Hypoactivity and neurochemical alterations in the basal ganglia of female Sprague-Dawley rats after repeated exposure to atrazine.

The herbicide atrazine (ATR) has been one of the most widely used herbicides worldwide. However, due to its indiscriminate use, it has been considered an environmental contaminant. Several studies have classified ATR as an endocrine disruptor, and it has been found to have neurotoxic effects on behavior, along with alterations in the dopaminergic, GABAergic, and glutamatergic systems in the basal ganglia of male rodents. These findings suggest that these neurotransmitter systems are targets of this herbicide. However, there are no studies evaluating the neurotoxicity of ATR in female rodents. Our study aimed to assess the effects of repeated IP injections of 100 mg ATR/kg or a vehicle every other day for 2 weeks (six injections) on the locomotor activity, content of monoamines, GABA, glutamate, and glutamine in the striatum, nucleus accumbens, ventral midbrain, and prefrontal cortex, and tyrosine hydroxylase (TH) protein levels in striatum and nucleus accumbens of female rats. Repeated 100 mg ATR/kg injections immediately decreased all the locomotor activity parameters evaluated, and such hypoactivity persisted for at least 48 h after the last ATR administration. The ATR administration increased dopamine and DOPAC content in the nucleus accumbens and the dopamine and DOPAC and serotonin and 5-HIAA content in the ventral midbrain. In contrast, the TH protein levels in the striatum and nucleus accumbens were similar between groups. Meanwhile, GABA, glutamine, and glutamate levels remained unaltered in all brain regions evaluated. The observed behavioral alterations could be associated with the monoamine changes presented by the rats. These data reveal that the nucleus accumbens and ventral midbrain are susceptible to repeated ATR exposure in female rats.

Early and transitory hypoactivity and olfactory alterations after chronic atrazine exposure in female Sprague-Dawley rats.

Several studies have shown that chronic exposure to the herbicide atrazine (ATR) causes alterations in locomotor activity and markers of the dopaminergic systems of male rats. However, few studies have evaluated the sex-dependent effects of atrazine exposure. The aim of the present study was to evaluate whether chronic ATR exposure causes alterations in behavioral performance and dopaminergic systems of female rats. At weaning, two groups of rats were exposed to 1 or 10 mg ATR/kg body weight daily thorough the food, while the control group received food without ATR for 14 months. Spontaneous locomotor activity was evaluated monthly for 12 months, while anxiety, egocentric and spatial memory, motor coordination, and olfactory function tasks were evaluated between 13 and 14 months of ATR exposure. Tyrosine hydroxylase (TH) and monoamine content in brain tissue were assessed at the end of ATR treatment. Female rats treated with 1 or 10 mg ATR showed vertical hypoactivity compared to the control group only in the first month of ATR exposure. Impairments in olfactory functions were found due to ATR exposure. Nevertheless, no alterations in anxiety, spatial and egocentric memory, or motor coordination tasks were observed, while the levels of TH and dopamine and its metabolites in brain tissue were similar among groups. These results suggest that female rats could present greater sensitivity to the neurotoxic effects of ATR on spontaneous locomotor activity in the early stages of development. However, they are unaffected by chronic ATR exposure later in life compared to male rats. More studies are necessary to unravel the sex-related differences observed after chronic ATR exposure.

Contribution and functional connectivity between cerebrum and cerebellum on sub-lexical and lexical-semantic processing of verbs.

Language comprehension involves both sub-lexical (e.g., phonological) and lexical-semantic processing. We conducted a task using functional magnetic resonance imaging (fMRI) to compare the processing of verbs in these two domains. Additionally, we examined the representation of concrete-motor and abstract-non-motor concepts by including two semantic categories of verbs: motor and mental. The findings indicate that sub-lexical processing during the reading of pseudo-verbs primarily involves the left dorsal stream of the perisylvian network, while lexical-semantic representation during the reading of verbs predominantly engages the ventral stream. According to the embodied or grounded cognition approach, modality-specific mechanisms (such as sensory-motor systems) and the well-established multimodal left perisylvian network contribute to the semantic representation of both concrete and abstract verbs. Our study identified the visual system as a preferential modality-specific system for abstract-mental verbs, which exhibited functional connectivity with the right crus I/lobule VI of the cerebellum. Taken together, these results confirm the dissociation between sub-lexical and lexical-semantic processing and provide neurobiological evidence of functional coupling between specific visual modality regions and the right cerebellum, forming a network that supports the semantic representation of abstract concepts. Further, the results shed light on the underlying mechanisms of semantic processing and contribute to our understanding of how the brain processes abstract concepts.

A novel task to evaluate irony comprehension and its essential elements in Spanish speakers.

An ironic statement transmits the opposite meaning to its literal counterpart and is one of the most complex communicative acts. Thus, it has been proposed to be a good indicator of social communication ability. Prosody and facial expression are two crucial paralinguistic cues that can facilitate the understanding of ironic statements. The primary aim of this study was to create and evaluate a task of irony identification that could be used in neuroimaging studies. We independently evaluated three cues, contextual discrepancy, prosody and facial expression, and selected the best cue that would lead participants in fMRI studies to identify a stimulus as ironic in a reliable way. This process included the design, selection, and comparison of the three cues, all of which have been previously associated with irony detection. The secondary aim was to correlate irony comprehension with specific cognitive functions. Results showed that psycholinguistic properties could differentiate irony from other communicative acts. The contextual discrepancy, prosody, and facial expression were relevant cues that helped detect ironic statements; with contextual discrepancy being the cue that produced the highest classification accuracy and classification time. This task can be used successfully to test irony comprehension in Spanish speakers using the cue of interest. The correlation of irony comprehension with cognitive functions did not yield consistent results. A more heterogeneous sample of participants and a broader battery of tests may be needed to find reliable cognitive correlates of irony comprehension.

Theory of Mind and Its Elusive Structural Substrate.

Is brain structure related to function? Can one predict the other? These are questions that are still waiting to be answered definitively. In this paper we seek to investigate these questions, in particular, we are interested in the relation between brain structure and theory of mind (ToM). ToM is defined as the ability to attribute mental states to others. Previous studies have observed correlations between performance on ToM tasks, and gray-matter size/volume in dorsomedial prefrontal cortex (dmPFC), temporoparietal junction (TPJ) and precuneus (PCu). Despite these findings, there are concerns about false positive results and replicability issues. In this study we used two different tasks to evaluate ToM, Reading the Mind in the Eyes Test (RMET), and the Short Story Task (SST). Performance in these tasks was correlated to brain anatomy measures including voxel-based morphometry (VBM) and cortical thickness (CT) analysis, from ninety-one neurotypical participants. High-resolution structural brain images were acquired, and whole-brain and region of interest (ROI) analyses were implemented. The analyses did not show statistically significant associations between ToM performance and brain structural measures after correction. Significant associations between performance on ToM tests and a widespread array of regions loosely associated with ToM were observed only for whole brain uncorrected analysis (p < 0.001). These results do not replicate a previous study with neurotypical participants. We tested two different ToM tests, two different softwares for VBM and CT, and we used two samples, one with 91 and a sub-sample with 69 participants. Neither of these conditions made a difference in the results obtained. Consequently, these results suggest that if the population is neurotypical and homogenous, it is unlikely that a reliable association between brain anatomy measures and ToM performance, as measured with these tasks, may be found.

Chronic Exposure to Arsenic in Drinking Water Causes Alterations in Locomotor Activity and Decreases Striatal mRNA for the D2 Dopamine Receptor in CD1 Male Mice.

Arsenic exposure has been associated with sensory, motor, memory, and learning alterations in humans and alterations in locomotor activity, behavioral tasks, and neurotransmitters systems in rodents. In this study, CD1 mice were exposed to 0.5 or 5.0 mg As/L of drinking water for 6 months. Locomotor activity, aggression, interspecific behavior and physical appearance, monoamines levels, and expression of the messenger for dopamine receptors D1 and D2 were assessed. Arsenic exposure produced hypoactivity at six months and other behaviors such as rearing and on-wall rearing and barbering showed both increases and decreases. No alterations on aggressive behavior or monoamines levels in striatum or frontal cortex were observed. A significant decrease in the expression of mRNA for D2 receptors was found in striatum of mice exposed to 5.0 mg As/L. This study provides evidence for the use of dopamine receptor D2 as potential target of arsenic toxicity in the dopaminergic system.

The herbicide glyphosate causes behavioral changes and alterations in dopaminergic markers in male Sprague-Dawley rat.

Glyphosate (Glyph) is the active ingredient of several herbicide formulations. Reports of Glyph exposure in humans and animal models suggest that it may be neurotoxic. To evaluate the effects of Glyph on the nervous system, male Sprague-Dawley rats were given six intraperitoneal injections of 50, 100, or 150 mg Glyph/kg BW over 2 weeks (three injections/week). We assessed dopaminergic markers and their association with locomotor activity. Repeated exposure to Glyph caused hypoactivity immediately after each injection, and it was also apparent 2 days after the last injection in rats exposed to the highest dose. Glyph did not decrease monoamines, tyrosine hydroxylase (TH), or mesencephalic TH+ cells when measured 2 or 16 days after the last Glyph injection. In contrast, Glyph decreased specific binding to D1 dopamine (DA) receptors in the nucleus accumbens (NAcc) when measured 2 days after the last Glyph injection. Microdialysis experiments showed that a systemic injection of 150 mg Glyph/kg BW decreased basal extracellular DA levels and high-potassium-induced DA release in striatum. Glyph did not affect the extracellular concentrations of 3,4-dihydroxyphenylacetic acid or homovanillic acid. These results indicate that repeated Glyph exposure results in hypoactivity accompanied by decreases in specific binding to D1-DA receptors in the NAcc, and that acute exposure to Glyph has evident effects on striatal DA levels. Additional experiments are necessary in order to unveil the specific targets of Glyph on dopaminergic system, and whether Glyph could be affecting other neurotransmitter systems involved in motor control.

Repeated exposure to the herbicide atrazine alters locomotor activity and the nigrostriatal dopaminergic system of the albino rat.

Atrazine (ATR) is used as a pre- and post-emergent herbicide; although banned in several countries of the European Community, it is still used extensively around the world. A recent study in rats has shown that chronic, daily exposure to 10 mg ATR/kg BW causes hyperactivity, disrupts motor coordination and learning of behavioral tasks, and decreases dopamine levels in the brain. In order to evaluate the short-term effect of ATR exposure on locomotor activity, monoamine markers, and antioxidants, adult male Sprague-Dawley rats received six IP injections of 100 mg ATR/kg BW or vehicle over two weeks. After every ATR injection we found hypoactivity that lasted up to five days, and it was accompanied by reductions in levels of striatal DA, DOPAC, and HVA without any alteration in the striatal expression of the mRNAs for Mn-SOD, Trx-1, DAR-D(1), or DAR-D(2). In contrast, in the nucleus accumbens no changes in monoamine markers were observed, and a down-regulation of Trx-1 expression was detected shortly after the ATR treatment. Moreover, in the ventral midbrain, we found that ATR induced a down-regulation of mRNA for Th and DAT, but it increased VMAT2 mRNA expression. Decreases of monoamine levels and of locomotor activity disappeared three months after ATR treatment; however, an amphetamine challenge (1 mg/kg) given two months after the ATR treatment resulted in a significant stimulation in the exposed group, revealing hidden effects of ATR on dopaminergic systems. These results indicate that ATR exposure differentially modifies the dopaminergic systems, and these modifications may underlie the behavioral changes observed.

Evaluation of plasmid permanence in transfected cells after transplantation into the rat brain.

Plasmid retention after long-term transplantation has been one of the major technical limitations for transplantation studies. This study describes the use of a modified protocol of Hirt and a SYBR Green-based quantitative real-time PCR (qPCR) to recover and quantify a vector containing a specific transgene in transfected cells after brain transplantation. We compared various methods for sample processing and recovery of extrachromosomal DNA suitable for qPCR. The modified protocol of Hirt was the most reliable for optimal plasmid recovery from transplanted tissue with minimal loss of plasmid DNA compared to a commercial kit or TRIzol(®) protocols. The PCR protocol for plasmid and transgene detection included the design of two highly specific primer sets to detect the sequence for the human glutamate decarboxylase 1 (hGAD(67)) transgene by SYBR Green-based qPCR, and to confirm the presence of vector pREP10 hGAD(67) by end-point PCR. We used a standard curve constructed from serial dilutions of pure plasmid pREP10 hGAD(67) as reference in qPCR experiments to determine the number of plasmid copies recovered from cultured cells and tissue samples after Hirt extraction. Then, plasmid permanence was evaluated in transplanted tissues after different time intervals, and plasmid loss in the tissue of interest was found to be time dependent. In this study we describe an easy, highly specific, low-cost, and reliable method for plasmid recovery and quantification of a transgene of interest in long-term brain transplantation studies; use of this method may be extended to other transplantation models.

Intranigral transplants of a GABAergic cell line produce long-term alleviation of established motor seizures.

We have previously shown that intranigral transplants of immortalized GABAergic cells decrease the number of kainic acid-induced seizures [Castillo CG, Mendoza S, Freed WJ, Giordano M. Intranigral transplants of immortalized GABAergic cells decrease the expression of kainic acid-induced seizures in the rat. Behav Brain Res 2006;171:109-15] in an animal model. In the present study, recurrent spontaneous behavioral seizures were established by repeated systemic injections of this excitotoxin into male Sprague-Dawley rats. After the seizures had been established, cells were transplanted into the substantia nigra. Animals with transplants of control cells (without hGAD67 expression) or with sham transplants showed a death rate of more than 40% over the 12 weeks of observation, whereas in animals with M213-2O CL-4 transplants, the death rate was reduced to less than 20%. The M213-2O CL-4 transplants significantly reduced the percentage of animals showing behavioral seizures; animals with these transplants also showed a lower occurrence of stage V seizures than animals in the other groups. In vivo and in vitro analyses provided evidence that the GABAergic cells show sustained expression of both GAD67 and hGAD67 cDNA, as well as increased gamma-aminobutyric acid (GABA) levels in the ventral mesencephalon of transplanted animals. Therefore, transplantation of GABA-producing cells can produce long-term alleviation of behavioral seizures in an animal model.

Long-term effects of striatal lesions on c-Fos immunoreactivity in the pedunculopontine nucleus.

The basal ganglia are a group of subcortical nuclei classically thought to be involved in the control of movement, and they have reciprocal connections with the cortex, thalamus and structures in the brainstem. Recent findings suggest that the basal ganglia interact with structures involved in the control of the sleep-waking cycle. The pedunculopontine tegmental nucleus (PPN) maintains a close relationship with the basal ganglia and is intimately involved in the regulation of wakefulness and REM sleep. This study evaluated changes in activity of PPN neurons following striatal kainic acid-induced lesions. Rats were injected in the anterodorsal striatum with either kainic acid or vehicle and allowed to recover for 7 or 30 days. The results showed an increase in the number of c-Fos+ cells in the PPN 30 days but not 7 days after the striatal lesion, when motor hyperactivity was no longer detected. In addition, we found a significant correlation between the ventricular brain ratio, as an indicator of lesion size, and the number of c-Fos+ cells in the PPN. Furthermore, the spatial distribution of cell types suggested that most c-Fos+ cells in the PPN were not cholinergic. These results provide new insights into the functional relationship between the basal ganglia and the PPN and suggest that the striatum, through its indirect influence on the PPN, may contribute to the regulation of wakefulness and cortical activation.

Induction of c-fos in nucleus accumbens in naive male Balb/c mice after wheel running.

Immunocytochemical labeling of the transcription factor c-Fos was used to determine whether wheel running (WR) activates nucleus accumbens (NAcc) neurons in naive male Balb/c mice. The results indicate compared to novel environment exploration and forced locomotor activity, WR increases c-Fos labeling in the middle level of the core but not the shell of the NAcc. These data show that WR activates middle-core NAcc neurons more effectively than other locomotor activities. The middle core of the NAcc may be a critical region for the self-reinforcing properties of WR.

Striatal dopaminergic stimulation produces c-Fos expression in the PPT and an increase in wakefulness.

Striatal activation can modify activity in cortical areas related to specific striatal functions possibly through a process of disinhibition within the basal ganglia. Anatomical studies have shown substantial GABAergic innervation from these nuclei to the pedunculopontine tegmental nucleus (PPT). Thus, dopaminergic stimulation of the striatum could produce PPT disinhibition and result in non-specific cortical activation. To test this hypothesis, d-amphetamine was infused both into the striatum of freely moving rats for motor and electrocorticographic recordings, and into the striatum of animals under deep anesthesia for c-Fos immunohistochemistry. The results show that intrastriatal amphetamine increases wakefulness independent of motor activity, and it increases c-Fos expression in the PPT and adjacent areas. They also suggest that the striatum participates in non-specific cortical activation probably as a result of its relationship with the PPT.

Changes in sleep-waking cycle after striatal excitotoxic lesions.

Huntington's disease (HD) patients show severe diurnal choreic movements, while during slow-wave sleep (SWS) abnormal movements subside. Sleep disturbances in HD, including irregular delta activity and decreases in SWS, have also been reported. Striatal excitotoxic lesions have been shown to induce increased nocturnal spontaneous locomotor activity in rodents. In order to characterize the changes in circadian activity and sleep patterns and their correlation with motor activity after striatal excitotoxic lesions, Sprague-Dawley rats were implanted and lesioned; their locomotor and EEG activities were recorded for either 4 or 24 h during baseline or 7 and 30 days post-lesion. Locomotor activity increased significantly at 7 days post-lesion during the dark phase of the light-dark cycle. In contrast, total time spent in wakefulness (W) increased at 30 days post-lesion during the light phase of the cycle. This increase was at the expense of SWS duration. No disruption of the circadian curves was observed. Increases in the number of W-bouts and decreases in the duration of SWS-bouts were also observed. These results suggest the possible participation of the striatum in the regulation of the sleep-waking cycle, independent of locomotor activity. The increase in W could be due to loss of inhibition of target structures involved in regulation of the sleep-waking cycle.

Insulin-like growth factor I partly prevents axon elimination in the neonate rat optic nerve.

Developmental neuronal death ensues after access of innervating neurons to target-derived neurotrophic factors is restricted. Recent evidence suggests, however, that growth factors such as those of the insulin family modulate neuronal death through autocrine/paracrine mechanisms. In rats, retinal ganglion neurons (RGNs) undergo massive death during early postnatal life. During this same period, the expression of various members of the insulin-like growth factor I (IGF-I) protein family is down regulated. To evaluate whether ocular IGF-I might modulate RGN death, we administered IGF-I in the posterior chamber of the eye of newborn rats. Optic nerve fiber number was estimated in control and IGF-I treated animals at postnatal day 5 when RGN death peaks. Intraocular IGF-I treatment at birth partly prevented optic nerve fiber elimination. Because the axon number in the optic nerve correlates to some extent with the RGN number, these results suggest that IGF-I may modulate RGN death in vivo through local interactions.