlocomotion of neonatal 6-hydroxydopamine (6-OHDA)-lesioned rats, an animal model of attention deficit hyperactivity disorder (ADHD). Because serotonin (5-HT) neurons mediate some DA agonist effects, we investigated the possible role of 5-HT neurons on locomotor activity. Rats were treated at 3 days after birth with vehicle or 6-OHDA (134 μg ICV; desipramine pretreatment, 20 mg/kg IP, 1 h), and at 10 weeks with vehicle or 5,7-dihydroxytryptamine (5,7-DHT; 75 μg ICV; pretreatment with desipramine and pargyline, 75 mg/kg IP, 30 min), to destroy DA and/or 5-HT fibers. Intense spontaneous hyperlocomotor activity was produced in rats lesioned with both 6-OHDA and 5,7-DHT. Locomotor time in this group was 550 ± 17 s in a 600 s session, vs. 127 ± 13 s in the 6-OHDA group and <75 s in 5,7-DHT and intact control groups (p < 0.001). Oral activity dose-effect curves established that 5,7-DHT attenuated DA D₁ receptor supersensitivity and further sensitized 5-HT_2c receptors. Acute treatment with dextroamphetamine (0.25 mg/kg SC) reduced locomotor time in 6-OHDA+5,7-DHT-lesioned rats to 76 ± 37 s (p < 0.001). Striatal DA was reduced by 99% and 5-HT was reduced by 30% (vs. 6-OHDA group). Because combined 6-OHDA (to neonates) and 5,7-DHT (to adults) lesions produce intense hyperlocomotion that is attenuated by amphetamine, we propose this as a new animal model of ADHD. The findings suggest that hyperactivity in ADHD may be due to injury or impairment of both DA and 5-HT neurons.     

Some central pharmacological effects of (+)- and (−)-oxaprotiline

Summary The central action of oxaprotiline (OXA) enantiomers, administered in a single dose, was studied in rats and mice. (+)-OXA and (–)-OXA attenuated reserpine- and apomorphine-induced hypothermia [(+)-OXA in a more potent manner] in mice and reduced the immobility time in the behavioural despair test in rats. Both OXA enantiomers inhibited locomotor activity in mice and rats, and enhanced and prolonged amphetamine- and apomorphine-induced stereotypy in rats. (–)-OXA potentiated the amphetamine hyperactivity in rats, but not in mice. Nomifensine hyperactivity in rats was unaffected by either enantiomer, and locomotor hypoactivity induced by low doses of apomorphine was also unchanged, as was L-DOPA-induced locomotor hyperactivity in mice. Apomorphine-induced climbing in mice was attenuated by (+)-OXA. Clonidine locomotor hypoactivity and hypothermia were unchanged, and clonidine-induced aggressiveness was attenuated by (+)-OXA. Neither OXA enantiomer affected the action of oxotremorine. In some tests the effect of OXA was stronger at 3 h than at 1 h after administration. The above results indicate that both OXA enantiomers—in particular (–)-OXA—increase some dopaminergic behavioural effects in rats.     

Effects of different doses of apomorphine on GAD activity in rat substantia nigra

The effects of different doses of the dopamine (DA) receptor agonist apomorphine on the activity of the γ-aminobutyric acid (GABA)-synthesizing enzyme glutamic acid decarboxylase (GAD, EC 4.1.1.15) were investigated in rat substantia nigra in comparison with haloperidol and sulpiride, two DA receptor blocking agents. Results obtained show that low doses (10,35 μg/kg, s.c.) of apomorphine induce a decrease in nigral GAD activity whilst an opposite effect is observed with the highest dose (1000 μg/kg, s.c.). No significant change is observed following injection of the intermediate doses (100 and 500 μg/kg, s.c.). Moreover, sulpiride at the dose used (2 mg/kg, i.p.) induces an increase in GAD activity whilst no effect follows systemic injection of the same dose of haloperidol. The results are discussed in light of recent neurochemical and behavioral data.

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Sommario Sono stati studiati gli effetti comparativi dell' apomorfina, agonista recettoriale dopaminergico, e di due dopamino-bloccanti, l'aloperidolo ed il sulpiride, sull'attività glutamato decarbossilasica nella sostanza nera di ratto. I risultati ottenuti dimostrano che basse dosi (10,35 μg/kg, s.c.) di apomorfina inducono un decremento dell'attività GAD nigrale mentre un effetto opposto è indotto dalla dose più alta (1000 μg/kg, s.c.). Le dosi intermedie (100 e 500 μg/kg, s.c.) non inducono alcuna significativa modificazione. Il sulpiride alla dose usata (2 mg/kg, i.p.) causa un incremento dell'attività GAD mentre nessun effetto si osserva a seguito della somministrazione sistematica di aloperidolo. I risultati sono discussi alla luce di recenti dati neurochimici e comportamentali.

     

Hyperactivity and hypoactivity produced by lesions to the mesolimbic dopamine system

Spontaneous locomotor activity and the locomotor response to amphetamine and apomorphine were studied in rats subjected to either radiofrequency (RF), 6-hydroxydopamine (6-OHDA) or both RF and 6-OHDA lesions of the mesolimbic dopamine (DA) system. Large 6-OHDA lesions of the ventral tegmental area (VTA) or of the nucleus accumbens (N.Acc.) produced hypo-activity in the open field, a complete blockade of the locomotor stimulating effects of D-amphetamine and a profound supersensitive response to apomorphine as measured by a significant increase in locomotor activity as compared to sham-operated animals. In contrast, smaller 6-OHDA lesions of the VTA produced significant increases in spontaneous daytime and nocturnal activity with the biggest effect occurring at the lowest dose. RF lesions to the VTA produced even greater hyperactivity which was blocked by the addition of a 6-OHDA lesion to the N.Acc. The rats with RF lesions to VTA alone that were spontaneously hyperactive remained hyperactive after injection of amphetamine, whereas apomorphine produced a significant decrease in this hyperactivity. In contrast, the rats with the combined RF lesion and N.Acc. 6-OHDA lesion showed a blockade of the locomotor stimulating effects of D-amphetamine and a potentiated response to apomorphine identical to that observed with a N.Acc. lesion alone. All lesion groups revealed massive depletion of DA in the N.Acc. and anterior striatum with significantly greater depletions in those groups showing hypoactivity and hypo-responsiveness to amphetamine. All groups except the N.Acc. 6-OHDA alone group showed significant depletions of DA in the posterior striatum. Thus, limited destruction of the mesolimbic DA system can produce hyperactivity, but more extensive destruction of this system in the region of the N.Acc. and anterior striatum can reverse this hyperactivity and produce a hypo-responsiveness to the locomotor stimulating effects of amphetamine. These results suggest an essential role for dopamine in the expression of spontaneous and stimulant-induced activity. Furthermore, the much larger increase in spontaneous activity in the RF-VTA lesion group as compared to the VTA-6-OHDA groups suggests the presence of an, as yet unidentified, powerful inhibitory influence to the mesolimbic DA system within the midbrain tegmentum.     

Postnatal iron overload destroys NA-DA functional interactions

Summary. C57/BL6 mice were administered either postnatal iron (Fe²⁺ 7.5 mg/kg, on postnatal days 10–12) or vehicle, followed by administration of either DSP4 (50 mg/kg, s.c., 30 min after injection of zimeldine, 20 mg/kg, s.c.) or vehicle (saline) at 63 days of age. Three weeks later, iron/vehicle treated, DSP4/vehicle treated mice were injected with either a low dose of MPTP (2 × 20 mg/kg, with a 24-hr interval between injections) or vehicle. Behaviour testing took place a further three weeks (spontaneous behaviour and L-Dopa induced) and two weeks (clonidine-L-Dopa induced) later. Postnatal iron administration exacerbated the bradykinesia induced by MPTP and virtually abolished all spontaneous motor activity in NA-denervated mice that were MPTP-treated. Postnatal iron administration reduced markedly the restoration of motor activity by suprathreshold L-Dopa (20 mg/kg) following a 60-min habituation to the test chambers. Pretreatment with DSP4 effectively eliminated the restorative effect of L-Dopa in the MPTP mice. The synergistic effects of co-administration of clinidine (1 mg/kg) with a subthreshold dose of L-Dopa (5 mg/kg) in elevating the motor activity of MPTP mice were reduced markedly by postnatal iron administration, as well as by pretreatment with DSP4. NA-denervation by DSP4, after postnatal iron treatment, totally abolished the activity-elevating effects of the α-adrenoceptor agonist + DA-precursor combination in MPTP mice, and virtually eliminated these effects in saline (non-MPTP) mice. Postnatal iron administration caused enduring higher levels of total iron content in all the groups with an increased level in mice treated with DSP4 followed by MPTP. These divergent findings confirm the direct influence of NA innervation upon dopaminergic functional expression and indicate a permanent vulnerability both in the noradrenergic and dopaminergic pathways following the postnatal infliction of an iron overload.     

Subchronic administration of haloperidol influences the functional deficits of postnatal iron administration in Mice

C57/BL6 mice were administered either 7.5 mg Fe(2+) (II)/ kg or vehicle (saline) postnatally on Days 10-12 after birth. From 64 days of age onwards for 24 days, groups of mice were administered either haloperidol (0.25 or 1 or 2 mg/kg, s.c.) or vehicle (Tween-80). Twenty-four hours after the final injection of either neuroleptic compound or vehicle, spontaneous motor activity was measured over a 60-min interval. Postnatal Fe(2+)-treatment (7.5 mg/kg, postnatally) reduced motor activity parameters during the initial 20-min periods (0-20 and 20-40 min) and then induced hyperactivity during the final 20-min period over all three parameters of activity, confirming previous observations. Subchronic administration of haloperidol, at the 1 and 2 mg/kg doses, and to a lesser extent the 0.25 mg/kg dose, increased the levels of activity in all three motor activity parameters in postnatal iron-treated mice: locomotion (1st and 2nd 20 min periods), rearing (1st and 2nd 20 min periods) and total activity (1st 20 min period). All three doses of haloperidol abolished the later hyperactivity in iron-treated mice, with the exception of the 0.25 mg/kg dose with regard to rearing behaviour. Apomorphine (1 mg/kg, s.c.)-induced activity was elevated by postnatal iron administration and by subchronic administration of apomorphine at the higher dose levels. In the context of these and other observations, it is suggested that subchronic administration of haloperidol interacting with postnatal iron induces different expressions of dopamine neuron comorbidity underlying movement disorder.     

Neurobehavioural deficits associated with apoptotic neurodegeneration and vulnerability for ADHD

Several studies involving postnatal administration of the N-methyl-D-aspartate (NMDA) antagonists, dizocilpine (MK-801; 3 x 0.5 mg/kg, at 08.00, 16.00 and 24.00 h) on Postnatal day 11, or Ketamine (1 × 50 mg/kg) or Ethanol (1 × 2.5 g/kg, Ethanol-Low, or 2 × 2.5 g/kg, 2-h interval, Ethanol-High) on Postnatal day 10, are described. Some mice from each treatment/vehicle group were sacrificed 24 h after NMDA antagonist treatment and brain regions were taken for fluoro-jade staining analysis. Functional analysis was initiated at 60 days of age. All three treatments inducing an antagonistic action at NMDA receptors, MK-801, Ketamine and Ethanol-High induced a similar pattern of initial hypoactivity followed by marked and lasting hyper-activity in the motor activity test chambers. In each case, the basal hyperactivity level was abolished by acute treatment with a low dose of D-amphetamine (0.25 mg/kg). All three treatments, MK-801, Ketamine and Ethanol-High, induced a deficit in acquisitive performance in the radial arm maze test of instrumental learning. The deficit induced by postnatal MK-801 was abolished by acute treatment with the low dose of D-amphetamine. All three treatments, MK-801, Ketamine and Ethanol-High, resulted in normal acquisitive performance during the first three test days in the circular swim with the submerged platform maintained in a constant position, but on the fourth test day, with the platform position shifted to a different “quadrant”, induced marked deficits. Fluoro-jade staining analyses indicated a devastating cell degeneration in several brain regions of mice administered NMDA antagonists postnatally, including the hippocampus, frontal cortex, parietal cortex, and cerebellum. Severe cell degeneration in the laterodorsal thalamus due to Ethanol or diazepam (5 mg/kg) appeared not to affect the different aspects of function. The pattern of dysfunctional outcome and apoptotic cell loss following postnatal NMDA antagonist treatment offers a plausible similarity to the major aspects of ‘syndromatic continuity’ in ADHD, hyperactivity, inattention and impulsivity, thereby providing an interesting animal model of the disorder.     

Hyperactivity following postnatal NMDA antagonist treatment: Reversal by D-amphetamine

Three experiments were performed to study the effects of neonatal administration of glutamate receptor antagonists, on either Day 11 (dizocilpine=MK-801, 3×0.5 mg/kg, s.c., injectioned at 0800, 1600 and 2400 h) or Day 10 (Ketamine, 1×50 mg/kg, s.c., or Ethanol-Low, 1×2.5 mg/kg, or, Ethanol-High, 2×2.5 mg/kg, s.c., with 2-h interval) to male mice pups, on spontaneous motor behavior, habituation to a novel situation, and D-amphetamine-induced activity in the adult animals. Mice administered MK-801 showed initial hypoactivity followed by hyperactivity over the later (20–40 and 40–60 min) periods of testing. Mice administered Ketamine and Ethanol-High similarly displayed an initial hypoactivity followed by hyperactivity over the later time (20–60 min) of testing. Habituation to the novel activity test chambers was reduced drastically in the MK-801 mice compared with vehicletreated mice. Similarly, mice administered Ketamine and Ethanol-High dislayed too drastically reduced habituation behavior. The low dose ofd-amphetamine (0.25 mg/kg) reduced hyperactivity of neonatal MK-801-treated mice, particularly from 30–60 min onwards, and elevated the activity level of the vehicle-treated mice. Similarly, the low dose ofd-amphetamine (0.25 mg/kg) reduced the hyperactivity of neonatally Ketamine-treated and Ethanol-High-treated mice, particularly from 30–60 min onwards, and elevated the activity level of the respective vehicle-treated mice. Fluoro-jade staining per mm² regional brain tissue of MK-801 mice pups expressed as percent of vehicle mice pups showed also that the extensiveness of staining was markedly greater in the parietal cortex, hippocampus, frontal cortex, and lesser so in the laterodorsal thalamus. Ketamine-treated mice showed cell degeneration mainly in the parietal cortex, whereas the Ethanol-High mice showed marked cell degeneration in both the parietal and laterodorsal cortex. The present findings that encompass a pattern of regional neuronal degeneration, disruptions of spontaneous motor activity, habituation deficits and reversal of hyperactivity by a low dose ofd-amphetamine suggest a model of Attention Deficit Hyperactivity Disorder that underlines the intimate role ofN-methyl-d-aspartate (NMDA) receptors in the developing brain.     

Inhibitory effect of the DA uptake blocker GBR 12909 on sodium channels of hippocampal neurons

The effect of the selective dopamine uptake inhibitor GBR 12909 on TTX-sensitive sodium channels of cultured hippocampal neurons was investigated using whole cell patch-clamp technique. GBR 12909 dose-dependently inhibited sodium currents evoked by trains of depolarizing pulses with an IC50 of 6.3 microM. A weaker inhibition (IC50 = 17-35 microM) could be observed when currents were evoked by either single pulse depolarization or from hyperpolarized holding membrane potential. These data indicate that the extent of inhibition caused by GBR 12909 depends on the physiological activity pattern of neurons. Our results suggest that caution is needed for the interpretation of data when GBR 12909 is used for the inhibition of dopamine uptake at concentrations above the submicromolar range.     

Clinically available iron chelators induce neuroprotection in the 6-OHDA model of Parkinson’s disease after peripheral administration

The iron content of the substantia nigra pars compacta increases in the brains of Parkinson’s disease patients. Hence, its removal by iron chelators may retard the progression of the disease. However, information on the ability of clinically available iron chelators to cross the blood brain barrier and be neuroprotective is limited. In this present study three iron chelators, which are currently approved for clinical use, namely the hexadendate, deferrioxamine, the bidentate deferiprone and the tridendate chelator deferasirox have been investigated for their efficacy to induce neuroprotection. Previous studies have shown that both deferiprone and deferrioxamine exert neuroprotection in the 6-hydroxy dopamine (6-OHDA) model but no such studies have investigated deferasirox. Focal administration of deferasirox (0.5, 2 and 10 μg) into the substantia nigra pars compacta of rats significantly attenuated the loss of dopaminergic neurons and striatal dopamine content resulting from 6-OHDA toxicity. Systemic administration of deferasirox (20 mg/kg), deferiprone (10 mg/kg) or deferrioxamine (30 mg/kg), to the 6-OHDA rat model of Parkinson’s disease, significantly attenuated the loss of dopaminergic neurons and striatal dopamine content. Further studies to comprehend the action of these chelators showed that local application of either 0.4 mM deferrioxamine, or 1 mM deferasirox, via a microdialysis probe into the striatum, prior to that of 200 μM 6-OHDA, prevented the generation of hydroxyl radicals. Our results confirm that the administration of these chelators show therapeutic efficacy and should be considered as therapeutic agents for the treatment of Parkinson’s disease.