Cristina Richichi

Neuron-restrictive silencer factor-mediated hyperpolarization-activated cyclic nucleotide gated channelopathy in experimental temporal lobe epilepsy.

Enduring, abnormal expression and function of the ion channel hyperpolarization‐activated cyclic adenosine monophosphate gated channel type 1 (HCN1) occurs in temporal lobe epilepsy (TLE). We examined the underlying mechanisms, and investigated whether interfering with these mechanisms could modify disease course.

Localization of HCN1 Channels to Presynaptic Compartments: Novel Plasticity That May Contribute to Hippocampal Maturation

Increasing evidence supports roles for the current mediated by hyperpolarization-activated cyclic nucleotide-gated (HCN) channels, Ih, in hippocampal maturation and specifically in the evolving changes of intrinsic properties as well as network responses of hippocampal neurons. Here, we describe a novel developmental plasticity of HCN channel expression in axonal and presynaptic compartments: HCN1 channels were localized to axon terminals of the perforant path (the major hippocampal afferent pathway) of immature rats, where they modulated synaptic efficacy. However, presynaptic expression and functions of the channels disappeared with maturation. This was a result of altered channel transport to the axons, because HCN1 mRNA and protein levels in entorhinal cortex neurons, where the perforant path axons originate, were stable through adulthood. Blocking action potential firing in vitro increased presynaptic expression of HCN1 channels in the perforant path, suggesting that network activity contributed to regulating this expression. These findings support a novel developmentally regulated axonal transport of functional ion channels and suggest a role for HCN1 channel-mediated presynaptic Ih in hippocampal maturation.

Activity-dependent heteromerization of the hyperpolarization-activated, cyclic-nucleotide gated (HCN) channels: role of N-linked glycosylation

Formation of heteromeric complexes of ion channels via co‐assembly of different subunit isoforms provides an important mechanism for enhanced channel diversity. We have previously demonstrated co‐association of the hyperpolarization activated cyclic‐nucleotide gated (HCN1/HCN2) channel isoforms that was regulated by network (seizure) activity in developing hippocampus. However, the mechanisms that underlie this augmented expression of heteromeric complexes have remained unknown. Glycosylation of the HCN channels has been implicated in the stabilization and membrane expression of heteromeric HCN1/HCN2 constructs in heterologous systems. Therefore, we used in vivo and in vitro systems to test the hypothesis that activity modifies HCN1/HCN2 heteromerization in neurons by modulating the glycosylation state of the channel molecules. Seizure‐like activity (SA) increased HCN1/HCN2 heteromerization in hippocampus in vivo as well as in hippocampal organotypic slice cultures. This activity increased the abundance of glycosylated HCN1 but not HCN2‐channel molecules. In addition, glycosylated HCN1 channels were preferentially co‐immunoprecipitated with the HCN2 isoforms. Provoking SA in vitro in the presence of the N‐linked glycosylation blocker tunicamycin abrogated the activity‐dependent increase of HCN1/HCN2 heteromerization. Thus, hippocampal HCN1 molecules have a significantly higher probability of being glycosylated after SA, and this might promote a stable heteromerization with HCN2.

Functional role of proinflammatory and anti-inflammatory cytokines in seizures

Recent evidence has shown that proinflammatory and anti-inflammatory molecules are synthesized during epileptic activity in glial cells in CNS regions where seizures initiate and spread. These molecules are released and interact with specific receptors on neurons. Since various cytokines have been shown to affect neuronal excitability, this led to the hypothesis that they may have a role in altering synaptic transmission in epileptic conditions. Indeed, intracerebral application of IL-1beta enhances epileptic activity in experimental models while its naturally occurring receptor antagonist (IL-1Ra) mediates anticonvulsant actions. Transgenic mice overexpressing IL-1Ra in astrocytes are less susceptible to seizures, indicating that endogenous IL-1 has proconvulsant activity. Several studies indicate a central role of IL-1beta for the exacerbation of brain damage after ischemic, traumatic or excitotoxic insults, suggesting that it may also contribute to neuronal cell injury associated with seizures. Finally, a functional polymorphism in the IL-1beta gene promoter, possibly associated with enhanced ability to produce this cytokine, has been specifically found in temporal lobe epilepsy patients with hippocampal sclerosis and in children with febrile seizures. Thus, the IL-1 system may represent a novel target for controlling seizure activity and/or the associated long-term sequelae. Furthermore, these studies suggest that other inflammatory and anti-inflammatory molecules produced in the CNS may have a role in the pathophysiology of seizure disorders.

The transcription factor NRSF contributes to epileptogenesis by selective repression of a subset of target genes.

The mechanisms generating epileptic neuronal networks following insults such as severe seizures are unknown. We have previously shown that interfering with the function of the neuron-restrictive silencer factor (NRSF/REST), an important transcription factor that influences neuronal phenotype, attenuated development of this disorder. In this study, we found that epilepsy-provoking seizures increased the low NRSF levels in mature hippocampus several fold yet surprisingly, provoked repression of only a subset (∼10%) of potential NRSF target genes. Accordingly, the repressed gene-set was rescued when NRSF binding to chromatin was blocked. Unexpectedly, genes selectively repressed by NRSF had mid-range binding frequencies to the repressor, a property that rendered them sensitive to moderate fluctuations of NRSF levels. Genes selectively regulated by NRSF during epileptogenesis coded for ion channels, receptors, and other crucial contributors to neuronal function. Thus, dynamic, selective regulation of NRSF target genes may play a role in influencing neuronal properties in pathological and physiological contexts. DOI: http://dx.doi.org/10.7554/eLife.01267.001

Plasticity of somatostatin and somatostatin sst2A receptors in the rat dentate gyrus during kindling epileptogenesis

Increasing evidence suggests that somatostatin may control neuronal excitability during epileptogenesis. In the hippocampus, sst2A receptors are likely to mediate somatostatin inhibitory actions but little is known about their status in kindled tissues. In the present study, sst2A receptor and somatostatin immunoreactivity were examined by confocal microscopy in the hippocampus during and after kindling acquisition. In control rats, somatostatin‐positive axon terminals were mainly found in the stratum lacunosum moleculare of CA1 area and in the outer molecular layer of the dentate gyrus. sst2A receptor immunoreactivity was diffusely distributed in the strata radiatum and oriens of CA1 and in the stratum moleculare of the dentate gyrus. Immunogold electron microscopy revealed that sst2A receptors were predominantly localized postsynaptically, at the plasma membrane of dendritic shafts and spines of principal neurons. During kindling epileptogenesis, qualitative and semiquantitative analysis revealed a progressive decrease of sst2A immunoreactivity in the outer molecular layer, which was spatially associated with an increase in somatostatin immunoreactivity. No obvious changes in sst2A receptor immunoreactivity were observed in other hippocampal subfields. These results suggest that the decrease of sst2A receptor immunoreactivity in the outer molecular layer reflects receptor down‐regulation in distal dendrites of granule cells in response to chronic somatostatin release. Because the sst2A receptor appears to mediate anticonvulsant and antiepileptogenic effects of somatostatin, this may represent a pivotal mechanism contributing to epileptogenesis.