Integrating adult neurogenesis and human brain organoid models to advance epilepsy and associated behavioral research.

Epilepsy is a chronic neurological disorder characterized by recurring, unprovoked seizures, asymmetrical electroencephalogram patterns, and other pathological abnormalities. The hippocampus plays a pivotal role in learning, memory consolidation, attentional control, and pattern separation. Impairment of hippocampal network circuitry can induce long-term cognitive and memory dysfunction. In this review, we discuss how aberrant adult neurogenesis and plasticity collectively alter the network balance for information processing within the hippocampal neural network. Subsequently, we explore the potential of human brain organoids integrated into microelectrode array technology as an electrophysiological tool. We also discuss the utilization of a closed-loop platform that connects the brain organoid to a mobile robot in a virtual environment. While in vivo models provide valuable insights into some aspects of epileptogenesis, such as the impact of adult neurogenesis on hippocampal function, brain organoids are indispensable for comprehensively studying epileptogenesis involving genetic mutations that underlie human epilepsy. More importantly, a combinational approach using brain organoids on MEA paves the way for studying impaired plasticity and abnormal information processing within epileptic neural networks. This innovative in vitro approach may provide a new pathway for investigating the behavioral outcomes of aberrant neural networks when integrated with a mobile robot, closing the loop between the neural network in brain organoids and the mobile robot. In this review, we aim to discuss the use of each model to study the behavioral changes in epilepsy and highlight the benefits of both in vivo and in vitro models for understanding the behavioral aspects of epilepsy.

Dual effects of ARX poly-alanine mutations in human cortical and interneuron development.

Mutations in ARX , an X-linked gene, are implicated in a wide spectrum of neurological disorders including patients who have intellectual disability and epilepsy. Mouse models have shown that Arx is critical for cortical development and interneuron migration, however they do not recapitulate the full phenotype observed in patients. Moreover, the epilepsy in many patients with poly-alanine tract expansion (PAE) mutations in ARX show pharmacoresistance, emphasizing the need to develop new treatments. Here, we used human neural organoid models to study the consequences of PAE mutations, one of the most prevalent mutations in ARX . We found that PAE mutations result in an early increase in radial glia cells and intermediate progenitor cells, and premature differentiation leading to a loss of cortical neurons at later timepoints. Moreover, ARX expression is upregulated in CO derived from patient at 30 DIV which alters the expression of CDKN1C , SFRP1 , DLK1 and FABP7 , among others. We also found a cell autonomously enhanced interneuron migration, which can be rescued by CXCR4 inhibition. Furthermore, ARX PAE assembloids had hyper-activity and synchrony evident from the early stages. These data provide novel insights to the pathogenesis of these and likely related human neurological disorders and identifies a critical window for therapeutic interventions.

Dopamine Receptors Gene Expression Pattern and Locomotor Improvement Differ Between Female and Male Zebrafish During Spinal Cord Auto Repair.

The dopaminergic system, a spinal cord (SC) motor circuit regulator, is administrated by sexual hormones and evolutionary conserved in all vertebrates. Accordingly, we hypothesized that the dopamine receptor (DAR) expression pattern may be dissimilar in female and male zebrafish SC auto repair. We implemented an uncomplicated method to induce spinal cord injury (SCI) on fully reproductive adult zebrafish, in both genders. SCI was induced using a 28-gauge needle at 9th-10th vertebra without skin incision. Thereupon, lesioned SC was harvested for DAR gene expression analysis; zebrafish were tracked routinely for any improvement in swim distance, speed, and their roaming capabilities/preference. Our findings revealed discrepancies between drd2a, drd2b, drd3, drd4a, and drd4b expression patterns at 1, 7, and 14 days postinjury (DPI) between female and male zebrafish. The receptors were mostly upregulated at 7 DPI in both genders, whereas drd2a and drd2b were mostly maximized in females. Surprisingly, drd3 was measured greater even in intact SC in males. In addition, female zebrafish were able to swim farther distances more accelerated, in multiple directions, by engaging more caudal muscles compared with males, of course with no statistical significance. Indeed, females were able to generate whole-body rotation and move forward using the muscles downstream to the lesion site, whereas the coordinated movement in males was accomplished by rostral muscles. In conclusion, there are differences in DAR gene expression pattern throughout SC autonomous recovery between adult female and male zebrafish, and also, female locomotion seems to ameliorate more rapidly.

In vitro characterization of subventricular zone isolated neural stem cells, from adult monkey and rat brain.

Neural stem cells (NSCs) are multipotent, self-renewable cells who are capable of differentiating into neurons, astrocytes, and oligodendrocytes. NSCs reside at the subventricular zone (SVZ) of the adult brain permanently to guarantee a lifelong neurogenesis during neural network plasticity or undesirable injuries. Although the specious inaccessibility of adult NSCs niche hampers their in vivo identification, researchers have been seeking ways to optimize adult NSCs isolation, expansion, and differentiation, in vitro. NSCs were isolated from rhesus monkey SVZ, expanded in vitro and then characterized for NSCs-specific markers expression by immunostaining, real-time PCR, flow cytometry, and cell differentiation assessments. Moreover, cell survival as well as self-renewal capacity were evaluated by TUNEL, Live/Dead and colony assays, respectively. In the next step, to validate SVZ-NSCs identity in other species, a similar protocol was applied to isolate NSCs from adult rat's SVZ as well. Our findings revealed that isolated SVZ-NSCs from both monkey and rat preserve proliferation capacity in at least nine passages as confirmed by Ki67 expression. Additionally, both SVZ-NSCs sources are capable of self-renewal in addition to NESTIN, SOX2, and GFAP expression. The mortality was measured meager with over 95% viability according to TUNEL and Live/Dead assay results. Eventually, the multipotency of SVZ-NSCs appraised authentic after their differentiation into neurons, astrocytes, and oligodendrocytes. In this study, we proposed a reliable method for SVZ-NSCs in vitro maintenance and identification, which, we believe is a promising cell source for therapeutic approach to recover neurological disorders and injuries condition.

Effect of Transcranial Direct Current Stimulation on Dorsolateral Prefrontal Cortex to Reduce the Symptoms of the Obsessive-Compulsive Disorder.

INTRODUCTION: Obsessive-Compulsive Disorder (OCD) is one of the most common debilitating mental disorders with a prevalence rate of 2% to 3% in the general population. Previous studies have indicated abnormalities in the dorsolateral prefrontal cortex (DLPFC) of OCD patients; thus, we decided to use transcranial Direct Current Stimulation (tDCS) to decline these patients' symptoms. METHODS: A total of 24 patients with OCD participated in this study with the hope of improvement after the application of tDCS. The subjects were randomly assigned to three groups of Sham, Right DLPFC, and Left DLPFC. tDCS was applied for five consecutive days and in each session, patients were subjected to 2 mA current flow for two 15 minutes followed by a 10-minute rest in between (every session lasted for 40 minutes). RESULTS: Subsequently, the changes in obsessive-compulsive level and cognitive functions were evaluated via Yale-Brown Obsessive-Compulsive Scale (Y-BOCS) and Depression, Anxiety, and Stress Scale 21 (DASS-21) by comparing the results before (pre-test) and after (post-test) tDCS treatment. CONCLUSION: Ultimately, the scores of the Yale-Brown scale in the Left DLPFC group showed significant changes after treatment with tDCS (mean difference compared to the sham group: -6.18 and P≤0.05). Hereupon, this study demonstrated that transcranial direct current stimulation may cause improvements in symptoms of OCD.

Protocol for purification and culture of astrocytes: useful not only in 2 days postnatal but also in adult rat brain.

Astrocytes play the key roles in the physiology and pathology of the CNS. Thereupon, in this manuscript, we aim to demonstrate that the protocol for purification and culture of astrocytes is useful not only in 2 days postnatal but also in adult rat brain. Also, the mentioned protocol is a simple and efficient primary cell culture technique. The whole-brain was isolated from the skull and the meninges were removed carefully. Afterward, the cerebral hemispheres were mechanically and enzymatically digested. Then, the cell suspension was seeded in T25 culture flask and was incubated at 37 °C in the CO2 incubator. The first shaking was performed after 7-8 days and on day 14, second shaking was done. After 2-3 passage, the culture was analyzed. By passaging, the majority of extracted cells were astrocytes presenting with a polygonal to fusiform and flat morphology that expressed GFAP, GLAST, and S100ß. The expression of neural, neuronal and oligodendrocyte markers was not detected in extracted cells. The patch-clamp recording comfirmed the purity of isolated astrocytes as well. The isolated cells from adult rat brain were astrocytes that expressed specific astrocyte markers after 3 and 10 passages. This method is suggested to obtain a population of astrocytes that may provide a beneficial tool for different neurophysiological and pathophysiological studies.

Combinational therapy of lithium and human neural stem cells in rat spinal cord contusion model.

A large number of treatment approaches have been used for spinal cord injury improvement, a medically incurable disorder, and subsequently stem cell transplantation appears to be a promising strategy. The main objective of this study is to ascertain whether combinational therapy of human neural stem cells (hNSCs) together with lithium chloride improves cell survival, proliferation, and differentiation in a rat spinal contusion model, or not. Contusive spinal cord injury was implemented on Wistar male rats. Experimental groups comprised of: control, hNSCs transplanted, lithium chloride (Li), and hNSCs and lithium chloride (hNSCs + Li). In every experimental group, locomotor activity score and motor evoked potential (MEP) were performed to evaluate motor recovery as well as histological assessments to determine mechanisms of improvement. In accordance with our results, the hNSCs + Li and the Li groups showed significant improvement in locomotor scores and MEP. Also, Histological assessments revealed that transplanted hNSCs are capable of differentiation and migration along the spinal cord. Although NESTIN-positive cells were proliferated significantly in the Lithium group in comparison with control and the hNSCs + Li groups, the quantity of ED1 cells in the hNSCs + Li was significantly larger than the other two groups. Our results demonstrate that combinational therapy of hNSCs with lithium chloride and lithium chloride individually are adequate for ameliorating more than partial functional recovery and endogenous repair in spinal cord-injured rats.

Development of membrane ion channels during neural differentiation from human embryonic stem cells.

OBJECTIVE: For human embryonic stem cells (hESCs) to differentiate into neurons, enormous changes has to occur leading to trigger action potential and neurotransmitter release. We attempt to determine the changes in expression of voltage gated channels (VGCs) and their electrophysiological properties during neural differentiation. MATERIALS AND METHODS: The relative expressions of α-subunit of voltage gated potassium, sodium and calcium channels were characterized by qRT-PCR technique. Patch clamp recording was performed to characterize the electrophysiological properties of hESCs during their differentiation into neuron-like cells. RESULTS: Relative expression of α-subunit of channels changed significantly. 4-AP and TEA sensitive outward currents were observed in all stages, although TEA sensitive currents were recorded once in rosette structure. Nifedipine and QX314 sensitive inward currents were recorded only in neuron-like cells. CONCLUSION: K+ currents were recorded in hESCs and rosette structure cells. Inward currents, sensitive to Nifedipine and QX314, were recorded in neuron-like cells.