Michel Soares Mesquita

Centrally injected tityustoxin produces the systemic manifestations observed in severe scorpion poisoning.

In this work we submitted adult male Wistar rats to intracerebroventricular (icv) and iv microinjections of the fraction tityostoxin (TsTX) from the Tityus serrulatus scorpion venom, to address whether the CNS could account for the systemic alterations previously reported: cardiac arrhythmias, lung edema, and seizures. Animals were injected icv, total volume of 1.0 microl, with either sterile saline (n = 4) or differing doses of TsTX (1.74, n = 5; 0.174, n = 4; 0.087, n = 5; and 0.058 microg, n = 4). The peripheral effect of the highest dose of TsTX used (1.74 microg) was tested through iv injections in the femoral vein (n = 4). All animals were recorded by a Video EEG/ECG system for a maximum period of 90 mins or until death. After recording, the lungs were harvested and weighed to evaluate edema (lung/body wt x100). Our results show that icv injections of TsTX, but not iv injections, were able to provoke heart arrhythmias, lung edema, and seizures. Furthermore, the toxin was capable of producing epileptiform discharges in all animals injected with 1.74 microg of the toxin. In conclusion, the action of TsTX in the CNS may solely account for the peripheral alterations observed in severe cases of Tityus serrulatus scorpion poisoning.

Distinct temporal patterns of electrical stimulation influence neural recruitment during PTZ infusion: An fMRI study

Our working hypothesis is that constant inter-pulse interval (IPI) electrical stimulation (ES) would resonate with endogenous epileptogenic reverberating circuits, favoring seizure, while random inter-interval ES protocol would promote desynchronization of such neural networks, interfering with the abnormal recruitment of neural structures. Male Wistar rats were stereotaxically implanted with a monopolar ES carbon-fiber electrode (minimizing fMRI artifact) in the amygdala. A 7T fMRI scanner was used to evaluate brain activity during ES, fixed four pulses per second ratio, using either a periodic IPI (ES-P) or random IPI (non-periodic ES-NP) stimulation paradigm. Appropriate imaging protocols were used to compare baseline BOLD (blood oxygen level dependent) MRI with scans during ES. A second series of experiments, both without stimuli and under the same ES paradigms, were evaluated during continuous infusion of pentylenetetrazole (PTZ, 4 mg/ml/min) through an i.v. catheter. Our results show that temporal lobe activation during ES-P or ES-NP did not present any statistical differences during ES. However, during PTZ infusion, PTZ-P facilitated recruitment of the temporal lobe ipsilateral to ES while PTZ-NP showed significantly less activation ipsilateral to ES and, in turn, less inter-hemispheric differences. Altogether, our results support the hypothesis of reverberating circuits being synchronized by ES-P and desynchronized by ES-NP. Time-coded low frequency stimulation may be an interesting alternative treatment for patients with refractory epilepsy.

Electroencephalographic evidence of brainstem recruitment during scorpion envenomation.

Scorpion envenomation is a public health problem in Brazil, with most severe cases occuring in children under the age of 5 years (0.6% lethality). In fact, the toxic fractions of the Tityus serrulatus scorpion venom (TSSV) have greater permeability across the BBB of weanling rats when compared to adults. Although EEG alterations have been reported in up to 75% of pediatric severe cases, the role of the CNS in envenomation morbidity is still in debate. Our working hypothesis is that the neural substrates that play a major role in morbidity generate activity undetectable from EEG scalp leads. Twenty one-day-old rats (n=18) were injected s.c. with the deadliest toxic fraction of the TSSV, tityustoxin (TsTX; 2xDL50=6 mg/kg). EEG leads were stereotaxicaly implanted in the nucleus of the solitary tract (NTS) and left parietal cortex. EEG and ECG were continuously monitored by a video EEG system until death or for a maximum period of 240 min. An experimental group pre-treated with carbamazepine (CBZ) was added in order to better access the cause-effect relationship between neural discharges and the systemic ECG alterations. High amplitude discharges in the NTS, which correlated to cardiac alterations, were recorded soon after administration of TsTX. Abnormal electrographic activity spread throughout the cortex only later in the recording. As expected, the CBZ treatment increased the latency for the first epileptiform discharge, decreased EEG/ECG alterations and increased the general survival time. In summary: peripheral scorpion toxin inoculation recruits brainstem involved in cardiovascular control and initial electrographic activity was undetectable from the cortical electrode.

Brainstem Structures Are Primarily Affected in an Experimental Model of Severe Scorpion Envenomation

Severe scorpion envenoming (SSE) is more frequent in children and is characterized by systemic dysfunctions with a mortality rate of up to 9%. Recent evidence shows that the central nervous system (CNS) plays a key role in triggering the cascade of symptoms present in SSE. The age-dependent role of the CNS in SSE lethality may be summarized in 3 hypotheses: (1) the shown increased blood brain barrier permeability of infants to the toxins would especially and primarily compromise neurovegetative control areas, (2) the neurons within these areas have high affinity to the toxins, and (3) the neurovascular interaction is such that SSE metabolically compromises proper function of toxin-targeted areas. A pharmacological magnetic resonance imaging paradigm was used to evaluate localized hemodynamic changes in relative cerebral blood volume (rCBV) for 30 min after the injection of TsTX, the most lethal toxin from the venom of the Tityus serrulatus scorpion. The brainstem showed significant rCBV reduction 1 min after TsTX administration, whereas rostral brain areas had delayed increase in rCBV (confirmed by laser Doppler measurements of cortical cerebral blood flow). Moreover, metabolic activity by 14C-2-deoxyglucose autoradiography showed the highest relative increase at the brainstem. To test whether TsTX has high affinity to brainstem neurons, the lateral ventricle was injected with Alexa Fluor 568 TsTX. Although some neurons showed intense fluorescence, the labeling pattern suggests that specific neurons were targeted. Altogether, these results suggest that brainstem areas involved in neurovegetative control are most likely within the primary structures triggering the cascade of symptoms present in SSE.

Characterization of gray matter atrophy following 6-hydroxydopamine lesion of the nigrostriatal system

BACKGROUND The unilaterally-lesioned 6-hydroxydopamine (6-OHDA) rat is one of the most commonly used experimental models of Parkinsons disease (PD). Here we investigated whether magnetic resonance imaging (MRI) that is widely used in human PD research, has the potential to non-invasively detect macroscopic structural brain changes in the 6-OHDA rat in ways translatable to humans. METHODS We measured the gray matter (GM) composition in the unilateral 6-OHDA rat in comparison to sham animals using whole-brain voxel-based morphometry (VBM) - an unbiased MR image analysis technique. The number of nigral dopamine (DA) neurons and the density of their cortical projections were examined post-mortem using immunohistochemistry. RESULTS VBM revealed widespread bilateral changes in gray matter volume (GMV) on a topographic scale in the brains of 6-OHDA rats, compared to sham-operated rats. The greatest changes were in the lesioned hemisphere, which displayed reductions of GMV in motor, cingulate and somatosensory cortex. Histopathological results revealed dopaminergic cell loss in the substantia nigra (SN) and a denervation in the striatum, as well as in the frontal, somatosensory and cingulate cortices. CONCLUSION Unilateral nigrostriatal 6-OHDA lesioning leads to widespread GMV changes, which extend beyond the nigrostriatal system and resemble advanced Parkinsonism. This study highlights the potential of structural MRI, and VBM in particular, for the system-level phenotyping of rodent models of Parkinsonism and provides a methodological framework for future studies in novel rodent models as they become available to the research community.

Alterations in brain microstructure in rats that develop abnormal aggression following peripubertal stress

Exposure to early adversity is implicated in the development of aggressive behaviour later in life in some but not all individuals. The reasons for the variability in response to such experiences are not clear but may relate to pre‐existing individual differences that influence their downstream effects. Applying structural magnetic resonance imaging (MRI) to a rat model of abnormal aggression induced by peripubertal stress, we examined whether individual differences in the development of an aggressive phenotype following stress exposure were underpinned by variation in the structure of aggression‐associated, corticolimbic brain regions. We also assessed whether responsiveness of the hypothalamic–pituitary–adrenal axis to stress was associated with neurobehavioural outcome following adversity. A subset of the rats exposed to peripubertal stress developed an aggressive phenotype, while the remaining rats were affected in other behavioural domains, such as increased anxiety‐like behaviours and reduced sociability. Peripubertal stress led to changes in tissue microstructure within prefrontal cortex, amygdala and hippocampal formation only in those individuals displaying an aggressive phenotype. Attenuated glucocorticoid response to stress during juvenility predicted the subsequent development of an aggressive phenotype in peripubertal stress‐exposed rats. Our study establishes a link between peripubertal stress exposure in rats and structural deviations in brain regions linked to abnormal aggression and points towards low glucocorticoid responsiveness to stress as a potential underlying mechanism. We additionally highlight the importance of considering individual differences in behavioural response to stress when determining neurobiological correlates.