Repeated PTZ treatment at 25-day intervals leads to a highly efficient accumulation of doublecortin in the dorsal hippocampus of rats.

BACKGROUND: Neurogenesis persists throughout life in the adult mammalian brain. Because neurogenesis can only be assessed in postmortem tissue, its functional significance remains undetermined, and identifying an in vivo correlate of neurogenesis has become an important goal. By studying pentylenetetrazole-induced brain stimulation in a rat model of kindling we accidentally discovered that 25±1 days periodic stimulation of Sprague-Dawley rats led to a highly efficient increase in seizure susceptibility. METHODOLOGY/PRINCIPAL FINDINGS: By EEG, RT-PCR, western blotting and immunohistochemistry, we show that repeated convulsive seizures with a periodicity of 25±1 days led to an enrichment of newly generated neurons, that were BrdU-positive in the dentate gyrus at day 25±1 post-seizure. At the same time, there was a massive increase in the number of neurons expressing the migratory marker, doublecortin, at the boundary between the granule cell layer and the polymorphic layer in the dorsal hippocampus. Some of these migrating neurons were also positive for NeuN, a marker for adult neurons. CONCLUSION/SIGNIFICANCE: Our results suggest that the increased susceptibility to seizure at day 25±1 post-treatment is coincident with a critical time required for newborn neurons to differentiate and integrate into the existing hippocampal network, and outlines the importance of the dorsal hippocampus for seizure-related neurogenesis. This model can be used as an in vivo correlate of neurogenesis to study basic questions related to neurogenesis and to the neurogenic mechanisms that contribute to the development of epilepsy.

Prolonged gaseous hypothermia prevents the upregulation of phagocytosis-specific protein annexin 1 and causes low-amplitude EEG activity in the aged rat brain after cerebral ischemia.

In aged humans, stroke is a major cause of disability for which no neuroprotective measures are available. In animal studies of focal ischemia, short-term hypothermia often reduces infarct size. Nevertheless, efficient neuroprotection requires long-term, regulated lowering of whole-body temperature. Previously, we reported that post-stroke exposure to hydrogen sulfide (H(2)S) effectively lowers whole-body temperature and confers neuroprotection in aged animals. In the present study using magnetic resonance imaging, electroencephalogram recording, DNA arrays, reverse transcriptase polymerase chain reaction, western blotting and immunofluorescence, we characterized the central nervous system response to H(2)S-induced hypothermia and report, for the first time, that annexin A1, a major pro-inflammatory protein that is upregulated after stroke, was consistently downregulated in polymorphonuclear cells in the peri-lesional cortex of post-ischemic, aged rat brain after 48 hours of hypothermia induced by exposure to H(2)S. Our data suggest that long-term hypothermia may be a viable clinical approach to protecting the aged brain from cerebral injury. Our findings further suggest that, in contrast to monotherapies that have thus far uniformly failed in clinical practice, hypothermia has pleiotropic effects on brain physiology that may be necessary for effective protection of the brain after stroke.

Brain Aging and Regeneration after Injuries: an Organismal approach.

Aging is associated with a decline of locomotor, sensory and cognitive performance in humans and experimental animals. The rate and pattern of organismal senescence may be regulated in part by changes in multiple genes involved in multiple processes. While this theory is supported by genetic data in lower organisms, a lack of direct experimental evidence in higher organisms has contributed to a broader acceptance of the "stochastic aging" model, in which accumulating, random damaging biological events play an important role. However, these insults alone cannot account for the inexorable deterioration and loss of function that characterizes aging. The higher the complexity of a system, the less obvious is the effect of genetic regulation on aging and the life span, indicating that epigenetic factors play an important role in aging. Most importantly, we present evidence that aging systems do retain some capacity for regeneration and functional recovery after injuries to the central nervous system like cerebral ischemia.

Long-term hypothermia reduces infarct volume in aged rats after focal ischemia.

UNLABELLED: In aged humans, stroke is a major cause of disability for which no neuroprotective measures are available. A viable alternative to conventional drug-based neuroprotective therapies is brain/body cooling, or hypothermia. In animal studies of focal ischemia, short-term hypothermia consistently reduces infarct size. Nevertheless, efficient neuroprotection requires long-term, regulated lowering of whole body temperature. Focal cerebral ischemia was produced by reversible occlusion of the right middle cerebral artery in 17-month-old male Sprague-Dawley rats. After stroke, the aged rats were exposed for 2 days to a mixture of air and a mild inhibitor of oxidative phosphorylation, hydrogen sulfide (H(2)S), which resulted in sustained, deep hypothermia (30.8+/-0.7 degrees C). Long-term hypothermia led to a 50% reduction in infarct size with a concomitant reduction in the number of phagocytic cells. At the transcription level, hypothermia caused a reduction in the mRNA coding for caspase 12, NF-kappa B and grp78 in the peri-infarcted region, suggesting an overall decrease in the transcriptional activity related to inflammation and apoptosis. Behaviorally, hypothermia was associated with better performance on tests that require complex sensorimotor skills, in the absence of obvious neurological deficits or physiological side effects, in aged rats. CONCLUSIONS: Prolonged, H(2)S-induced hypothermia is a simple and efficacious method to limit the damage inflicted by stroke in aged rats.

Premature cellular proliferation following cortical infarct in aged rats.

Old age is associated with an enhanced susceptibility to stroke and poor recovery from brain injury, but the cellular mechanisms underlying such phenomena are not known. Using BrdU-labeling, quantitative immunohistochemistry and 3D-reconstruction of confocal images in a rat model of mild cerebral ischemia, we found that aged rats are highly susceptible to develop an early infarct that is associated with premature cellular proliferation originating from the vascular tree. In aged rats we also found a rapid delimitation of the infarct area by capillary-derived neuroepithelial cells and an early incorporation of these cells into the glial scar. Since most proliferating cells at the infarct site are microglia or nestin-positive cells derived from the vascular wall, we conclude that the vasculature plays a hitherto unrecognized role as a source of proliferating neuroepithelial cells after stroke. Age-associated alterations in the timing and origin of the cytogenic response to cerebral ischemia may underlie the poor functional recovery from stroke. Clarifying the molecular basis of these phenomena could yield novel approaches to enhancing neurorestoration in the elderly. Studies of stroke in experimental animals have demonstrated the neuroprotective efficacy of a variety of interventions, but most of the strategies that have been clinically tested failed to show benefit in aged humans. One possible explanation for this discrepancy between laboratory and clinical investigations is the role that age plays in the recovery of the brain from insult. Although it is well known that aging is a risk factor for stroke (Barnett HJ, 2002), the majority of experimental studies of stroke have been performed on young animals, and therefore may not fully replicate the effects of ischemia on neural tissue in aged subjects (Wang LC et al., 1995; Davies M et al., 1995; Sutherland GR et al., 1996; Popa-Wagner A et al., 1998, 1999a). Hence, the aged post-acute animal model is clinically most relevant to stroke rehabilitation and cellular studies (Lindner MD et al., 2003; Brown AW et al., 2003; Badan I et al., 2003).