Blockade of adenosine A2A receptors inhibits Tremulous Jaw Movements as well as expression of zif-268 and GAD65 mRNAs in brain motor structures.

Tremor is one of the motor symptoms of Parkinson's disease (PD), present also in neuroleptic-induced parkinsonism. Tremulous Jaw Movements (TJMs) are suggested to be a well-validated rodent model of PD resting tremor. TJMs can be induced by typical antipsychotics and are known to be reduced by different drugs, including adenosine A2A receptor antagonists. The aim of the present study was to search for brain structures involved in the tremorolytic action of SCH58261, a selective A2A receptor antagonist, in TJMs induced by subchronic pimozide. Besides TJMs, we evaluated in the same animals the expression of zif-268 mRNA (neuronal responsiveness marker), and mRNA levels for glutamic acid decarboxylase 65-kDa isoform (GAD65) and vesicular glutamate transporters 1 and 2 (vGluT1/2) in selected brain structures, as markers of GABAergic and glutamatergic neurons, respectively. We found that SCH58261 reduced the pimozide-induced TJMs. Pimozide increased the zif-268 mRNA level in the striatum, nucleus accumbens (NAc) core, and substantia nigra pars reticulata (SNr). Additionally, it increased GAD65 mRNA in the striatum and SNr, and vGluT2 mRNA levels in the subthalamic nucleus (STN). A positive correlation between zif-268, GAD65 and vGluT2 mRNAs and TJMs was found. SCH58261 reversed the pimozide-increased zif-268 mRNA in the striatum and NAc core and GAD65 mRNA in the striatum and SNr. In contrast, SCH58261 did not influence vGluT2 mRNA in STN. The present study suggests an importance of the striato-subthalamo-nigro-thalamic circuit in neuroleptic-induced TJMs. The tremorolytic effect of A2A receptor blockade seems to involve this circuit bypassing, however, STN.

The influence of preconditioning with low dose of LPS on paraquat-induced neurotoxicity, microglia activation and expression of α-synuclein and synphilin-1 in the dopaminergic system.

BACKGROUND: Prolonged inflammation, oxidative stress, and protein aggregation are important factors contributing to Parkinson's disease (PD) pathology. A known ROS generator, pesticide paraquat (PQ), was indicated as an environmental substance potentially increasing the incidence of PD and is used to model this disease. We investigated if a combination of inflammation and oxidative stress in subthreshold doses would exacerbate the modelled neuropathology. METHODS: We examined the late effects of acute or repeated peripheral inflammation induced by low dose of LPS (10 µg/kg, ip) on PQ toxicity in the rat nigrostriatal dopaminergic pathway, microglial activation markers and expression of major Lewy bodies proteins, α-synuclein and synphilin-1. RESULTS: We observed that LPS increased, while PQ decreased body temperature and microglia CD11b expression in the SN. Single LPS pretreatment, 3 h before repeated weekly PQ injections (4×) slightly aggravated neuronal degeneration in the SN. Moreover, degeneration of dopaminergic neurons after weekly repeated inflammation itself (4×) was observed. Interestingly, repeated LPS administration combined with each PQ dose counteracted such effect. The expression of α-synuclein decreased after repeated LPS injections, while only combined, repeated LPS and PQ treatment lowered the levels of synphilin-1. Therefore, α-synuclein and synphilin-1 expression change was influenced by different mechanisms. Concomitantly, decreased levels of the two proteins correlated with decreased degeneration of dopaminergic neurons and with a normalized microglia activation marker. CONCLUSIONS: Our results indicate that both oxidative insult triggered by PQ and inflammation caused by peripheral LPS injection can individually induce neurotoxicity. Those factors act through different mechanisms that are not additive and not selective towards dopaminergic neurons, probably implying microglia. Repeated, but small insults from oxidative stress and inflammation when administered in significant time intervals can counteract each other and even act protective as a preconditioning effect. The timing of such repetitive insults is also of essence.

Antiparkinsonian-like effects of CPL500036, a novel selective inhibitor of phosphodiesterase 10A, in the unilateral rat model of Parkinson's disease.

Phosphodiesterase 10A (PDE10A), the enzyme which catalyzes hydrolysis of cyclic adenosine monophosphate (cAMP) and cyclic guanosine monophosphate (cGMP), is located almost exclusively in striatal γ-amino-butyric acid (GABA)ergic medium spiny neurons (MSNs). Since dopaminergic deficiency in Parkinson's disease (PD) leads to functional imbalance of striatal direct and indirect output pathways formed by MSNs, PDE10A seems to be of special interest as a potential therapeutic target in PD. The aim of the present study was to examine the influence of 7-{5,8-dimethyl-[1,2,4]triazolo[1,5-a]pyrazin-2-yl}-2-phenylimidazo[1,2-a]pyrimidine (CPL500036), a novel selective inhibitor of PDE10A, on sensorimotor deficits and therapeutic effects of L-3,4-dihydroxyphenylalanine (L-DOPA) in hemiparkinsonian rats. Animals were unilaterally lesioned with 6-hydroxydopamine, and their sensorimotor deficits were examined in the stepping, cylinder, vibrissae and catalepsy tests. CPL500036 (0.1 and 0.3 mg/kg) was administered either acutely or chronically (2 weeks), alone or in combination with L-DOPA/benserazide (6 mg/kg/6 mg/kg). Acute treatment with CPL500036 reversed the lesion-induced impairments of contralateral forelimb use in the stepping and cylinder tests but did not influence deficits in the vibrissae test and the lesion-induced catalepsy. Moreover, CPL500036 did not diminish the therapeutic effects produced by acute and chronic treatment with L-DOPA in these tests. The present study suggests a potential use of CPL500036 as a co-treatment to L-DOPA in PD therapy.

Pramipexole Reduces zif-268 mRNA Expression in Brain Structures involved in the Generation of Harmaline-Induced Tremor.

Essential tremor is one of the most common neurological disorders, however, it is not sufficiently controlled with currently available pharmacotherapy. Our recent study has shown that pramipexole, a drug efficient in inhibiting parkinsonian tremor, reduced the harmaline-induced tremor in rats, generally accepted to be a model of essential tremor. The aim of the present study was to investigate brain targets for the tremorolytic effect of pramipexole by determination of the early activity-dependent gene zif-268 mRNA expression. Tremor in rats was induced by harmaline administered at a dose of 15 mg/kg ip. Pramipexole was administered at a low dose of 0.1 mg/kg sc. Tremor was measured by Force Plate Actimeters where four force transducers located below the corners of the plate tracked the animal's position on a Cartesian plane. The zif-268 mRNA expression was analyzed by in situ hybridization in brain slices. Harmaline induced tremor and increased zif-268 mRNA levels in the inferior olive, cerebellar cortex, ventroanterior/ventrolateral thalamic nuclei and motor cortex. Pramipexole reversed both the harmaline-induced tremor and the increase in zif-268 mRNA expression in the inferior olive, cerebellar cortex and motor cortex. Moreover, the tremor intensity correlated positively with zif-268 mRNA expression in the above structures. The present results seem to suggest that the tremorolytic effect of pramipexole is related to the modulation of the harmaline-increased neuronal activity in the tremor network which includes the inferior olive, cerebellar cortex and motor cortex. Potential mechanisms underlying the above pramipexole action are discussed.

Potential antipsychotic action of the selective agonist of adenosine A1 receptors, 5'-Cl-5'-deoxy-ENBA, in amphetamine and MK-801 rat models.

BACKGROUND: Disturbances of dopaminergic and glutamatergic transmissions have been suggested to be involved in the pathomechanisms underlying psychotic symptoms of schizophrenia. In line with this concept, hyperlocomotion induced by the dopaminomimetic amphetamine and the uncompetitive antagonist of NMDA receptors MK-801 (dizocilpine) in rodents is a generally established model for screening of new potential antipsychotic drugs. Since recent studies have indicated that receptors for adenosine may be targets for antipsychotic therapy, the aim of the present study was to investigate an influence of 5'-Cl-5'-deoxy-ENBA, a potent and selective adenosine A1 receptor agonist, on hyperlocomotion induced by amphetamine and MK-801. METHODS: Locomotor activity was measured by Force Plate Actimeters where four force transducers located below the corners of the floor of the cage tracked the animal position on a Cartesian plane at each time point. RESULTS: Hyperlocomotion induced by either amphetamine (1 mg/kg sc) or MK-801 (0.3 mg/kg ip) was inhibited by 5'-Cl-5'-deoxy-ENBA (0.1 mg/kg ip). The effect of 5'-Cl-5'-deoxy-ENBA on the amphetamine- and MK-801-induced hyperlocomotion was antagonized by the selective antagonist of adenosine A1 receptor DPCPX at doses of 1 and 2 mg/kg ip, respectively. CONCLUSION: The present study suggests that stimulation of adenosine A1 receptors may produce antipsychotic effects.

Inhibition of Excessive Glutamatergic Transmission in the Ventral Thalamic Nuclei by a Selective Adenosine A1 Receptor Agonist, 5'-Chloro-5'-Deoxy-(±)-ENBA Underlies its Tremorolytic Effect in the Harmaline-Induced Model of Essential Tremor.

The primary cause of harmaline tremor, which is a model of essential tremor (ET) in animals, is excessive activation of olivocerebellar glutamatergic climbing fibers. Our recent study indicated that 5'-chloro-5'-deoxy-(±)-N6-(±)-(endo-norborn-2-yl)adenosine (5'Cl5'd-(±)-ENBA), a potent and selective adenosine A1 receptor (A1) agonist, inhibited harmaline tremor. The present study was aimed to evaluate the role of glutamatergic transmission system in 5'Cl5'd-(±)-ENBA tremorolytic action in the harmaline model in rats, by analyzing glutamate release in the motor nuclei of the thalamus and mRNA expression of glutamatergic neuron markers (vGlut1/2) in reference to the general neuronal activity marker (zif-268) in different brain structures. The extracellular glutamate level in the motor thalamus was evaluated by in vivo microdialysis and the vGlut1/vGlut2 and zif-268 mRNA expression was analyzed by in situ hybridization. The intensity of tremor was measured automatically using Force Plate Actimeters (FPAs). 5'Cl5'd-(±)-ENBA (0.5 mg/kg) given 30 min before harmaline (30 mg/kg) decreased the harmaline-induced excessive glutamate release in the motor thalamus and reversed harmaline-induced molecular effects, such as elevation of the vGlut1 mRNA expression in the inferior olive (IO) and decrease in the motor cortex, as well as an increase of the zif-268 mRNA expression in the IO, motor thalamus and motor cortex. Moreover, 5'Cl5'd-(±)-ENBA reduced harmaline tremor by lowering its power in 9-15 Hz frequency band. Our findings show that A1 stimulation decreases glutamate release in the motor thalamic nuclei in the harmaline model of ET, suggesting that A1 receptors, especially in this structure, may be a potential therapeutic target in this disorder.

Zona incerta as a therapeutic target in Parkinson's disease.

The zona incerta has recently become an important target for deep-brain stimulation (DBS) in Parkinson's disease (PD). The present review summarizes clinical, animal and anatomical data which have indicated an important role of this structure in PD, and discusses potential mechanisms involved in therapeutic effects of DBS. Animal studies have suggested initially some role of neurons as well as GABAergic and glutamatergic receptors of the zona incerta in locomotion and generation of PD signs. Anatomical data have indicated that thanks to its multiple interconnections with the basal ganglia, thalamus, cerebral cortex, brainstem, spinal cord and cerebellum, the zona incerta is an important link in a neuronal chain transmitting impulses involved in PD pathology. Finally, clinical studies have shown that DBS of this structure alleviates parkinsonian bradykinesia, muscle rigidity and tremor. DBS of caudal zona incerta seemed to be the most effective therapeutic intervention, especially with regard to reduction of PD tremor as well as other forms of tremor.

Tremorolytic effect of 5'-chloro-5'-deoxy-(±)-ENBA, a potent and selective adenosine A1 receptor agonist, evaluated in the harmaline-induced model in rats.

AIM: The aim of this study was to examine the role of adenosine A1 receptors in the harmaline-induced tremor in rats using 5'-chloro-5'-deoxy-(±)-ENBA (5'Cl5'd-(±)-ENBA), a brain-penetrant, potent, and selective adenosine A1 receptor agonist. METHODS: Harmaline was injected at a dose of 15 mg/kg ip and tremor was measured automatically in force-plate actimeters by an increased averaged power in the frequency band of 9-15 Hz (AP2) and by tremor index (a difference in power between AP2 and averaged power in the frequency band of 0-8 Hz). The zif-268 mRNA expression was additionally analyzed by in situ hybridization in several brain structures. RESULTS: 5'Cl5'd-(±)-ENBA (0.05-0.5 mg/kg ip) dose dependently reduced the harmaline-induced tremor and this effect was reversed by 8-cyclopentyl-1,3-dipropylxanthine (DPCPX), a selective antagonist of adenosine A1 receptors (1 mg/kg ip). Harmaline increased the zif-268 mRNA expression in the inferior olive, cerebellar cortex, ventroanterior/ventrolateral thalamic nuclei, and motor cortex. 5'Cl5'd-(±)-ENBA reversed these increases in all the above structures. DPCPX reduced the effect of 5'Cl5'd-(±)-ENBA on zif-268 mRNA in the motor cortex. CONCLUSION: This study suggests that adenosine A1 receptors may be a potential target for the treatment of essential tremor.

Adaptation within mitochondrial oxidative phosphorylation supercomplexes and membrane viscosity during degeneration of dopaminergic neurons in an animal model of early Parkinson's disease.

In Parkinson's disease (PD) motor symptoms are not observed until loss of 70% of dopaminergic neurons in substantia nigra (SN), preventing early diagnosis. Mitochondrial dysfunction was indicated in neuropathological process already at early PD stages. Aging and oxidative stress, the main factors in PD pathogenesis, cause membrane stiffening, which could influence functioning of membrane-bound oxidative phosphorylation (OxPhos) complexes (Cxs) in mitochondria. In 6-OHDA rat model, medium-sized dopaminergic lesion was used to study mitochondrial membrane viscosity and changes at the level of OxPhos Cxs and their higher assembled states-supercomplexes (SCxs), during the early degeneration processes and after it. We observed loss of dopaminergic phenotype in SN and decreased dopamine level in striatum (STR) before actual death of neurons in SN. Behavioural deficits induced by lesion were reversed despite progressing neurodegeneration. Along with degeneration process in STR, mitochondrial Cx I performance and amount decreased in almost all forms of SCxs. Also, progressing decrease of Cx IV performance in SCxs (I1III2IV3-1, I1IV2-1) in STR was observed during degeneration. In SN, SCxs containing Cx I increased protein amount and a shifted individual Cx I1 into superassembled states. Importantly, mitochondrial membrane viscosity changed in parallel with altered SCxs performance. We show for the first time changes at the level of mitochondrial membrane viscosity influencing SCxs function after dopaminergic system degeneration. It implicates that altered mitochondrial membrane viscosity could play an important role in regulation of mitochondria functioning and pathomechanisms of PD. The data obtained are also discussed in relation to compensatory processes observed.

Pramipexole at a Low Dose Induces Beneficial Effect in the Harmaline-induced Model of Essential Tremor in Rats.

AIMS: The aim of the study was to examine the effects of preferential agonists of dopamine D3 receptors: pramipexole and 7-OH-DPAT on the harmaline-induced tremor in rats (a model of essential tremor, ET). To study receptor mechanisms of these drugs, rats were pretreated with dopamine D3 receptor antagonists--SB-277011-A and SR-21502, an antagonist of presynaptic D2/D3 receptors--amisulpride, or a nonselective antagonist of D2-like receptors, haloperidol, at a postsynaptic dose. METHODS: For tremor measurement, fully automated force plate actimeters were used and data were analyzed using fast Fourier transform. RESULTS: Harmaline (15 mg/kg ip)-triggered tremor was manifested by an increase in the power within 9-15 Hz band (AP2). Pramipexole administered at a low (0.1 mg/kg sc), but not higher doses (0.3 and 1 mg/kg sc), and 7-OH-DPAT (0.1, 0.3, and 1 mg/kg sc) reversed the harmaline-increased AP2. None of the examined dopamine antagonists: SB-277011-A (10 mg/kg ip), SR-21502 (15 mg/kg ip), haloperidol (0.5 mg/kg ip), or amisulpride (1 mg/kg ip) influenced the above effect of dopamine agonists. CONCLUSION: The present study indicates that pramipexole reduces the harmaline-induced tremor, which may suggest its beneficial effects in ET patients. However, mechanisms underlying its action are still unclear and need further examination.

Apomorphine enhances harmaline-induced tremor in rats.

BACKGROUND: Harmaline-induced tremor is a well-known model of essential tremor in humans. The aim of the present study was to examine the influence of apomorphine, a non-selective dopamine receptor agonist, on the tremor induced by harmaline in rats. Propranolol (a first-line drug in essential tremor) was used as a reference compound. METHODS: Tremor, locomotor activity and focused stereotypy were measured objectively using force plate actimeters. Tremor was analyzed using a Fourier transform to generate power spectra for rhythmic behavior. RESULTS: The tremor induced by harmaline administered at a dose of 15 mg/kg ip was associated with an increase in power in the 9-15 Hz band (AP2) and in the tremor index, calculated as a difference between AP2 and power in the 0-8 Hz band (AP1). Propranolol injected at a dose of 20mg/kg ip reversed both of these effects of harmaline. Apomorphine administered at the doses of 0.5 and 1mg/kg sc further enhanced AP2 and at the lower dose also the tremor index elevated by harmaline. This increase in AP2 was stronger than enhancement of locomotor activity induced by apomorphine in the harmaline-treated animals. CONCLUSIONS: The present study suggests that the dopamine agonist apomorphine enhances the tremor induced by harmaline, and this effect is at least partly independent of hyperactivity.

Alterations of BDNF and trkB mRNA expression in the 6-hydroxydopamine-induced model of preclinical stages of Parkinson's disease: an influence of chronic pramipexole in rats.

Our recent study has indicated that a moderate lesion of the mesostriatal and mesolimbic pathways in rats, modelling preclinical stages of Parkinson's disease, induces a depressive-like behaviour which is reversed by chronic treatment with pramipexole. The purpose of the present study was to examine the role of brain derived neurotrophic factor (BDNF) signalling in the aforementioned model of depression. Therefore, we investigated the influence of 6-hydoxydopamine (6-OHDA) administration into the ventral region of the caudate-putamen on mRNA levels of BDNF and tropomyosin-related kinase B (trkB) receptor. The BDNF and trkB mRNA levels were determined in the nigrostriatal and limbic structures by in situ hybridization 2 weeks after the operation. Pramipexole (1 mg/kg sc twice a day) and imipramine (10 mg/kg ip once a day) were injected for 2 weeks. The lesion lowered the BDNF and trkB mRNA levels in the hippocampus [CA1, CA3 and dentate gyrus (DG)] and amygdala (basolateral/lateral) as well as the BDNF mRNA content in the habenula (medial/lateral). The lesion did not influence BDNF and trkB expression in the caudate-putamen, substantia nigra, nucleus accumbens (shell and core) and ventral tegmental area (VTA). Chronic imipramine reversed the lesion-induced decreases in BDNF mRNA in the DG. Chronic pramipexole increased BDNF mRNA, but decreased trkB mRNA in the VTA in lesioned rats. Furthermore, it reduced BDNF and trkB mRNA expression in the shell and core of the nucleus accumbens, BDNF mRNA in the amygdala and trkB mRNA in the caudate-putamen in these animals. The present study indicates that both the 6-OHDA-induced dopaminergic lesion and chronic pramipexole influence BDNF signalling in limbic structures, which may be related to their pro-depressive and antidepressant activity in rats, respectively.

Pramipexole but not imipramine or fluoxetine reverses the "depressive-like" behaviour in a rat model of preclinical stages of Parkinson's disease.

Depression is a frequent comorbid disorder in Parkinson's disease and may antedate its motor symptoms. However, mechanisms underlying Parkinson's disease-associated depression are unknown and its current medication is insufficient. The aim of the present study was to compare antidepressant-like effects of imipramine, fluoxetine and pramipexole in a model of preclinical stages of Parkinson's disease in rats. 6-Hydroxydopamine was bilaterally injected into the ventrolateral region of the caudate-putamen in rats. This treatment induced moderate decreases in the levels of dopamine and its metabolites in the caudate-putamen, nucleus accumbens and frontal cortex and reduced the density of tyrosine hydroxylase-immunoreactive neurons in the substantia nigra pars compacta and ventral tegmental area. The lesion increased immobility measured in the forced swimming test without influencing locomotor activity. Chronic (13 days) administration of pramipexole (1mg/kg sc/twice a day) reversed prolongation of the immobility time in lesioned animals but did not stimulate their locomotion. Chronic pramipexole activated dopaminergic transmission in the brain structures which might contribute to its effectiveness in the forced swimming test. In contrast, the 13-day administration of imipramine (10mg/kg ip/day) and fluoxetine (10mg/kg ip/day) did not shorten the immobility time in lesioned rats but reduced their locomotion. The present study indicates that already a moderate lesion of dopaminergic neurons induces "depressive-like" behaviour in animals which is reversed by chronic administration of the antiparkinsonian drug, pramipexole.

Lu AF21934, a positive allosteric modulator of mGlu4 receptors, reduces the harmaline-induced hyperactivity but not tremor in rats.

Harmaline induces tremor in animals resembling essential tremor which has been suggested to result from activation of the glutamatergic olivo-cerebellar projection. The aim of the present study was to examine the effects of systemic administration of Lu AF21934, a brain-penetrating positive allosteric modulator of the metabotropic glutamate receptor 4 (mGlu4), on the harmaline-induced tremor and other forms of motor activity in rats using fully automated Force Plate Actimeters. The influence of harmaline on the mGlu4 mRNA expression in the cerebellum and inferior olive was analysed by in situ hybridization. Harmaline at a dose of 15 mg/kg (ip) triggered tremor which was manifested by an increase in the power within 9-15 Hz band and in the tremor index (a difference in power between bands 9-15 Hz and 0-8 Hz). Harmaline induced a biphasic effect on mobility, initially inhibiting the exploratory locomotor activity of rats (0-30 min after administration), followed by an increase in their basic activity. Lu AF21934 (0.5-5 mg/kg sc) did not influence tremor but at doses of 0.5 and 2.5 mg/kg reversed harmaline-induced hyperactivity. MGlu4 mRNA expression was high in the cerebellar cortex and low in the inferior olive. Repeated harmaline (15 mg/kg ip once a day for 5 days] decreased mGlu4 mRNA in the cerebellum and inferior olive. The present study indicates that the mGlu4 stimulation counteracts hyperactivity induced by harmaline which suggests the involvement of cerebellar glutamatergic transmission in this process. In contrast, neuronal mechanisms involved in tremor seem to be insensitive to the stimulation of mGlu4.

A partial lesion of the substantia nigra pars compacta and retrorubral field decreases the harmaline-induced glutamate release in the rat cerebellum.

The aim of the present study was to examine the influence of a partial lesion of both the substantia nigra pars compacta (SNC) and retrorubral field (RRF) on the glutamatergic transmission in the cerebellum and tremor induced by harmaline in rats. 6-Hydroxydopamine (6-OHDA, 8 µg/2 µl) was injected unilaterally into the region of the posterior part of the SNC and RRF. Harmaline was administered in a dose of 30 mg/kg ip on the 8th day after the operation and the extracellular level of glutamate was measured by microdialysis in vivo in the cerebellar vermis. Harmaline induced glutamate release in the cerebellum. The lesion which encompassed 23-37% neurons in the anterior SNC, 52-54% in the posterior SNC and 47-55% in the RRF did not influence the basal extracellular glutamate level but decreased the harmaline-induced release of this neurotransmitter. Tremor evoked by harmaline was also visibly inhibited by the above lesion. The results of the present study seem to indicate that midbrain dopaminergic neurons influence glutamatergic transmission in the cerebellum which may be important for generation of the tremor induced by harmaline.

Depression in Parkinson's disease.

Depression is a frequent comorbid disorder of Parkinson's disease; however, little is known about its pathomechanisms. Although depression is an important factor negatively affecting the quality of life of parkinsonian patients, it often remains undiagnosed and therefore untreated. Furthermore, antidepressant therapy is problematic because of the need to combine antidepressant drugs with antiparkinsonian treatments. The present paper gives an overview of characteristic features of Parkinson's disease-associated depression, experimental studies on its animal models, potential mechanisms involved in its occurrence and possible strategies for treatment.

6-OHDA injections into A8-A9 dopaminergic neurons modelling early stages of Parkinson's disease increase the harmaline-induced tremor in rats.

The aim of the present study was to examine the influence of a unilateral 6-hydroxydopamine (6-OHDA)-induced partial lesion of both the substantia nigra pars compacta (SNc, A9) and retrorubral field (RRF, A8) on the tremor evoked by harmaline. 6-OHDA (8µg/2µl) was injected unilaterally into the region of the posterior part of the SNc and RRF. Harmaline was administered in a dose of 7.5mg/kg ip on the eighth day after the operation and tremor of forelimbs, head and trunk was measured. We found that the lesion increased intensity of the tremor induced by harmaline but did not influence its character. Stereological examination of the lesion extent revealed losses of dopaminergic (tyrosine hydroxylase-immunoreactive) neurons in the anterior (30%) and posterior (72%) SNc, as well as in RRF (72% on the average). Levels of dopamine and all its metabolites, as well as noradrenaline concentrations, were ipsilaterally moderately decreased in the caudate-putamen in the lesioned animals, however, dopamine and DOPAC in the anterior cerebellum were increased. In the caudate-putamen, the ipsi/contra ratio of dopamine level correlated negatively, while that of dopamine turnover positively with the tremor intensity. However, in the anterior cerebellum an inverse relationship was found. Moreover, this symptom correlated positively with the serotonin level and negatively with the 5-HIAA/serotonin ratio on the contralateral side of the posterior cerebellum. The present results seem to indicate that the modulation of dopaminergic and serotonergic transmissions by the lesion modelling early stages of Parkinson's disease may influence tremor triggered in the cerebellum.

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.

Glycogen synthase kinase 3ß and its phosphorylated form (Y216) in the paraquat-induced model of parkinsonism.

Parkinson's disease is a slowly progressing disease, due to a lesion of dopaminergic neurons in the substantia nigra and a dramatic loss of dopamine in the striatum. It is now accepted that several environmental agents including the herbicide paraquat (PQ) may contribute to its pathogenesis. However, till now nothing is known about the role of glycogen synthase kinase-3ß (GSK-3ß) in the PQ toxicity. Therefore, the aim of this study was to examine the influence of 37-week administration of PQ in rats on the immunohistochemically measured levels of the total GSK-3ß and its active, tyrosine 216 (pY216)-phosphorylated form in subcellular fractions of the midbrain with pons, as well as of the striatum. The present results revealed that the long-term PQ administration increased the levels of total and active forms of GSK-3ß in the midbrain with pons, whereas decreased them in the striatum. Examination of the lesion extent showed a decrease in the number of tyrosine-immunoreactive neurons in the substantia nigra pars compacta, ventral tegmental area, and locus coeruleus, as well as lower DOPAC/dopamine ratio and noradrenaline level in the striatum in rats treated with PQ. The long-term PQ administration disturbed also motor activity of rats. Summarizing, the present data indicate that the long-term exposure of rats to PQ, a commonly used herbicide, diversely alters levels of GSK-3ß in different brain structures, which may be associated with their vulnerability to its toxicity.