Rat Midbrain Research Articles

The effects of antipsychotics on β‐catenin, glycogen synthase kinase‐3 and dishevelled in the ventral midbrain of rats

Protein kinase B and glycogen synthase kinase-3 have been identified as susceptibility genes for schizophrenia and altered protein and mRNA levels have been detected in the brains of schizophrenics post-mortem. Recently, we reported that haloperidol, clozapine and risperidone alter glycogen synthase kinase-3 and beta-catenin protein expression and glycogen synthase kinase-3 phosphorylation levels in the rat prefrontal cortex and striatum. In the current study, beta-catenin, adenomatous polyposis coli, Wnt1, dishevelled and glycogen synthase kinase-3 were examined in the ventral midbrain and hippocampus using western blotting. In addition, beta-catenin and GSK-3 were examined in the substantia nigra and ventral tegmental area using confocal and fluorescence microscopy. The results indicate that repeated antipsychotic administration results in significant elevations in glycogen synthase kinase-3, beta-catenin and dishevelled-3 protein levels in the ventral midbrain and hippocampus. Raclopride causes similar changes in beta-catenin and GSK-3 in the ventral midbrain, suggesting that D2 dopamine receptor antagonism mediated the changes observed following antipsychotic administration. In contrast, amphetamine, a drug capable of inducing psychotic episodes, had the opposite effect on beta-catenin and GSK-3 in the ventral midbrain. Collectively, the results suggest that antipsychotics may exert their beneficial effects through modifications to proteins that are associated with the canonical Wnt pathway.

Glutaric Acid Administration Impairs Energy Metabolism in Midbrain and Skeletal Muscle of Young Rats

A genetic mice model of glutaric acidemia type I (GAI) has recently been developed, however affected animals do not develop the striatal damage characteristic of patients with this disorder. Therefore, the initial aim of the present work was to induce high glutaric acid (GA) concentrations in rat brain similar to those found in GAI patients through subcutaneous injection of GA. High brain GA concentrations (up to 0.60 micromol/g congruent with 0.60mM) were achieved by a single subcutaneous injection of saline-buffered GA (5 micromol/g body weight) to Wistar rats of 7-22 days of life. GA brain levels were about 10-fold lower than in plasma and 5-fold lower than in skeletal and cardiac muscles, indicating that the permeability of the blood brain barrier to GA is low. We also aimed to use this model to investigate neurochemical parameters in the animals. Thus, we evaluated the effect of this model on energy metabolism parameters in midbrain, in which the striatum is localized, as well as in peripheral tissues (skeletal and cardiac muscles) of 22-day-old rats. Control rats were treated with saline in the same volumes. We verified that CO2 production from glucose was not altered in midbrain of rats treated with GA, indicating a normal functioning of the tricarboxylic acid cycle. Creatine kinase activity was also not changed in midbrain, skeletal and cardiac muscles. In contrast, complex I-III activity of the respiratory chain was inhibited in midbrain (25%), while complexes I-III (25%) and II-III (15%) activities were reduced in skeletal muscle, with no alterations found in cardiac muscle. These data indicate that GA administration moderately impairs cellular energy metabolism in midbrain and skeletal muscle of young rats.

Metabolism and Receptor Binding of Serotonin in Brain Structures During Performance of a Conditioned Passive Avoidance Response

The levels of serotonin and its metabolite 5-hydroxyindoleacetic acid, monoamine oxidase activity, and the specific binding of the radioligand [3H]serotonin were measured in the prefrontal cortex, striatum, amygdaloid complex, hippocampus, and periacqueductal gray matter of the midbrain in rats at different time points after training to a conditioned passive avoidance reaction. Changes in serotoninergic activity were found to be characteristic only for the process of reproducing the conditioned reaction. The metabolism and serotonin receptor binding in these brain structures did not change immediately after the training period or one day after this, or in conditions of failure to reproduce the reaction because of amnesia, or in untrained animals. The involvement of the brain serotoninergic system in the process of performing the conditioned reaction was found to demonstrate a spatial-structural selectivity: the metabolism and receptor binding of serotonin changed in the amygdaloid complex, periacqueductal gray matter, and the striatum, while no changes were seen in the hippocampus or prefrontal cortex. All three brain structures showed decreases in [3H]serotonin receptor binding of. Serotonin levels did not change, though the amygdaloid complex and periacqueductal gray matter showed increases in oxidative deamination of serotonin and increases in the active transport of the metabolite, while the striatum showed decreases in serotonin catabolism. The differences in the catabolism of this neurotransmitter suggest that the decrease in serotonin receptor binding in these brain structures depends on different synaptic processes--presynaptic in the striatum and postsynaptic in the amygdaloid complex and periacqueductal gray matter. It is concluded that the decrease in the functional activity of serotoninergic transmission in the amygdaloid complex and periacqueductal gray matter is one of the mechanisms involved in activation of the emotiogenic system triggering the process of reproduction of the memory trace.

Differential survival patterns among midbrain dopaminergic cells of MPTP-treated monkeys and 6OHDA-lesioned rats

We explore the patterns of survival among dopaminergic cells of the midbrain in MPTP-treated macaque monkeys and 6OHDA-lesioned Sprague-Dawley rats. For the monkeys, animals were injected intramuscularly with MPTP for 8 days consecutively and then allowed to survive for 21 days. For the rats, 6OHDA was injected into the midbrain and then allowed to survive for either 7, 28 or 84 days. Brains were processed for tyrosine hydroxylase (TH) and calbindin immunocytochemistry to label populations in the ventral and dorsal tiers of midbrain dopaminergic cells. In monkeys, while there was a decrease in the TH+ cell number in the ventral tier of MPTP-treated cases (65%), there was an overall increase (22%) in the TH+ and calbindin+ cell number in the dorsal tier. Double labelling studies indicate that approximately 50% of TH+ cells of the dorsal tier contain calbindin also. In rats, there was a decrease in TH+ cell number in the ventral tier of 6OHDA-lesioned cases (97%), and to a lesser extent, in the TH+ and calbindin+ cell number in the dorsal tier ( approximately 40%). In conclusion, we show a surprising increase in TH+ and calbindin+ cell number in the dorsal tier in response to MPTP insult; such an increase was not evident after 6OHDA insult. We suggest that the increase in antigen expression relates to the dopaminergic reinnervation of the striatum in MPTP-treated cases. We also suggest that the greater loss of dopaminergic cells in the ventral tier when compared to the dorsal tier relates to glutamate toxicity.

The role of 5-HT2A receptor antagonism in amphetamine-induced inhibition of A10 dopamine neurons in vitro

The role of the 5-HT2A receptor in modulating amphetamine-induced inhibition of dopamine neuronal firing in A9 and A10 was investigated in rat midbrain slices. The antipsychotic drugs olanzapine and clozapine more potently reversed the amphetamine-induced inhibition in A10 neurons compared to A9 neurons. Risperidone (0.03 and 0.1 μM) reversed amphetamine-induced inhibition of firing activity similarly in A9 and A10. The dopamine D2 receptor antagonist (−)sulpiride (0.05 and 1 μM) reversed the amphetamine (10 μM)-induced inhibition of firing activity in A9 and A10 neurons. The selective 5-HT2A receptor antagonist MDL100907 (0.05 μM), strongly enhanced the reversal of amphetamine-induced inhibition by (−)sulpiride in A10, but its effectiveness was much smaller in A9 dopamine neurons.We conclude that 5-HT2A receptor antagonism enhanced reversal of amphetamine-induced inhibition by dopamine D2 antagonism in A10, suggesting that dopamine D2 receptor antagonism combined with 5-HT2A receptor antagonism may play a role in antipsychotic drug atypicality.

Inhibition of cell differentiation by quinolones in micromass cultures of rat embryonic limb bud and midbrain cells

Micromass cultures (MMC) of rat embryonic limb bud (LB) and midbrain (CNS) cells were applied to compare the developmental toxicity of three quinolone antimicrobials: norfloxacin (Nor), enrofloxacin (Enr) and ciprofloxacin (Cip). Cultures were exposed for 5 days to seven concentrations of drugs. Cytotoxicity was assessed by quantifying neutral red uptake; differentiation—by quantifying alcian blue uptake (LB) or by image analysis of Gill’s haematoxylin stained foci (CNS). Both, LB and CNS cultures showed dose-dependent reduction in total cell number and differentiation. To distinguish specific effect on differentiation, IC 50 for proliferation ( P) and differentiation ( D) were calculated and P/ D ratios were compared. In LB cultures all three drugs were cytotoxic ( P/ D ratios were ⩽1). In CNS cultures P/ D ratios were ⩾1 (up to 2.7 for Nor, up to 4.4 for Enr and up to 16 for Cip) what can suggest specific action on differentiation. Ciprofloxacin was the most toxic and CNS cells were more sensitive than LB. The ranges of IC 50-D values (μg/ml) were as follows: Nor (79–14), Enr (127–179), Cip (91–101) in LB cultures; Nor (22–52), Enr (38–91), Cip (3–17) in CNS cultures. With one exception (Cip in CNS culture) all drugs were classified as weak embryotoxic.

Tissue‐specific expression of tryptophan hydroxylase mRNAs in the rat midbrain: anatomical evidence and daily profiles

Serotonin (5-HT) is involved in both photic and non-photic synchronization of the mammalian biological clock located in the suprachiasmatic nuclei (SCN). We have previously demonstrated that tryptophan hydroxylase protein (TPH), the rate-limiting enzyme of 5-HT synthesis, shows circadian rhythmicity in the pathways projecting from the raphe nuclei to the intergeniculate leaflets of the thalamus on one hand, and to the SCN on the other hand. In this study, we investigate whether the circadian rhythmicity in TPH protein could result from the rhythmic expression of tph gene in the raphe nuclei. We thus cloned specific tph1 and tph2 partial cDNAs and assessed the daily profiles of TPH mRNA levels by in situ hybridization in the rat raphe nuclei. Our results demonstrate that: (i) tph2 gene is exclusively expressed in the raphe nuclei, whereas tph1 gene is expressed in the pineal gland; (ii) under light-dark cycle (LD), TPH2 mRNA levels present daily variation within both median and dorsal raphe nuclei; (iii) under constant darkness TPH2 mRNA levels in both nuclei exhibit the same variation reported under LD cycle. These data show that the circadian 5-HT synthesis within the serotonergic neurons projecting to the circadian system might be explained by the rhythmic transcription of the tph2 gene in raphe nuclei. Taking our result with previous data into consideration, we further suggest that 5-HT synthesis and release within the circadian system could be directly or indirectly under the control of the SCN.

Postsynaptic actions of substance P on rat periaqueductal grey neurons in vitro

The postsynaptic actions of substance P on rat midbrain periaqueductal grey (PAG) neurons were examined using whole-cell patch-clamp recordings in brain slices. Substance P produced an inward current in a subpopulation (60%) of PAG neurons. The substance P induced current was concentration dependent (EC 50=27 nM) and was reduced by the NK1, NK2 and NK3 antagonists L-732,138 (20 μM), GR 159897 (3 μM) and SB 218795 (3 μM). The selective NK1, NK2 and NK3 agonists [Sar 9,Met(O 2) 11]-Substance P (100 nM), GR 64349 (300–500 nM) and senktide (300 nM) also produced inward currents in subpopulations of neurons. A greater proportion of substance P-sensitive neurons (70%) than substance P-insensitive neurons (31%) responded to the μ/δ opioid agonist met-enkephalin (10 μM). Substance P reduced the outward current produced by met-enkephalin. The reversal potential of the substance P induced current varied from −5 mV to below −140 mV in the absence of met-enkephalin, and was −105 mV in the presence of met-enkephalin. These results indicate that substance P acts via NK1, NK2 and NK3 receptors to excite subpopulations of opioid-sensitive and insensitive PAG neurons by increasing a non-selective cation conductance and by reducing a K + current. In addition, substance P has anti-opioid actions that are largely mediated by a reduction in the opioid induced K + current.

Expression of ORL 1 mRNA in some brain nuclei in neuropathic pain rats

The present study was designed to investigate changes of opioid receptor like 1 receptor (ORL 1, NOP) mRNA expression in some pain-related brain nuclei of neuropathic pain rats using in situ hybridization technique. Nociceptin/orphanin FQ (N/OFQ), the endogenous ligand of ORL 1, plays an important role in neuropathic pain through its receptor. There are ORL 1 mRNA expression in the nucleus of raphe magnus (NRM), ventrolateral periaqueductal gray (vlPAG) and dorsal raphe nucleus (DRN) of rat mesencephalon. In the sciatic nerve chronic constriction injury (CCI)-induced neuropathic pain model, a significant increase of ORL 1 mRNA expression was observed in these three regions on the 7th day after operation, and the changes lasted for 2 weeks. The result indicated that ORL 1 synthesis was increased in NRM, vlPAG and DRN of neuropathic pain rats, suggesting that ORL 1 was involved in nociceptive transmission of neuropathic pain.

GABAB receptor modulators potentiate baclofen‐induced depression of dopamine neuron activity in the rat ventral tegmental area

2,6-Di-tert-butyl-4-(3-hydroxy-2,2-dimethyl-propyl)-phenol (CGP7930) is a recently reported positive allosteric modulator of gamma-aminobutyric acid (GABA)(B) receptors. In this study, we assessed the ability of CGP7930 to modulate the baclofen-induced depression of dopamine (DA) neuron activity via the activation of GABA(B) receptors in the ventral tegmental area in rat midbrain slices. The selective GABA(B) receptor agonist, baclofen, depressed the spontaneous firing rate of DA neurons in a concentration-dependent manner (EC50 = 0.27 microM, n = 11). CGP7930 (30 microM) significantly (P < 0.05) shifted the baclofen concentration-response curve to the left (EC50 = 0.15 microM, n = 5). The effects of baclofen alone or baclofen coapplied with CGP7930 were fully blocked by 1 microM (2S)-3-[[(1S)-1-(3,4-dichloropheny)ethyl]amino-2-hydroxypropyl] (phenylmethyl) phosphinic acid (CGP55845), a potent and selective GABA(B) receptor antagonist. In similar experiments, N-[3,3-diphenylpropyl]-alpha-methylbenzylamine (fendiline) (30 or 50 microM), a compound shown to potentiate GABA(B) receptor-mediated cortical hyperpolarisation, also significantly enhanced the inhibitory effect of baclofen. It is therefore concluded that the recently reported GABA(B) receptor modulators, CGP7930 and fendiline, can enhance GABA(B) receptor-mediated depression of DA neuronal activity. This finding suggests a therapeutic potential for GABA(B) potentiators for the treatment of diseases associated with a hyperfunctional mesocorticolimbic system.

Acanthopanax senticosus Harms as a prophylactic for MPTP-induced Parkinson's disease in rats

The aim of this study was to determine whether Acanthopanax senticosus Harms (ASH) offers protection against Parkinson's disease (PD) and its related depressive behaviors in rats administered 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP). We examined how ASH affected the MPTP-induced loss of tyrosine hydroxylase (TH)-positive neurons in the midbrain of rats. Extract from the stem bark of ASH prepared with hot water was dissolved in distilled water. Rats were then orally administered ASH (250 mg/kg) once a day for 2 weeks before ASH administration plus an intraperitoneal injection of MPTP (20 mg/kg). The pole test and catalepsy test were used to evaluate the effects of ASH administration on bradykinesia and depressive behaviors in the PD model of rats given MPTP for 2 weeks. Treatment with ASH for 2 weeks resulted in prophylactic effects on MPTP-induced Parkinsonian bradykinesia and catalepsy. Immunohistochemistical analysis using TH antibody showed that ASH provided cytoprotective effects against MPTP-induced loss of dopamine (DA) cells. The present results suggest that it may be possible to use ASH for the prevention of nigral degenerative disorders, e.g., PD with depression, caused by exposure to toxic substances.

Actions of methylphenidate on dopaminergic neurons of the ventral midbrain

Methylphenidate has been suggested to exert its therapeutic effect mainly by blocking the dopamine transporter. In spite of the importance of this interaction, no detailed information is available yet on its actions on single dopaminergic neurons. We examined the effects of methylphenidate on dopaminergic neurons using electrophysiological recordings from rat midbrain slices. Methylphenidate inhibited spontaneous firing and caused a membrane hyperpolarization in current clamp or an outward current in voltage clamp. These effects were antagonized by the D(2) receptor antagonist sulpiride. An acute dopamine-depleting treatment of the slices with the dopa-decarboxylase inhibitor carbidopa significantly reduced the effects of methylphenidate. This drug potentiated, in a concentration-dependent manner, cellular responses to exogenous dopamine application. Our electrophysiological data are consistent with the hypothesis that methylphenidate inhibits dopamine transporter and suggest that the depression of firing is mediated by the release of newly synthesized dopamine which accumulates extracellularly due to inhibition of its reuptake.

Trace Amines Depress GABAB Response in Dopaminergic Neurons by Inhibiting G-βγ-Gated Inwardly Rectifying Potassium Channels

Trace amines (TAs) are present in the central nervous system in which they up-regulate catecholamine release and are implicated in the pathogenesis of addiction, attention-deficit/hyper-activity disorder, Parkinson's disease, and schizophrenia. By using intracellular and patch-clamp recordings from dopaminergic cells in the rat midbrain slices, we report a depressant postsynaptic action of two TAs, beta-phenylethylamine (beta-PEA) and tyramine (TYR) on the GABA(B)-mediated slow inhibitory postsynaptic potential and baclofen-activated outward currents. beta-PEA and TYR activated G-proteins, interfering with the coupling between GABA(B) receptors and G-betagamma-gated inwardly rectifying potassium channels. This is the first demonstration that beta-PEA and TYR depress inhibitory synaptic potentials in neurons of the central nervous system, supporting their emerging role as neuromodulators.

A role for p38 mitogen-activated protein kinase in the regulation of the serotonin transporter: evidence for distinct cellular mechanisms involved in transporter surface expression.

The serotonin transporter (SERT) is regulated by various signaling mechanisms that may operate to maintain appropriate levels of synaptic serotonin (5-HT). We demonstrate that one of the mitogen-activated protein kinases (MAPKs), p38 MAPK, regulates SERT. Treatment of rat midbrain synaptosomes with p38 MAPK-specific inhibitors, PD169316 [4-(4-fluorophenyl)-2-(4-nitrophenyl)-5-(4-pyridyl)-1H-imidazole] or SB203580 [4-(4-fluorophenyl)-2-(4-methylsulfinylphenyl)-5-(4-pyridyl)-1H-imidazole], reduced 5-HT uptake. An additive SERT inhibition by PD169316 and beta-phorbol 12-myristate 13-acetate (beta-PMA) indicated the involvement of a protein kinase C (PKC)-independent MAPK pathway. Kinetic studies indicated a significant decrease in the transport capacity (V(max)) after PD169316 treatment of synaptosomes. Biotinylation studies showed reduced SERT proteins in the plasma membrane of synaptosomes after p38 MAPK inhibition and PKC activation. Phosphorylation studies using synaptosomes revealed decreased SERT phosphorylation by PD169316 but increased phosphorylation by beta-PMA. d-Amphetamine enhanced SERT basal phosphorylation and PD169316 blocked this effect. SERT interaction with protein phosphatase 2A catalytic subunit and syntaxin 1A decreased after PD169316 or beta-PMA treatment of synaptosomes. In synaptosomes, PKC activation but not p38 MAPK inhibition resulted in SERT redistribution from cholesterolrich lipid raft fractions to nonlipid raft fractions. The presence of phospho-p38 MAPK in synaptosomes and human embryonic kidney 293 (HEK-293) cells suggested the presence of constitutively active p38 MAPK in these preparations. Cotransfection of HEK-293 cells with SERT and a constitutively active form of MAP kinase kinase 3b(E) [MKK3b(E)] increased 5-HT transport, and RNA interference targeted to p38 MAPK inhibited 5-HT uptake, confirming the involvement of active p38 MAPK in SERT expression. Although PD169316 inhibited SERT insertion to the plasma membrane, beta-PMA increased SERT internalization in HEK-293 cells. Together, these results indicate a distinct role of p38 MAPK in SERT regulation.

Effect of Sesamin in Acanthopanax senticosus HARMS on Behavioral Dysfunction in Rotenone-Induced Parkinsonian Rats

The aim of this study was to determine whether sesamin, a component from Acanthopanax senticosus HARMS (ASH) pharmacologically offers protection against Parkinson's disease (PD) and its related depressive behavior in rats administered rotenone. We also examined how sesamin affected the rotenone-induced loss of tyrosine hydroxylase (TH) or glial cell line-derived neurotrophic factor (GDNF)-positive neurons in the midbrain of rats. Rats were orally administered sesamin (3, 30 mg/kg) once a day for 2 weeks before an intraperitoneal injection of rotenone (2.5 mg/kg). The pole test and catalepsy test were used to evaluate the effects of sesamin administration on bradykinesia and depressive behaviors in the PD model of rats given rotenone for 5 weeks. Those effects were compared with the ASH administrated group (250 mg/kg). Treatment with sesamin for seven weeks resulted in prophylactic effects on rotenone-induced parkinsonian bradykinesia and catalepsy, and the effects were equivalent to ASH effects. Immunohistochemistical analysis using TH or GDNF antibody showed that sesamin provided cytoprotective effects against rotenone-induced loss of DA cells. The results suggest that it may be possible to use the ASH and sesamin for the prevention of nigral degenerative disorders, e.g., PD with depression, caused by exposure to pesticide or environmental neurotoxins in general.

Supervivencia in vitro de neuronas dopaminérgicas hibernadas durante siete días

The use of fresh foetal tissue in neurotransplants entails considerable problems of logistics that limit its clinical applicability, something that can be resolved by the development of optimal tissue storage procedures that do not affect in vivo viability and survival of dopamine. AIMS. To determine whether 7 days' hibernation affects the survival of mesencephalic tissue in vitro, and to compare it to fresh tissue. The midbrains of rats were hibernated for 1, 3, 5 and 7 days at 4 degrees C. A cellular suspension was prepared for culture throughout a 7-day period. The number of TH+ cells present in the fresh and hibernated cultures was determined. The morphology of the hibernated and cultured dopaminergic neurons was very similar to that of the fresh cells. Comparing the viability of the hibernated and fresh cells did not reveal any significant differences. No significant differences between the numbers of TH+ neurons were observed at any of the hibernation times. The lowest rate of TH+ cell survival was reached at seven days' hibernation. Significant differences (p < 0.05) were found between the number of TH+ neurons for fresh and hibernated tissue. Hibernation at 4 degrees C for up to five days guarantees the survival of TH+ cells in vitro, but it is affected by longer times. This procedure could be considered useful for preserving human tissue in clinical transplant applications. These results refer to in vitro conditions; therefore, studies must be conducted to investigate the survival and functionality of hibernated and transplanted neurons in animal models to enable us to evaluate its applicability in neurorestorative therapy.

Reduction in 5-HT1A receptor density, 5-HT1A mRNA expression, and functional correlates for 5-HT1A receptors in genetically defined aggressive rats

The present experiments tested the hypothesis that one of the critical mechanisms underlying genetically defined aggressiveness involves brain serotonin 5-HT1A receptors. 5-HT1A receptor density, the receptor mRNA expression in brain structures, and functional correlates for 5-HT1A receptors identified as 8-OH-DPAT-induced hypothermia and lower lip retraction (LLR) were studied in Norway rats bred for 59 generations for the lack of aggressiveness and for high affective aggressiveness with respect to man. Considerable differences between the highly aggressive and the nonaggressive rats were shown in all three traits. A significant decrease in B(max) of specific receptor binding of [3H]8-OH-DPAT in the frontal cortex, hypothalamus, and amygdala and a reduction in 5-HT1A receptor mRNA expression in the midbrain of aggressive rats were found. 5-HT1A receptor agonist 8-OH-DPAT (0.5 mg/kg, i.p.) produced a distinct hypothermic reaction in nonaggressive rats and did not affect significantly the body temperature in aggressive rats. Similar differences were revealed in 8-OH-DPAT-induced LLR: LLR was expressed much more in nonaggressive than in aggressive animals. Additionally, 8-OH-DPAT (0.5 mg/kg i.p.) treatment significantly attenuated the aggressive response to man. The results demonstrated an association of aggressiveness with reduced 5-HT1A receptor expression and function, thereby providing support for the view favoring the idea that brain HT1A receptor contributes to the genetically defined individual differences in aggressiveness.

Administration of Myo-inositol Plus Ethanolamine Elevates Phosphatidylethanolamine Plasmalogen in the Rat Cerebellum

Plasmalogens are ether-linked phospholipids highly abundant in nervous tissue. Previously we demonstrated that acute administration of myo-inositol (myo-Ins) + [2-(13)C] ethanolamine ([2-(13)C]Etn) significantly elevated phosphatidylethanolamine plasmalogen (PlsEtn) in rat whole brain. Current experiments investigated the effects of acute myo-Ins+[2-(13)C]Etn administration on [PlsEtn] and the biosynthesis of new Etn lipids using NMR spectroscopy in rat cerebral cortex, hippocampus, brainstem, midbrain and cerebellum. Treated rats received a single dose of myo-Ins + [2-(13)C]Etn and controls received saline rather than myoIns. Data reveal that the cerebellum is the brain region most affected by treatment, which resulted in a 22% increase in [PlsEtn] and 89% increase in newly synthesized Etn lipids relative to controls (P < 0.05). Furthermore, the cerebellar PlsEtn/phosphatidylethanolamine ratio and molar percentage of PlsEtn were significantly elevated by 12% and 8%, respectively (P < 0.05). These data suggest that myo-Ins influences Etn lipid metabolism in brain, particularly in the cerebellum where there is a stimulation in the biosynthesis of new Etn lipids with a preference towards PlsEtn.

Histochemical study of effects of weak electromagnetic field on structures of the rat midbrain

Histochemical activity of the mitochondrial enzyme cytochrome oxidase (CO) in auditory and non-auditory structures of rat midbrain was studied after exposure to low electromagnetic field (EMF). An increase of the enzyme activity was shown in several midbrain structures of the experimental animals.

DOPAMINE CELL MORPHOLOGY AND GLIAL CELL HYPERTROPHY AND PROCESS BRANCHING IN THE NIGROSTRIATAL SYSTEM AFTER STRIATAL 6-OHDA ANALYZED BY SPECIFIC STEROLOGICAL TOOLS

Morphological changes in the dopamine neurons and glial cells of the rat midbrain ascending dopamine pathways were investigated after a partial lesion induced by unilateral striatal injection of a small dose of 6-hydroxydopamine (6-OHDA). Fourteen days after lesion, animals showed contralateral rotation induced by apomorphine injection. After behavioral analysis, rats were killed and their brains processed for the immunohistochemistry tyrosine hydroxylase (TH), a marker for dopamine cells, as well as glial fibrillary acidic protein (GFAP) and OX-42, markers for astrocyte and microglia, respectively. Stereological tools were employed in the quantifications. The volumes of the regions of the striatal TH immunoreactive disappearance, as well as the astroglial and microglial activation were several folds increased compared to control saline-injected rats. The optical disector detected decreases in the estimated total number of dopamine cells in the entire ipsilateral pars compacta of the substantia nigra (SNc) and ventral tegmental area (VTA) as well as in the estimated total number of varicosity profiles in the entire ipsilateral neostriatum. The stereological tool rotator showed no changes either in the mean or in the histogram distribution of the cytoplasmic volume of the nigral and VTA dopamine cells of 6-OHDA lesioned rats. Increases in the estimated total number of GFAP positive astrocytes were found in the entire neostriatum bilaterally as well as in the ipsilateral entire SNc and VTA of 6-OHDA lesioned rats. The estimated total number of OX-42 immunoreactive microglial profiles was elevated only in the ipsilateral entire neostriatum of the lesioned rats. The rotator detected cytoplasmic hypertrophy in the astrocytes, and also a shift to the right of the gaussian curves of the normal distribution of the logarithmic plotted values of the astroglial cell body volumes,of the neostriatum bilaterally as well as in the ipsilateral SNc and VTA of the striatal 6-OHDA injected rats. Cytoplasmic hypertrophy of microglia, and also a shift to the right of the gaussian curves of the values of microglia cell body volumes were seen only in the ipsilateral neostriatum; however, the point intercepts revealed an increased amount of microglial processes in the ipsilateral SNc and VTA of the lesioned rats. Specific stereological methods can be applied on detection of regionally different forms of cellular astroglial and microglial reaction after a partial lesion of dopamine pathway.