Modulating Expression of Endogenous Interleukin 1 Beta in the Acute Phase of the Pilocarpine Model of Epilepsy May Change Animal Survival.

The pilocarpine-induced (PILO) model has helped elucidate the electrophysiological and molecular aspects related to mesial temporal lobe epilepsy. It has been suggested that the extensive cell death and edema observed in the brains of these animals could be induced by increased inflammatory responses, such as the rapid release of the inflammatory cytokine interleukin 1 beta (Il1b). In this study, we investigate the role of endogenous Il1b in the acute phase of the PILO model. Our aim is twofold. First, we want to determine whether it is feasible to silence Il1b in the central nervous system using a non-invasive procedure. Second, we aim to investigate the effect of silencing endogenous Il1b and its antagonist, Il1rn.We used RNA interference applied non-invasively to knockdown Il1b and its endogenous antagonist Il1rn. We found that knocking down Il1b prior to pilocarpine injection increased the mortality rate of treated animals. Furthermore, we observed that, when exposing the animals to more Il1b by silencing its endogenous antagonist Il1rn, there was a better response to status epilepticus with decreased animal mortality in the acute phase of the PILO model. Thus, we show the feasibility of using a novel, less invasive approach to study genes involved in the inflammatory response in the central nervous system. Furthermore, our results provide suggestive evidence that modulating endogenous Il1b improves animal survival in the acute phase of the PILO model and may have effects that extend into the chronic phase.

Asymptomatic leptospiral infection is associated with canine chronic kidney disease.

Canine leptospirosis is characterized by an acute or chronic disease. Some dogs may act as asymptomatic carriers, keeping the agent in the renal tubules and eliminating it in the urine for an extended period. Chronic kidney disease (CKD) is multifactorial and pathophysiology has been widely discussed. The aim of the study was to investigate whether the occurrence of CKD may possibly be associated with asymptomatic leptospiral infection in dogs in endemic regions. Serology and urine PCR were performed in 16 dogs with CKD and 48 healthy dogs from an endemic area. Dogs with CKD were more frequently shedders (75%) than non-CKD animals (20.8%). Therefore, our results demonstrate that asymptomatic leptospiral infection is associated with canine chronic kidney disease and that differential diagnosis is important for dogs from endemic areas presenting CKD. The early detection of shedders, besides the obvious impact on Public health may also help to improve the animal health and avoid the development of CKD.

Role of non-coding RNAs in non-aging-related neurological disorders.

Protein coding sequences represent only 2% of the human genome. Recent advances have demonstrated that a significant portion of the genome is actively transcribed as non-coding RNA molecules. These non-coding RNAs are emerging as key players in the regulation of biological processes, and act as "fine-tuners" of gene expression. Neurological disorders are caused by a wide range of genetic mutations, epigenetic and environmental factors, and the exact pathophysiology of many of these conditions is still unknown. It is currently recognized that dysregulations in the expression of non-coding RNAs are present in many neurological disorders and may be relevant in the mechanisms leading to disease. In addition, circulating non-coding RNAs are emerging as potential biomarkers with great potential impact in clinical practice. In this review, we discuss mainly the role of microRNAs and long non-coding RNAs in several neurological disorders, such as epilepsy, Huntington disease, fragile X-associated ataxia, spinocerebellar ataxias, amyotrophic lateral sclerosis (ALS), and pain. In addition, we give information about the conditions where microRNAs have demonstrated to be potential biomarkers such as in epilepsy, pain, and ALS.

Role of non-coding RNAs in non-aging-related neurological disorders

Protein coding sequences represent only 2% of the human genome. Recent advances have demonstrated that a significant portion of the genome is actively transcribed as non-coding RNA molecules. These non-coding RNAs are emerging as key players in the regulation of biological processes, and act as "fine-tuners" of gene expression. Neurological disorders are caused by a wide range of genetic mutations, epigenetic and environmental factors, and the exact pathophysiology of many of these conditions is still unknown. It is currently recognized that dysregulations in the expression of non-coding RNAs are present in many neurological disorders and may be relevant in the mechanisms leading to disease. In addition, circulating non-coding RNAs are emerging as potential biomarkers with great potential impact in clinical practice. In this review, we discuss mainly the role of microRNAs and long non-coding RNAs in several neurological disorders, such as epilepsy, Huntington disease, fragile X-associated ataxia, spinocerebellar ataxias, amyotrophic lateral sclerosis (ALS), and pain. In addition, we give information about the conditions where microRNAs have demonstrated to be potential biomarkers such as in epilepsy, pain, and ALS.

High number of asymptomatic dogs as leptospiral carriers in an endemic area indicates a serious public health concern.

Asymptomatic dogs can be potential hosts of leptospirosis. However, the extension of this phenomenon in endemic areas has not yet been clearly defined. This study is aimed at evaluating the role of asymptomatic dogs as carriers of Leptospira in an endemic area of Brazil. A total of 131 male dogs without apparent leptospirosis symptoms were included in the study based on clinical and hematologic exams. Serum and urine samples were collected for microscopic agglutination tests (MAT) and polymerase chain reactions (PCR) targeted the LipL32 gene, respectively. Forty-two dogs (32·1%) presented seroreactivity (titres ⩾100). The serogroup Icterohaemorrhagiae was predominant, representing 92·7% of the seropositive samples. Overall, leptospiral DNA was detected on 26 urine samples (19·8%). PCR positivity was more common (28·6%) on seropositive dogs than on seronegative (15·7%) ones. Nevertheless, MAT was not correlated to PCR (P > 0·05). Age was not associated with seroreactivity, but dogs older than 5 years of age had 4·07 more chances (odds ratio) of being carriers (PCR positive) than younger ones. Although the fact of knowing that asymptomatic dogs can act as leptospiral carriers is not new, the extension of this fact is impressive in an endemic region, and its role and impact on public health cannot be neglected.

RNA sequencing reveals region-specific molecular mechanisms associated with epileptogenesis in a model of classical hippocampal sclerosis.

We report here the first complete transcriptome analysis of the dorsal (dDG) and ventral dentate gyrus (vDG) of a rat epilepsy model presenting a hippocampal lesion with a strict resemblance to classical hippocampal sclerosis (HS). We collected the dDG and vDG by laser microdissection 15 days after electrical stimulation and performed high-throughput RNA-sequencing. There were many differentially regulated genes, some of which were specific to either of the two sub-regions in stimulated animals. Gene ontology analysis indicated an enrichment of inflammation-related processes in both sub-regions and of axonal guidance and calcium signaling processes exclusively in the vDG. There was also a differential regulation of genes encoding molecules involved in synaptic function, neural electrical activity and neuropeptides in stimulated rats. The data presented here suggests, in the time point analyzed, a remarkable interaction among several molecular components which takes place in the damaged hippocampi. Furthermore, even though similar mechanisms may function in different regions of the DG, the molecular components involved seem to be region specific.

Physical exercise prevents motor disorders and striatal oxidative imbalance after cerebral ischemia-reperfusion.

Stroke is the third most common cause of death worldwide, and most stroke survivors present some functional impairment. We assessed the striatal oxidative balance and motor alterations resulting from stroke in a rat model to investigate the neuroprotective role of physical exercise. Forty male Wistar rats were assigned to 4 groups: a) control, b) ischemia, c) physical exercise, and d) physical exercise and ischemia. Physical exercise was conducted using a treadmill for 8 weeks. Ischemia-reperfusion surgery involved transient bilateral occlusion of the common carotid arteries for 30 min. Neuromotor performance (open-field and rotarod performance tests) and pain sensitivity were evaluated beginning at 24 h after the surgery. Rats were euthanized and the corpora striata was removed for assay of reactive oxygen species, lipoperoxidation activity, and antioxidant markers. Ischemia-reperfusion caused changes in motor activity. The ischemia-induced alterations observed in the open-field test were fully reversed, and those observed in the rotarod test were partially reversed, by physical exercise. Pain sensitivity was similar among all groups. Levels of reactive oxygen species and lipoperoxidation increased after ischemia; physical exercise decreased reactive oxygen species levels. None of the treatments altered the levels of antioxidant markers. In summary, ischemia-reperfusion resulted in motor impairment and altered striatal oxidative balance in this animal model, but those changes were moderated by physical exercise.

Neuronal P2X3 receptor activation is essential to the hyperalgesia induced by prostaglandins and sympathomimetic amines released during inflammation.

We have demonstrated that the activation of P2X3 receptor on peripheral afferent neurons is critical to development of inflammatory hyperalgesia in peripheral tissue, although pharmacological administration of prostaglandin E(2) or sympathomimetic amines is enough to sensitize primary afferent neurons by acting directly in neuronal receptors. Therefore, to clarify this ambiguity this study verifies whether P2X3 receptor activation on primary afferent neurons enables the sensitization induced by prostaglandin E(2) or sympathomimetic amine. Initially, this study confirmed that co-administration of A317491 (60 µg/paw), a selective P2X3 receptor antagonist, or pre-treatment with dexamethasone (1 mg/mL/kg) prevents the mechanical hyperalgesia induced by carrageenan (300 µg/paw) in the rat's hind paw. Sub-threshold doses of PGE(2) (4 ng/paw) or dopamine (0.4 µg/paw), that do not induce hyperalgesia by themselves, when injected just following αßmeATP or carrageenan in rats treated with dexamethasone induced hyperalgesia, which is prevented by A317491 or treatment with periganglionar (DRG-L5) injections of ODN-antisense, against P2X3 receptor. Furthermore, because PKCɛ translocation induces an increase of neuronal susceptibility to inflammatory mediators, this study demonstrates that αßmeATP in peripheral tissue increases the expression of PKCɛ in cell membranes of DRG-L5, and in contrast, the administration of PKCɛ translocation inhibitor (1 µg/paw) in peripheral tissue 45 min before αßmeATP, prevented the hyperalgesia induced by sub-threshold dose of PGE(2) (4 ng/paw). In conclusion, this study suggests that neuronal P2X3 receptor activation and the consequent PKCɛ translocation increase the susceptibility of nociceptor to inflammatory mediators allowing the development of inflammatory hyperalgesia.

The antidepressive effect of the physical exercise correlates with increased levels of mature BDNF, and proBDNF proteolytic cleavage-related genes, p11 and tPA.

Clinical studies show an evident antidepressive effect of physical exercise and animal research corroborate such evidence. However, the neurobiological mechanisms underlying the antidepressive effect of exercise are not completely understood. Notwithstanding, it is known that exercise increases brain-derived neurotrophic factor (BDNF) expression in the hippocampus similarly to antidepressant drugs. BDNF is synthesized as a precursor molecule that undergoes a proteolytic cleavage to generate either a mature or a truncated isoform. Precursor and mature BDNF are assumed to elicit opposing biological effects in neuroplasticity. In the present study we investigated the effect of voluntary physical activity on precursor and mature brain-derived neurotrophic factor levels and on proBDNF cleavage related genes, p11 and tissue plasminogen activator (tPA), as well as the antidepressive and cognitive effects of voluntary physical activity. Mice had access to mobile or locked running wheels for 28 days and were submitted to forced-swim, tail suspension and water maze tests. Their hippocampi were dissected and analyzed by Western blot and real time RT-PCR. Voluntary physical activity, but not locked wheel exposure, induced a robust increase in hippocampal mature BDNF protein levels, as well as in p11 and tPA mRNA expression; and also promoted antidepressive effects and improved learning, when compared with sedentary mice. On the other hand, there were no significant differences between any groups in the expression of precursor or truncated isoforms of BDNF. Our data suggest that the antidepressive effect of the physical exercise may depend, at least in part, on changes in BDNF post-translational processing.

Effects of systemic administration of ciliary neurotrophic factor on Bax and Bcl-2 proteins in the lumbar spinal cord of neonatal rats after sciatic nerve transection.

Ciliary neurotrophic factor (CNTF) is a cytokine that plays a neuroprotective role in relation to axotomized motoneurons. We determined the effect of daily subcutaneous doses of CNTF (1.2 microg/g for 5 days; N = 13) or PBS (N = 13) on the levels of mRNA for Bcl-2 and Bax, as well as the expression and inter-association of Bcl-2 and Bax proteins, and the survival of motoneurons in the spinal cord lumbar enlargement of 2-day-old Wistar rats after sciatic nerve transection. Five days after transection, the effects were evaluated on histological and molecular levels using Nissl staining, immunoprecipitation, Western blot analysis, and reverse transcriptase-polymerase chain reaction. The motoneuron survival ratio, defined as the ratio between the number of motoneurons counted on the lesioned side vs those on the unlesioned side, was calculated. This ratio was 0.77 +/- 0.02 for CNTF-treated rats vs 0.53 +/- 0.02 for the PBS-treated controls (P < 0.001). Treatment with CNTF modified the level of mRNA, with the expression of Bax RNA decreasing 18% (with a consequent decrease in the level of Bax protein), while the expression of Bcl-2 RNA was increased 87%, although the level of Bcl-2 protein was unchanged. The amount of Bcl-2/Bax heterodimer increased 91% over that found in the PBS-treated controls. These data show, for the first time, that the neuroprotective effect of CNTF on neonatal rat axotomized motoneurons is associated with a reduction in free Bax, due to the inhibition of Bax expression, as well as increased Bcl-2/Bax heterodimerization. Thus, the neuroprotective action of the CNTF on axotomized motoneurons can be related to the inhibition of this apoptotic pathway.

Effects of systemic administration of ciliary neurotrophic factor on Bax and Bcl-2 proteins in the lumbar spinal cord of neonatal rats after sciatic nerve transection

Ciliary neurotrophic factor (CNTF) is a cytokine that plays a neuroprotective role in relation to axotomized motoneurons. We determined the effect of daily subcutaneous doses of CNTF (1.2 µg/g for 5 days; N = 13) or PBS (N = 13) on the levels of mRNA for Bcl-2 and Bax, as well as the expression and inter-association of Bcl-2 and Bax proteins, and the survival of motoneurons in the spinal cord lumbar enlargement of 2-day-old Wistar rats after sciatic nerve transection. Five days after transection, the effects were evaluated on histological and molecular levels using Nissl staining, immunoprecipitation, Western blot analysis, and reverse transcriptase-polymerase chain reaction. The motoneuron survival ratio, defined as the ratio between the number of motoneurons counted on the lesioned side vs those on the unlesioned side, was calculated. This ratio was 0.77 ± 0.02 for CNTF-treated rats vs 0.53 ± 0.02 for the PBS-treated controls (P < 0.001). Treatment with CNTF modified the level of mRNA, with the expression of Bax RNA decreasing 18 percent (with a consequent decrease in the level of Bax protein), while the expression of Bcl-2 RNA was increased 87 percent, although the level of Bcl-2 protein was unchanged. The amount of Bcl-2/Bax heterodimer increased 91 percent over that found in the PBS-treated controls. These data show, for the first time, that the neuroprotective effect of CNTF on neonatal rat axotomized motoneurons is associated with a reduction in free Bax, due to the inhibition of Bax expression, as well as increased Bcl-2/Bax heterodimerization. Thus, the neuroprotective action of the CNTF on axotomized motoneurons can be related to the inhibition of this apoptotic pathway.

Avoidance of smoking: the impact of warning labels in Brazil.

BACKGROUND: Research on human emotion shows that pictures drive the activity of specialised brain networks affecting attitude and behaviour. Pictorial warnings on cigarette packages are considered one of the most effective ways to convey information on the health consequences of smoking. However, few studies have evaluated the effectiveness of warning labels to elicit avoidance of smoking. OBJECTIVES: To investigate the impact of pictorial health warnings conveyed by the Brazilian tobacco control programme through a well-established psychometric tool designed for studies on emotion and behaviour. METHODS: Graphic Brazilian cigarette warnings labels were evaluated. They consisted of the two sets of warning pictures displayed in 2002-4 (n = 9) and 2004-8 (n = 10). Pleasant, unpleasant and neutral pictures selected from a standard catalogue were used as controls. Undergraduate students (n = 212, 18% smokers) evaluated the emotional content of each picture in two affective dimensions: hedonic valence and arousal. Participants were not provided with the sources of distinction between control and warning pictures. RESULTS: The judgements of hedonic content of the warning pictures ranged from neutral to very unpleasant. None was classified as highly arousing. Smokers judged warning pictures representing people smoking significantly more pleasant than pictures without smoking scenes, and significantly more so than non-smokers. No significant differences between smokers and non-smokers were found for warning pictures without these smoking scenes. CONCLUSION: Previous studies have shown that the most threatening and arousing pictures prompt the greatest evidence of defensive activation. Emotional ratings of Brazilian warning pictures described them as unpleasant but moderately arousing. To intensify avoidance of the packages, future graphic warnings should therefore generate more arousal. The ratings for the Brazilian warning pictures indicated that, except for those depicting people smoking, judgements by smokers and non-smokers were similar, suggesting a potential applicability in both prevention and cessation. Smoking cues, however, should be avoided.