ABSTRACT
Increased dosage of methyl-CpG-binding protein-2 (MeCP2) results in a dramatic neurodevelopmental phenotype with onset at birth. We generated induced pluripotent stem cells (iPSCs) from patients with the MECP2 duplication syndrome (MECP2dup), carrying different duplication sizes, to study the impact of increased MeCP2 dosage in human neurons. We show that cortical neurons derived from these different MECP2dup iPSC lines have increased synaptogenesis and dendritic complexity. In addition, using multi-electrodes arrays, we show that neuronal network synchronization was altered in MECP2dup-derived neurons. Given MeCP2 functions at the epigenetic level, we tested whether these alterations were reversible using a library of compounds with defined activity on epigenetic pathways. One histone deacetylase inhibitor, NCH-51, was validated as a potential clinical candidate. Interestingly, this compound has never been considered before as a therapeutic alternative for neurological disorders. Our model recapitulates early stages of the human MECP2 duplication syndrome and represents a promising cellular tool to facilitate therapeutic drug screening for severe neurodevelopmental disorders.
Subject(s)
Methyl-CpG-Binding Protein 2/genetics , Methyl-CpG-Binding Protein 2/physiology , Nerve Net/metabolism , Cell Differentiation , Dendrites/metabolism , Gene Dosage/physiology , Gene Duplication/genetics , Genetic Association Studies , Humans , Induced Pluripotent Stem Cells , Male , Neurogenesis , NeuronsABSTRACT
Proechimys (Rodentia: Echimyidae) is a neotropical rodent of the Amazon region that has been successfully colonized in the laboratory and used for experimental medicine. Preliminary studies indicated that Proechimys (casiragua) rodents express an atypical resistance to developing a chronic epileptic condition in common models of temporal lobe epilepsy. Moreover, previous investigation of our laboratory described a remarkably different Proechimy's cytoarchitecture organization of the hippocampal CA2 subfield. In the present study, we investigated the intrinsic neuronal properties and morphological characteristics of the Proechimys's hippocampal pyramidal neurons of the CA1 and CA2 areas. A comparative approach was performed using neurons recorded in Wistar rats. A striking finding in Proechimys rodents was the presence of large pyramidal-like neurons throughout the stratum oriens from CA2 to CA1 area. In order to confirm such distinctive feature of the Proechimys's hippocampus, we performed Nissl staining and immunohistochemistry for neurofilament protein SM311. CA2 pyramidal neurons in the stratum pyramidale of Proechimys exhibited a significantly higher input resistance and lower time constant when compared to corresponding cell groups in the same area of the Wistar rat's. This newly identified population of pyramidal-shaped neurons in stratum oriens of Proechimys exhibited distinct electrophysiological and morphological properties. This included larger capacitance, lower input resistance, larger rheobase, long latency to first action potential and slower firing frequency. In addition, the apical dendrites of these neurons were oriented in parallel to apical dendrites of regular pyramidal neurons in stratum pyramidale. Moreover, these neurons were immunoreactive to SM311 as the majority of the neurons of the pyramidal layer. The functional role of these hippocampal neurons of the rodent Proechimys deserves further investigation.
Subject(s)
CA1 Region, Hippocampal/cytology , CA1 Region, Hippocampal/physiology , CA2 Region, Hippocampal/cytology , CA2 Region, Hippocampal/physiology , Pyramidal Cells/physiology , Rodentia/physiology , Action Potentials/physiology , Animals , Electrophysiology/methods , Male , Neural Pathways/physiology , Neural Pathways/ultrastructure , Organ Culture Techniques , Pyramidal Cells/cytology , Rats , Rats, Wistar , Species Specificity , Synapses/physiology , Synapses/ultrastructureABSTRACT
Previous data of our laboratory have shown that the Amazonian rodents Proechimys do not present spontaneous seizures in different models of epilepsy, suggesting endogenous inhibitory mechanisms. Here, we describe a remarkably different Proechimy's cytoarchitecture organization of the hippocampal cornu Ammonis 2 (CA2) subfield. We identified a very distinctive Proechimy's CA2 sector exhibiting disorganized cell presentation of the pyramidal layer and atypical dispersion of the pyramidal-like cells to the stratum oriens, strongly contrasting to the densely packed CA2 cells in the Wistar rats. Studies showed that CA2 is the only cornu ammonis (CA) subfield resistant to the extensive pyramidal neural loss in mesial temporal lobe epilepsy (MTLE) associated to hippocampal sclerosis. Thus, in order to investigate this region, we used Nissl and Timm staining, stereological approach to count neurons and immunohistochemistry to neuronal nuclei (NeuN), parvalbumin (PV), calbindin (CB) and calretinin (CR). We did not notice statistically significant differences in the total number of neurons of the CA2 region between Proechimys and Wistar. However, Proechimys rodents presented higher CA2 volume than Wistar rats. Furthermore, no significant difference in the optical density of parvalbumin-immunoreactivity was found between subject groups. On the other hand, Proechimys presented significant higher density of calbindin and calretinin-immunoreactivity when compared to Wistar rats. In this context, this unique CA2 subfield seen in Proechimys opens up a new set of possibilities to explore the contribution of CA2 neurons in normal and pathological brain circuits.
