[The influence of acizolum to bioelements content in rat's blood plasma, parenchimal organs and brain]. / Vliianie atsizola na soderzhanie bioélementov v plazme krovi, parenkhimatoznykh organakh i golovnom mozge krys.

Zinc content in blood plasma and brain tissue of rats was studied by analytic mass-spectrometry with inductively coupled plasma. In control (saline-treated animal) zinc content in plasma was 3.6±1.4 mg/ml, in the liver - 12.5±2.5 mg/mg, in the spleen - 10.9±4.1 mg/mg, in the brain - 8.7±3.0 mg/mg. After a single intraperitoneal injection of zinc donator acizolum (24 mg/kg) zinc content decreased in all examined tissues, especially in brain. After a course of sequential acizolum injections (seven administrations during two weeks) essential elevation of zinc content in blood plasma and tissues investigated was detected. The maximal increase zinc concentration in blood plasma and liver was detected in 15 h after the last acizolum injections. Selen, calcium, copper and iron contents demonstrated a more complex behaviour. The obtained data suggest that prolonged acizolum administration has a significant impact on the bioelements content, and this should be taken into consideration when this zinc donator is used as a drug.

[The clinical features of Parkinson's disease in patients with mutations and polymorphic variants of GBA gene]. / Klinicheskie osobennosti bolezni Parkinsona u patsientov s mutatsiiami i polimorfnymi variantami gena GBA.

BACKGROUND: Mutations in the glucocerebrosidase gene (GBA) increase the risk of Parkinson's disease (PD) by 6-10 times in all populations and are associated with the early-onset of PD, development of cognitive impairment and presence of psychotic disorders. At the same time, polymorphic variants associated with the twofold increase in the risk of PD were also described in the GBA gene. AIM: To estimate the clinical features of PD in patients with mutations and polymorphic variants of the GBA gene. MATERIAL AND METHODS: Evaluation of motor, cognitive, emotional, psychotic and autonomic dysfunctions in patients with mutations (N370S, L444P) and polymorphic variants (E326K, T369M) in the GBA gene was performed using clinical scales. RESULTS: Patients with mutations (mGBA-PD), and with polymorphic variants (pGBA-PD) in the GBA gene were compared with the group of patients with sporadic PD (sPD). Compared to sPD, affective disorders (depression and anxiety) were more expressed in the mGBA-PD group (p=0.001) and the general GBA-PD group (p=0.001) assessed with Sheehan anxiety rating scale, in the pGBA-PD group (p=0.012) and the general GBA-PD group (p=0.05) assessed with the NPI, in the mGBA-PD (p=0.003), pGBA-PD (p=0.022), and general GBA-PD groups (p=0.001) assessed with the Hospital Anxiety and Depression scale (HADS 'A'), and in the pGBA-PD group (p=0.005) assessed with the HADS 'D'. Non-motor symptoms assessed with the PD-NMS were more expressed in the pGBA-PD patients (p=0.007) and in the total group with GBA-PD (p=0,014) compared to sPD. Cognitive impairment measured with MMSE was more marked in mGBA-PD patients (p=0.022). Differences in motor and non-motor clinical symptoms between pGBA-PD and mGBA-PD groups were not found. CONCLUSION: Thus, clinical features of non-motor symptoms were described both in carriers of GBA mutations and polymorphisms. Identification of the specific clinical phenotype of PD in carriers of GBA polymorphic variants is important due to their relatively high prevalence in PD patients.

[Mutation del 1,02kb in the CLN3 gene and extrapyramidal syndrome]. / Mutatsiya del 1,02 kb v gene CLN3 i sindrom ekstrapiramidnykh rasstroistv.

Mutations in the GBA and SMPD1 genes, which lead to the development of lysosomal storage diseases, are high risk factors for Parkinson's disease and dementia with Lewy bodies. We screened the mutations in the GALC and CLN3 genes in patients with Parkinson's disease and control subjects. A heterozygous CLN3 mutation (del 1.02 kb) carrier with clinical features of the unusual extrapyramidal syndrome was identified. A role of CLN3 mutations in the development of neurodegenerative disorders is discussed.

[SNCA rs356219 AND rs356165 VARIANTS ARE ASSOCIATED WITH PARKINSON'S DISEASE AND INCREASED ALPHA-SYNUCLEIN GENE EXPRESSION IN THE CD45(+)-BLOOD CELLS].

Impaired metabolism of alpha-synuclein (SNCA) and its aggregation are now implicated in the pathogenesis of Parkinson's disease (PD). Previous studies have found association between PD and gene locus, containing the SNCA gene. Meta-analysis have shown high significant association of single nucleotide polymorphisms (SNPs) rs356165 (A/G) and rs356219 (A/G) in the SNCA gene with PD. We genotyped these SNPs in 260 PD patients and 262 controls from north-western region of Russia. Alleles "G" of rs356165 and rs356219 were associated with increased risk of PD development. Linkage disequilibrium was shown between associated marker alleles. We studied the relationship between rs356165 and rs356219 and levels of mRNA SNCA and alpha-synuclein in CD45+ peripheral blood cells in drug-naive PD patients (n = 43) and controls (n = 39). Alleles "G" of rs356165 and rs356219 were associated with increased levels of SNCA expression (p = 0.046) and high alpha-synuclein levels (p = 0.039) in controls. Our data suggest that rs356165 and rs356219 variants might influence on PD development by upregulating SNCA expression.

Zinc Chloride and Zinc Acetate Injected into the Neostriatum Produce Opposite Effect on Locomotor Behavior of Rats.

Zinc chloride and zinc acetate solutions injected in a dose of 1 µg into the rostral neostriatum produced opposite effect on locomotor behavior of rats. Zink chloride disturbed conditioned avoidance and reduced spontaneous motor activity. Zink acetate virtually did not modify avoidance behavior and stimulated motor activity with elements of motor stereotypy. It was hypothesized that important factors here were the relationship between the effect and the level of metal released after salt dissociation and different reactivity of the synaptic substrate of the neostriatum to the presence of zinc ions.

Increased plasma oligomeric alpha-synuclein in patients with lysosomal storage diseases.

A link between lysosomal storage diseases (LSDs) and neurodegenerative disorders associated with accumulation of presynaptic protein alpha-synuclein has been shown. Particularly, Gaucher disease (GD) patients with a deficiency of the lysosomal enzyme glucocerebrosidase (GBA) and carriers of GBA mutations are at increased risk of Parkinson's disease (PD). It remains unclear whether this link is due to increased alpha-synuclein oligomerization. Here we show that level of oligomeric alpha-synuclein form, associated with PD development, is increased in plasma of GD patients (n=41, median=22.9pg/mL, range1.57-444.58pg/mL; controls (n=40, median=6.02pg/mL, range 1.05-103.14pg/mL, p<0.0001). This difference is absent in GD patients receiving enzyme replacement therapy (ERT) for more than 5 years. Moreover, the levels of alpha-synuclein oligomers in plasma are also higher in patients with other LSDs (Niemann-Pick type C, Krabbe disease, Wolman disease) compared to the median value in controls. Therefore, we suggest that mutations in the GBA gene and at least in several other LSDs genes may be associated with an increase in oligomeric alpha-synuclein in plasma. ERT applied for recovering of GBA functions in GD treatment might decrease formation of plasma oligomeric alpha-synuclein.

Effect of zinc chloride on picrotoxin-induced hyperkinesis depends on its concentration in solution injected into rat neostriatum.

The hyperkinekic effect (increase in spontaneous activity and development of choreomyoclonic hyperkinesis of the extremities and body) of picrotoxin injected into the rostral neostriatum of rats in a dose of 2 µg was reduced if the drug was injected together with ZnCl(2) in a concentration of 0.1 µg/µl. ZnCl(2) in a concentration of 1 µg/µl did not modulate the effects of picrotoxin, while in a concentration of 3 µg/µl it increased spontaneous motor activity in the open field test without affecting the symptoms of choreomyoclonic hyperkinesis.

Reduced content of α-synuclein in peripheral blood leukocytes of patients with LRRK2-associated Parkinson's disease.

Measurement of α-synuclein level in the peripheral blood was proposed as a diagnostic test for Parkinson's disease. However, the results of these studies remain contradictory, probably because the examined samples included patients with different etiology of Parkinson's disease. To verify this assumption we studied the levels of α-synuclein in peripheral blood leukocytes of patients with Parkinson's disease associated with mutations in the gene of leucine-rich kinase 2 (LRRK2). The mean α-synuclein level was significantly lower in patients with LRRK2-associated Parkinson's disease (N=8) than in patients with sporadic form of the disease (N=33; p<0.02) and in controls (N=18; p<0.05). On the other hand, we found no differences in the level of α-synuclein level between patients with sporadic form of the disease and controls. We hypothesize that the level of α-synuclein in the peripheral blood largely depends on the etiology of the disease and cannot be used as a universal diagnostic test for Parkinson's disease.

Screening for LRRK2 mutations in patients with Parkinson's disease in Russia: identification of a novel LRRK2 variant.

BACKGROUND AND PURPOSE: Mutations in LRRK2, encoding leucine-rich repeat kinase 2 (or Dardarin), cause autosomal dominant Parkinson's disease (AdPD) and are also found in sporadic PD (sPD). To investigate the frequency of LRRK2 mutations in a sample of Russian PD patients. METHODS: We sequenced the complete coding region of LRRK2 in 65 patients with AdPD and in 30 patients with sPD. Furthermore, in 20 patients with AdPD and in 159 patients with sPD we screened several common LRRK2 mutations (G2019S, R1441C/G/H, I2012T and I2020T). RESULTS: Five AdPD patients had the LRRK2 G2019S mutation (5.9%, 5/85). In addition, we discovered a novel LRRK2 variant V1613A in a family with a tremor dominant form of AdPD; this variant was not present in controls. We identified two patients with LRRK2 mutations in sPD: one with the G2019S mutation (0.5; 1/189) and another with the previously described R1441C mutation (0,5; 1/189). CONCLUSIONS: LRRK2 mutations are common amongst patients with PD in Russia. The results also show that the G2019S mutation is the most frequent. We identified one novel mutation in a functional region of LRRK2.

Intrastrial injection of magnesium after picrotoxin does not reduce motor hyperactivity in rats.

Magnesium chloride injected into rat rostral neostriatum before picrotoxin prevents motor hyperactivity induced by this GABAA receptor antagonist: prevents the increase in spontaneous motor activity in the open field test and reduces reproduction and duration of choreic hyperkinesia. Injection of magnesium 15 min after picrotoxin virtually did not modify the hyperkinetic effects of picrotoxin. The results indicate the role of calcium processes in the appearance of strial picrotoxin hyperactivity and principal possibility of its correction with magnesium only before the cascade of respective calcium ionic transformations is triggered in the brain.

Magnesium ions prevent the development of hyperkinesia evoked by administration of picrotoxin into the rat neostriatum.

Chronic experiments on rats were performed to study the effects of daily administration of the GABA(A) receptor antagonist picrotoxin (2 microg) into the rostral neostriatum in rats. Picrotoxin was injected in 1 microl of sterile apyrogenic physiological saline or in 1 microl of 1.0 M or 1.5 M MgCl(2); control animals received physiological saline or 1.0 M MgCl(2). Picrotoxin given in physiological saline induced impairments in conditioned reflex (avoidance) and free motor behavior and produced clear stereotypical imperative movements in the form of choreomyoclonic hyperkinesia of the paws, head, and trunk. These motor impairments resembled the manifestaitons of basal ganglia dysfunction typical of Huntingdon's chorea in humans. Magnesium ions prevented both the development of hyperkinesia and impairments of learned behavior. Given that magnesium is a nonspecific calcium channel antagonist, it can be suggested that one of the leading mechanisms of development of hyperkinesias is impairment of calcium homeostasis in striatal neurons.

Verapamil aggravates hyperkinesia produced by intrastriatal administration of picrotoxin in rats.

The effects of slow Ca2+ channel blocker verapamil and Ca2(+)-binding agent EDTA were studied on the model of choreic hyperkinesia induced by chronic intrastriatal microinjections GABAA receptor antagonist picrotoxin. Normal and pathological movements were recorded. The test preparations facilitated the effect of picrotoxin on spontaneous and learned behavior. They exerted a permissive effect on picrotoxin-induced hyperkinesia: increase reproducibility and duration of hyperkinesia and decrease the latency of this reaction. Our results indicate that Ca2+ channels are involved into the development and progression of hyperkinesia.

Motor behavior in rats after separate and combined administration of GABAergic agents into the neostriatum.

Chronic experiments were performed on rats to study the main inhibitory transmitter system of the neostriatum--the GABAergic system--in the regulation of normal and pathological motor behavior. Studies addressed the effects of separate and combined administration of GABA (45 microg) and A-type receptor antagonists, i.e., picrotoxin (1 microg) and bicuculline (5 microg), into the neostriatum on the performance by rats of spontaneous (including pathological) and conditioned reflex motor behavior (active avoidance reflex in a shuttle box). Agents were injected in a volume of 1 microl daily for three weeks; control animals received physiological saline. Activation of the GABAergic system of the neostriatum had no significant effect on behavior. Conditioned reflex avoidance behavior was impaired throughout the period of bicuculline administration (there was no significant change in spontaneous behavior); this recovered after the course of microinjections finished. Picrotoxin produced smaller negative effects on performance of the reflex, though rats showed clear imperative movements in the form of choreomyoclonic hyperkinesia. Simultaneous administration of picrotoxin and GABA into the neostriatum produced less hyperkinesia; administration of picrotoxin and bicuculline altered the nature of hyperkinesia. The importance of the GABAergic system for the antihyperkinetic activity of the basal ganglia is discussed, and it is suggested that the GABA-A subsystem is of critical importance in these functions.

Neostriatal glutamatergic system is involved in the pathogenesis of picrotoxin-induced choreomyoclonic hyperkinesis.

Administration of dizocilpine (MK-801, noncompetitive antagonist of NMDA glutamate receptors) into the neostriatum decreased the reproducibility and duration of hyperkinesis in rats induced by repeated microinjections of GABA(A) receptor antagonist picrotoxin. By contrast, glutamate potentiated the hyperkinetic and convulsive effect of picrotoxin and promoted the inhibition of conditioned avoidance response. Our results indicate that the striatal glutamatergic system is involved in the development of locomotor and cognitive disorders associated with deficiency of the neostriatal GABAergic system and playing a role in the pathogenesis of Huntington's chorea.

Structural and topical bases of picrotoxin-induced choreomyoclonic hyperkinesis.

Multiple bilateral microinjections of 1 g picrotoxin (blocker of GABAA-receptor chlorine channels) into the rostral neostriatum cause choreomyoclonic hyperkinesis of the paws, head, and body in rats. The most pronounced hyperkinesis was observed when the cannula was localized in the most anterior regions of the rostral neostriatum with irradiation of its action to the white matter (corpus callosum) and when the cannulas were located asymmetrically in the right and left striatum.

Analysis of striopallidal interactions in the control of avoidance behavior.

The effects of chronic administration of phenamine (15 microg) into the neostriatum and bicucculline (5 microg) in the globus pallidus of rats on the performance of avoidance behavior in a shuttle box were studied. Agents were injected daily for three weeks, and control animals received physiological saline. Activation of the dopaminergic system of the neostriatum compensated for decreases in avoidance behavior due to surgical trauma, increased the intensity of spontaneous activity in the "open field," and induced motor stereotypy. Administration of bicucculline into the globus pallidus produced sharp worsening of conditioned reflex avoidance behavior throughout the treatment period (spontaneous movement activity showed no significant change), with recovery only occurring two weeks after the last microinjection. Simultaneous treatment with phenamine into the neostriatum and bicucculline into the globus pallidus resulted in compensation for the inhibitory effects of bicucculline. Several phases were observed in behavioral changes, apparently reflecting the dynamics of interactions between the neo- and paleostriatum. The significance of the coordination of intrastriatal functions for the regulatory activity of the basal ganglia is discussed.

Food-procuring behavior of rats under the conditions of chronic activation and blockade of the neostriatal dopaminergic system.

Using the technique of intracerebral microinjections, the features of food-procuring behavior (the realization of a situational instrumental conditioned reflex in a Skinner box) were investigated in rats in experiments under the conditions of a pharmacological influence on the dopaminergic system of the neostriatum. Amphetamine in a dose of 15 and 45 micrograms and haloperidol in a dose of 5 mg were injected daily over the course of three weeks bilaterally into the rostral division of the neostriatum. A amphetamine dose of 15 micrograms was ineffective, but a increase in motor hyperactivity, a behavioral stereotypy, and a substantial acceleration of food-procuring movements were observed against the background of the injection of 45 micrograms of amphetamine. Stimulation of the dopaminergic system of the neostriatum qualitatively altered the behavioral strategy in animals with an initially low level of realization of the reflex; this fostered a stable activation of the instrumental skill enduring even after the cessation of the microinjections. Chronic blockade of dopamine receptors by haloperidol induced the reverse effect.

Behavior of rats during chronic activation and blockade of the neostriatal opiate system.

The effects of daily microinjections (MI), over of three weeks, bilaterally into the rostral striatum, of morphine, promedol, native leu-enkephalin and its synthetic tetrapeptide analogs were studied in experiments on rats. Naloxone was used as an antagonist. An active avoidance conditioned reflex was developed preliminary in a shuttle box. A decrease in the accuracy of the realization and an increase in the latent period of the reflex were observed after the first MI of morphine and enkephalins. The effect of the most stable aminated ornithine-containing tetrapeptides proved to be the strongest. A search stereotypy and increased motoric activity were recorded in the rats during the development of the chronic effects of the activators of the opiate system. A clear correlation was not found between the motor and conditioned reflex shifts. The blockade of the opiate receptors with naloxone did not lead to substantial changes in behavior. The data obtained confirm the current hypothesis regarding the important role of the enkephalinergic system of the neostriatum in the regulation of complex forms of behavior and its close functional association with the dopaminergic system.

Influence of glutamate on the content and metabolism of dopamine in the nigrostriatal system of rats distinguished by capacity for learning.

The effect of repeated (over the course of nine days) intrastriatal microinjections of glutamate (5 or 0.5 micrograms in 0.75 microliters of physiological solution daily) were investigated in 42 male Sprague-Dawley rats. Twenty-nine rats were preliminarily trained in a Skinner box using food reinforcement. It was demonstrated that the administration of glutamate to rats not subjected to training increases the content of homovanillic acid in the striatum. A similar influence in rats that are capable of learning leads to an increase in the content of dopamine and a decrease in the level of homovanillic acid in this nucleus, while it does not induce changes in the biochemical indicators under investigation in those rats that are incapable of learning. Microinjections of glutamate also do not alter the capacity for learning in any of the groups of animals. The possible causes for the different influence of intrastriatal microinjections of glutamate on the activity of the nigrostriatal system of rats differing by capacity for learning are discussed.

The influence of chronic activation and blockade of the dopamine- and enkephalinergic systems of the neostriatum on conditioned reflex behavior and dopamine metabolism in the rat nigrostriatal system.

The effects of daily injections, over the course of 14 days, of 45 micrograms of phenamine, 5 micrograms of haloperidol and naloxone, 15 micrograms of leu-enkephalin and its analog, a tetrapeptide, into the rostral neostriatum have been studied. The chronic stimulation of the dopaminergic system of the striatum induced facilitation of the realization of an active avoidance conditioned reflex, and stimulated exploratory stereotypy, while its blockade led to suppression of conditioned reflex activity against the background of a clearcut rigid akinetic syndrome. The microinjections of leu-enkephalin and naloxone did not substantially alter the behavior, but the injection of the tetrapeptide was accompanied by changes in behavior, with symptoms of catalepsy and hyperkinesia. Injections of phenamine and haloperidol were accompanied by a decrease in the content of dopamine in the striatum and an increase in the level of DOPAC; the injections of enkephalin and naloxone induced changes of the reverse order. The possible causes of the noncorrespondence of the behavioral and neurochemical shifts in the presence of a direct chronic pharmacological action on the mediator of the neostriatum.