Faculty Bibliography

Vascular endothelial growth factor (VEGF) is a protein factor which has been found to play a significant role in both normal and pathological states. Its role as an angiogenic factor is well-established. More recently, VEGF has been shown to protect neurons from cell death both in vivo and in vitro. While VEGF's potential as a protective factor has been demonstrated in hypoxia-ischemia, in vitro excitotoxicity, and motor neuron degeneration, its role in seizure-induced cell loss has received little attention. A potential role in seizures is suggested by Newton et al.'s [Newton SS, Collier EF, Hunsberger J, Adams D, Terwilliger R, Selvanayagam E, Duman RS (2003) Gene profile of electroconvulsive seizures: Induction of neurotrophic and angiogenic factors. J Neurosci 23:10841-10851] finding that VEGF mRNA increases in areas of the brain that are susceptible to cell loss after electroconvulsive-shock induced seizures. Because a linear relationship does not always exist between expression of mRNA and protein, we investigated whether VEGF protein expression increased after pilocarpine-induced status epilepticus. In addition, we administered exogenous VEGF in one experiment and blocked endogenous VEGF in another to determine whether VEGF exerts a neuroprotective effect against status epilepticus-induced cell loss in one vulnerable brain region, the rat hippocampus. Our data revealed that VEGF is dramatically up-regulated in neurons and glia in hippocampus, thalamus, amygdala, and neocortex 24 h after status epilepticus. VEGF induced significant preservation of hippocampal neurons, suggesting that VEGF may play a neuroprotective role following status epilepticus

In adult female rats, robust hippocampal changes occur when estradiol rises on the morning of proestrus. Whether estradiol mediates these changes, however, remains unknown. To address this issue, we used sequential injections of estradiol to simulate two key components of the preovulatory surge: the rapid rise in estradiol on proestrous morning, and the slower rise during the preceding day, diestrus 2. Animals were examined mid-morning of simulated proestrus, and compared to vehicle-treated or intact rats. In both simulated and intact rats, CA1-evoked responses were potentiated in hippocampal slices, and presynaptic mechanisms appeared to contribute. In CA3, multiple population spikes were evoked in response to mossy fiber stimuli, and expression of brain-derived neurotrophic factor was increased. Simulation of proestrous morning also improved performance on object and place recognition tests, in comparison to vehicle treatment. Surprisingly, effects on CA1-evoked responses showed a dependence on estradiol during simulated diestrus 2, as well as a dependence on proestrous morning. Increasing estradiol above the physiological range on proestrous morning paradoxically decreased evoked responses in CA1. However, CA3 pyramidal cell activity increased further, and became synchronized. Together, the results confirm that physiological estradiol levels are sufficient to profoundly affect hippocampal function. In addition: (i) changes on proestrous morning appear to depend on slow increases in estradiol during the preceding day; (ii) effects are extremely sensitive to the peak serum level on proestrous morning; and (iii) there are striking subfield differences within the hippocampus

Characterizing the responses of different mouse strains to experimentally-induced seizures can provide clues to the genes that are responsible for seizure susceptibility, and factors that contribute to epilepsy. This approach is optimal when sequenced mouse strains are available. Therefore, we compared two sequenced strains, DBA/2J (DBA) and A/J. These strains were compared using the chemoconvulsant pilocarpine, because pilocarpine induces status epilepticus, a state of severe, prolonged seizures. In addition, pilocarpine-induced status is followed by changes in the brain that are associated with the pathophysiology of temporal lobe epilepsy (TLE). Therefore, pilocarpine can be used to address susceptibility to severe seizures, as well as genes that could be relevant to TLE. A/J mice had a higher incidence of status, but a longer latency to status than DBA mice. DBA mice exhibited more hippocampal pyramidal cell damage. DBA mice developed more ectopic granule cells in the hilus, a result of aberrant migration of granule cells born after status. DBA mice experienced sudden death in the weeks following status, while A/J mice exhibited the most sudden death in the initial hour after pilocarpine administration. The results support previous studies of strain differences based on responses to convulsants. They suggest caution in studies of seizure susceptibility that are based only on incidence or latency. In addition, the results provide new insight into the strain-specific characteristics of DBA and A/J mice. A/J mice provide a potential resource to examine the progression to status. The DBA mouse may be valuable to clarify genes regulating other seizure-associated phenomena, such as seizure-induced neurogenesis and sudden death

Granule cell neurogenesis increases following seizures, and some newly born granule cells develop at abnormal locations within the hilus. These ectopic granule cells (EGCs) demonstrate regular bursts of action potentials that are synchronized with CA3 pyramidal cell burst discharges and the bursts of hilar neurons, including mossy cells. Such findings suggest that mossy cells may participate in circuits that activate EGCs. Electron microscopic immunolabeling was therefore used to determine if mossy cell axon terminals form synapses with hilar EGC dendrites, using animals that underwent pilocarpine-induced status epilepticus. Pilocarpine was administered to adult male rats, and those which developed status epilepticus were perfused 5-7 months later, after the period of EGC genesis. Hippocampal sections were processed for dual electron microscopic immunolabeling (using calcitonin gene-related peptide (CGRP) as a marker for mossy cells and calbindin (CaBP) as a marker for EGCs). Light microscopic analysis revealed large CGRP-immunoreactive cells in the hilus, with the appearance and distribution of mossy cells. Electron microscopic analysis revealed numerous CaBP-immunoreactive dendrites in the hilus, some of which were innervated by CGRP-immunoreactive terminals. The results suggest that mossy cells participate in the excitatory circuits which activate EGCs, providing further insight into the network rearrangements that accompany seizure-induced neurogenesis in this animal model of epilepsy

Glial cells provide energy substrates to neurons, in part from glycogen metabolism, which is influenced by glycogen phosphorylase (GP). To gain insight into the potential subfield and laminar-specific expression of GP, histochemistry can be used to evaluate active GP (GPa) or totalGP (GPa + GPb). Using this approach, we tested the hypothesis that changes in GP would occur under pathological conditions that are associated with increased energy demand, i.e. severe seizures (status epilepticus or 'status'). We also hypothesized that GP histochemistry would provide insight into changes in the days and weeks after status, particularly in the hippocampus and entorhinal cortex, where there are robust changes in structure and function. One hour after the onset of pilocarpine-induced status, GPa staining was reduced in most regions of the hippocampus and entorhinal cortex relative to saline-injected controls. One week after status, there was increased GPa and totalGP, especially in the inner molecular layer, where synaptic reorganization of granule cell mossy fibre axons occurs (mossy fibre sprouting). In addition, patches of dense GP reactivity were evident in many areas. One month after status, levels of GPa and totalGP remained elevated in some areas, suggesting an ongoing role of GP or other aspects of glycogen metabolism, possibly due to the evolution of intermittent, recurrent seizures at approximately 3-4 weeks after status. Taken together, the results suggest that GP is dynamically regulated during and after status in the adult rat, and may have an important role in the pilocarpine model of epilepsy

Epilepsy is a complex disease with diverse clinical characteristics that preclude a singular mechanism. One way to gain insight into potential mechanisms is to reduce the features of epilepsy to its basic components: seizures, epileptogenesis, and the state of recurrent unprovoked seizures that defines epilepsy itself. A common way to explain seizures in a normal individual is that a disruption has occurred in the normal balance of excitation and inhibition. The fact that multiple mechanisms exist is not surprising given the varied ways the normal nervous system controls this balance. In contrast, understanding seizures in the brain of an individual with epilepsy is more difficult because seizures are typically superimposed on an altered nervous system. The different environment includes diverse changes, making mechanistic predictions a challenge. Understanding the mechanisms of seizures in an individual with epilepsy is also more complex than understanding the mechanisms of seizures in a normal individual because epilepsy is not necessarily a static condition but can continue to evolve over the lifespan. Using temporal lobe epilepsy as an example, it is clear that genes, developmental mechanisms, and neuronal plasticity play major roles in creating a state of underlying hyperexcitability. However, the critical control points for the emergence of chronic seizures in temporal lobe epilepsy, as well as their persistence, frequency, and severity, are questions that remain unresolved

We have shown that neuropeptide Y (NPY) regulates neurogenesis in the normal dentate gyrus (DG) via Y(1) receptors (Howell, O.W., Scharfman, H.E., Herzog, H., Sundstrom, L.E., Beck-Sickinger, A. and Gray, W.P. (2003) Neuropeptide Y is neuroproliferative for post-natal hippocampal precursor cells. J Neurochem, 86, 646-659; Howell, O.W., Doyle, K., Goodman, J.H., Scharfman, H.E., Herzog, H., Pringle, A., Beck-Sickinger, A.G. and Gray, W.P. (2005) Neuropeptide Y stimulates neuronal precursor proliferation in the post-natal and adult dentate gyrus. J Neurochem, 93, 560-570). This regulation may be relevant to epilepsy, because seizures increase both NPY expression and precursor cell proliferation in the DG. Therefore, the effects of NPY on DG precursors were evaluated in normal conditions and after status epilepticus. In addition, potentially distinct NPY-responsive precursors were identified, and an analysis performed not only of the DG, but also the caudal subventricular zone (cSVZ) and subcallosal zone (SCZ) where seizures modulate glial precursors. We show a proliferative effect of NPY on multipotent nestin cells expressing the stem cell marker Lewis-X from both the DG and the cSVZ/SCZ in vitro. We confirm an effect on proliferation in the cSVZ/SCZ of Y(1) receptor(-/-) mice and demonstrate a significant reduction in basal and seizure-induced proliferation in the DG of NPY(-/-) mice

Granule cells of the mammalian dentate gyrus normally form a discrete layer, and virtually all granule cells migrate to this location. Exceptional granule cells that are positioned incorrectly, in 'ectopic' locations, are rare. Although the characteristics of such ectopic granule cells appear similar in many respects to granule cells located in the granule cell layer, their rare occurrence has limited a full evaluation of their structure and function. More information about ectopic granule cells has been obtained by studying those that develop after experimental manipulations that increase their number. For example, after severe seizures, the number of ectopic granule cells located in the hilus increases dramatically. These experimentally-induced ectopic granule cells may not be equivalent to normal ectopic granule cells necessarily, but the vastly increased numbers have allowed much more information to be obtained. Remarkably, the granule cells that are positioned ectopically develop intrinsic properties and an axonal projection that are similar to granule cells that are located normally, i.e., in the granule cell layer. However, dendritic structure and synaptic structure/function appear to differ. These studies have provided new insight into a rare type of granule cell in the dentate gyrus, and the plastic characteristics of dentate granule cells that appear to depend on the location of the cell body