Research ReportHippocampal seizures cause depolymerization of filamentous actin in neurons independent of acute morphological changes
Introduction
Patients with epilepsy often suffer from significant neurocognitive deficits, such as memory impairment, which may, in part, be caused by direct injurious effects of seizures on the brain (Dodrill, 2002, Elger et al., 2004). Many previous studies indicate that seizures can directly cause “excitotoxic” cell injury and neuronal death under some conditions (Holmes, 2002, Sutula et al., 2003); however, seizures may also induce more subtle “non-lethal” pathophysiological changes in neuronal structure and function, such as abnormalities in synaptic transmission. Dendritic spines receive a majority of the excitatory synaptic inputs to cortical neurons and are strongly implicated in mechanisms of synaptic plasticity and learning (Carlise and Kennedy, 2005, Segal, 2005). Studies of human epilepsy and animal models demonstrate that seizures may directly affect the morphological and functional properties of dendritic spines (reviewed in Swann et al., 2000, Wong, 2005), suggesting that seizure-related changes in spines may represent a mechanistic basis for cognitive deficits in epilepsy.
The molecular mechanisms by which physiological and pathological stimuli modulate dendritic spine structure and function are incompletely understood, but may involve regulation of the actin cytoskeleton (Carlise and Kennedy, 2005). Filamentous actin (F-actin), which is highly concentrated in dendritic spines (Matus et al., 1982, Allison et al., 1998, Capani et al., 2001), forms complex networks that provide structural support for dendrites and dendritic spines and also mediate non-structural spine functions, such as biochemical compartmentalization, gating of protein translocation into spines, and receptor trafficking and anchoring (Allison et al., 1998, Kennedy et al., 2005, Oertner and Matus, 2005, Ouyang et al., 2005). Previous studies have shown that physiological activation of neurons or induction of long-term potentiation (LTP) can cause changes in actin dynamics that may mediate synaptic plasticity (Kim and Lisman, 1999, Krucker et al., 2000, Fukazawa et al., 2003, Okamoto et al., 2004, Lin et al., 2005, Ouyang et al., 2005, Kramar et al., 2006). This activity-dependent regulation of actin dynamics involves modulation of actin-binding proteins, such as cofilin (Sarmiere and Bamburg, 2004), which causes changes in the polymerization state of actin and may result in either structural changes or non-structural, functional changes in dendritic spines.
In contrast to the regulation of actin dynamics by physiological stimuli, relatively little is known about the effects of more pathological activity, such as seizures, on actin networks. In the present study, we have investigated the short-term effects of pathophysiological seizure stimuli, as well as elevated potassium, on actin and cofilin dynamics in the mouse hippocampus in vivo and in vitro and examined by modern cellular imaging methods whether there is a gross structural correlate to these effects in dendritic spines. We report that seizures can modulate actin and cofilin dynamics acutely, although the ultimate structural or functional consequences of these changes may be dependent on the type and severity of the pathophysiological stimulus.
Section snippets
Hippocampal seizures in vivo and in vitro cause depolymerization of F-actin
For in vivo seizure experiments, stereotypical clinical-electrographic seizures were induced by injection of 4-aminopyridine (4-AP) into the CA1 region of hippocampus of mice (Fig. 1A). Behaviorally, within minutes of recovery from anesthesia after 4-AP injection, mice exhibited rapidly repetitive or continuous clinical symptoms, including head nodding, forearm clonus, rearing and falling, and occasional wild running and generalized convulsive activity (Racine stage 5), lasting 1 h until the
Discussion
Studies of both human epilepsy and animal models suggest that seizures may exert pathophysiological effects on dendritic spines (Swann et al., 2000, Wong, 2005), which could contribute to cognitive deficits of epilepsy patients. Although previous studies have demonstrated that seizures may activate a variety of types and mechanisms of brain injury involving dendrites, in the present study we focused on the hypothesis that seizures might disrupt dendritic structure or function, at least in part,
Animals and reagents
Two- to three-month-old male C57/BL6 wild-type mice (Charles River) and transgenic mice (with a C57/Bl6 background) expressing green fluorescent protein (GFP) in subpopulations of hippocampal neurons (line GFP-M, Feng et al., 2000) were used in this study. Wild-type C57/Bl6 mice were used in experiments for rhodamine-phalloidin labeling and Western blot analysis of actin and cofilin. GFP-M transgenic mice were utilized in imaging experiments involving analysis of dendritic spine density. Care
Acknowledgments
The authors thank Nicholas Rensing for technical assistance with some of these experiments. This work was supported by NIH K02 NS045583 (MW) and the Alafi Family Foundation.
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