New neurons in old brains: implications of age in the analysis of neurogenesis in post-mortem tissue.

Adult neurogenesis, the proliferation and integration of newly generated neurons, has been observed in the adult mammalian hippocampus of many species. Numerous studies have also found adult neurogenesis in the human hippocampus, but several recent high-profile studies have suggested that this process is considerably reduced in humans, occurring in children but not in adults. In comparison, rodent studies also show age-related decline but a greater degree of proliferation of new neurons in adult animals. These differences may represent biological species differences or could alternatively be explained by methodological differences in tissue handling and fixation. Here, we examine whether differences in the post-mortem interval between death and tissue fixation might impact subsequent detection of adult neurogenesis due to increased tissue degradation. Because there are fewer new neurons present in older subjects to begin with we hypothesized that, subject age might interact significantly with post-mortem interval in the detection of adult neurogenesis. We analyzed neurogenesis in the hippocampus of rats that were either perfusion-fixed or the brains extracted and immersion-fixed at various post-mortem intervals. We observed an interaction between animal age and the time delay between death and tissue fixation. While similar levels of neurogenesis were observed in young rats regardless of fixation, older rats had significantly fewer labeled neurons when fixation was not immediate. Furthermore, the morphological detail of the labeled neurons was significantly reduced in the delayed fixation conditions at all ages. This study highlights critical concerns that must be considered when using post-mortem tissue to quantify adult neurogenesis.

Assessment of brain oxygenation imbalance following soman exposure in rats.

Nerve agents (NAs) are potent organophosphorus (OP) compounds with applications in chemical warfare. OP compounds act by inhibiting acetylcholinesterase (AChE). Soman (O-pinacolyl methylphosphonofluoridate) is one of the most potent NAs. It is well known that small doses of NAs can be lethal, and that even non-lethal exposure leads to long-term mental debilitation/neurological damage. However, the neuropathology following exposure to sub-lethal nerve agents is not well understood. In this study, we examined changes in tissue oxygenation (pO2) in the cortex and hippocampus after a sub-lethal dose of soman [80-90 µg/kg; subcutaneous]. pO2 changes can provide information regarding oxygen delivery and utilization and may be indicative of a disruption in cerebral blood flow and/or metabolism. Changes in oxygenation were measured with chronically implanted oxygen sensors in awake and freely moving rats. Measurements were taken before, during, and after soman-induced convulsive seizures. Soman exposure resulted in an immediate increase in pO2 in the cortex, followed by an even greater increase that precedes the onset of soman-induced convulsive seizures. The rise in hippocampus pO2 was delayed relative to the cortex, although the general pattern of brain oxygenation between these two regions was similar. After convulsive seizures began, pO2 levels declined but usually remained hyperoxygenated. Following the decline in pO2, low frequency cycles of large amplitude changes were observed in both the cortex and hippocampus. This pattern is consistent with recurring seizures. Measuring real-time changes in brain pO2 provides new information on the physiological status of the brain following soman exposure. These results highlight that the measurement of brain oxygenation could provide a sensitive marker of nerve agent exposure and serve as a biomarker for treatment studies.

Increased excitability and molecular changes in adult rats after a febrile seizure.

Both early life inflammation and prolonged febrile seizures have been associated with increased excitation in the adult brain. We hypothesized this may be due in part to changes in the cation-chloride cotransporter system. Rat pups received saline or lipopolysaccharide/kainic acid (LPS/KA) resulting in inflammation, followed by a behavioral febrile seizure (FS) in approximately 50% of rats. Adult animals from the saline, inflammation, or inflammation + FS groups underwent the following: (1) in vitro electrophysiologic studies; (2) Western blotting or polymerase chain reaction; or (3) application of the Na-K-Cl cotransporter 1 (NKCC1) blocker bumetanide to determine its effect on reversing increased excitability in vitro. The inflammation and inflammation + FS groups demonstrated increased excitability in vitro and increased hippocampal protein expression of NR2B and GABAA α5 receptor subunits and mRNA expression of NKCC1. The inflammation + FS group also had decreased protein expression of GluR2 and GABAA α1 receptor subunits and mRNA and protein expression of KCC2. Bumetanide decreased in vitro 4-aminopyridine-induced inter-ictal activity in the inflammation and inflammation + FS groups. The results demonstrate early-life inflammation with or without a behavioral FS can lead to long-lasting molecular changes and increased excitability in the adult rat hippocampus, although some changes are more extensive when inflammation is accompanied by behavioral seizure activity. Bumetanide is effective in reversing increased excitability in vitro, providing evidence for a causal role for cation-chloride cotransporters and suggesting this drug may prove useful for treating epilepsy that develops after a FS.

Changes in (14)C-labeled 2-deoxyglucose brain uptake from nickel-induced epileptic activity.

Unilateral epidural applications of nickel solution to motor cortex were followed in about 1 h by contralateral forelimb myoclonus. In rats which displayed frequent myoclonal jerking during the 45-min 2-deoxyglucose (2-DG) uptake and clearing period, autoradiographic analysis showed that glucose utilization at the nickel implant site was greater in the supragranular and infragranular layers than in the granular layer (in normal cortex, activity is greatest in the granular layer), and was also greater in the substantia nigra and other subcortical centers. The same cortical and most of the subcortical changes in 2-DG uptake were also observed when metabolic activity was assessed 1 h after myoclonus had stopped, indicating that it may not have been the seizure activity itself that had altered metabolic activity, but some process engendered by the seizures - possibly a tissue response to excitotoxic damage. In fact, rats which displayed infrequent myoclonus showed negligible increases in cortical and subcortical uptake. These results do not support an earlier claim that increased glucose consumption is the metabolic signature of the interictal activity produced by seizure-inducing metals. Indeed, the findings raise the possibility that tissue damage is responsible for interictal hypermetabolism when it is observed in animal models of epilepsy.