scholarly journals Chemicogenetic recruitment of specific interneurons suppresses seizure activity

2018 ◽  
Author(s):  
Alexandru Călin ◽  
Mihai Stancu ◽  
Ana-Maria Zagrean ◽  
John G. Jefferys ◽  
Andrei S. Ilie ◽  
...  
Keyword(s):  
Mouse Model ◽  
Seizure Activity ◽  
Epileptiform Activity ◽  
Experimental Models ◽  
Inhibitory Neurons ◽  
Synaptic Inhibition ◽  
Inhibitory Mechanisms ◽  
Hippocampal Seizures

AbstractEnhancing the brain’s endogenous inhibitory mechanisms represents an important strategy for suppressing epileptic discharges. Indeed, drugs that boost synaptic inhibition can disrupt epileptic seizure activity, although these drugs generate complex effects due to the broad nature of their action. Recently developed chemicogenetic techniques provide the opportunity to pharmacologically enhance endogenous inhibitory mechanisms in a more selective manner. Here we use chemicogenetics to assess the anti-epileptic potential of enhancing the synaptic output from three major interneuron populations in the hippocampus: parvalbumin (PV), somatostatin (SST) and vasoactive intestinal peptide (VIP) expressing interneurons. Targeted pre- and post-synaptic whole cell recordings in an in vitro hippocampal mouse model revealed that all three interneuron types increase their firing rate and synaptic output following chemicogenetic activation. However, the interneuron populations exhibited different anti-epileptic effects. Recruiting VIP interneurons resulted in a mixture of pro-epileptic and anti-epileptic effects. In contrast, recruiting SST or PV interneurons produced robust suppression of epileptiform activity. PV interneurons exhibited the strongest effect per cell, eliciting at least a five-fold greater reduction in epileptiform activity than the other cell types. Consistent with this, we found that chemicogenetic recruitment of PV interneurons was effective in an in vivo mouse model of hippocampal seizures. Following efficient delivery of the chemicogenetic tool, pharmacological enhancement of the PV interneuron population suppressed a range of seizure-related behaviours and prevented generalized seizures. Our findings therefore support the idea that selective chemicogenetic enhancement of synaptic inhibitory pathways offers potential as an anti-epileptic strategy.Significance statementDrugs that enhance synaptic inhibition can be effective anticonvulsants but often cause complex effects due to their widespread action. Here we examined the anti-epileptic potential of recently developed chemicogenetic techniques, which offer a way to selectively enhance the synaptic output of distinct types of inhibitory neurons. A combination of in vitro and in vivo experimental models were used to investigate seizure activity in the mouse hippocampus. We find that chemicogenetically recruiting the parvalbumin-expressing population of inhibitory neurons produces the strongest anti-epileptic effect per cell, and that recruiting this cell population can suppress a range of epileptic behaviours in vivo. The data therefore support the idea that targeted chemicogenetic enhancement of synaptic inhibition offers promise for developing new treatments.

scholarly journals Epileptiform Discharges With In-Vivo-Like Features in Slices of Rat Piriform Cortex With Longitudinal Association Fibers

2001 ◽  
Vol 86 (5) ◽  
pp. 2445-2460 ◽  
Author(s):  
Rezan Demir ◽  
Lewis B. Haberly ◽  
Meyer B. Jackson
Keyword(s):  
Seizure Activity ◽  
Epileptiform Activity ◽  
Piriform Cortex ◽  
Brain Slices ◽  
Epileptiform Discharges ◽  
Local Circuitry ◽  
Association Fibers ◽  
Discharge Propagation

Brain slices serve as useful models for the investigation of epilepsy. However, the preparation of brain slices disrupts circuitry and severs axons, thus complicating efforts to relate epileptiform activity in vitro to seizure activity in vivo. This issue is relevant to studies in transverse slices of the piriform cortex (PC), the preparation of which disrupts extensive rostrocaudal fiber systems. In these slices, epileptiform discharges propagate slowly and in a wavelike manner, whereas such discharges in vivo propagate more rapidly and jump abruptly between layers. The objective of the present study was to identify fiber systems responsible for these differences. PC slices were prepared by cutting along three different nearly orthogonal planes (transverse, parasagittal, and longitudinal), and epileptiform discharges were imaged with a voltage-sensitive fluorescent dye. Interictal-like epileptiform activity was enabled by either a kindling-like induction process or disinhibition with bicuculline. The pattern of discharge onset was very similar in slices cut in different planes. As described previously in transverse PC slices, discharges were initiated in the endopiriform nucleus (En) and adjoining regions in a two-stage process, starting with low-amplitude “plateau activity” at one site and leading to an accelerating depolarization and discharge onset at another nearby site. The similar pattern of onset in slices of various orientations indicates that the local circuitry and neuronal properties in and around the En, rather than long-range fibers, assume dominant roles in the initiation of epileptiform activity. Subtle variations in the onset site indicate that interneurons can fine tune the site of discharge onset. In contrast to the mode of onset, discharge propagation showed striking variations. In longitudinal slices, where rostrocaudal association fibers are best preserved, discharge propagation resembled in vivo seizure activity in the following respects: propagation was as rapid as in vivo and about two to three times faster than in other slices; discharges jumped abruptly between the En and PC; and discharges had large amplitudes in superficial layers of the PC. Cuts in longitudinal slices that partially separated the PC from the En eliminated these unique features. These results help clarify why epileptiform activity differs between in vitro and in vivo experiments and suggest that rostrocaudal pyramidal cell association fibers play a major role in the propagation of discharges in the intact brain. The longitudinal PC slice, which best preserves these fibers, is ideally suited for the study their role.

scholarly journals Astrocytic Connexin43 Channels as Candidate Targets in Epilepsy Treatment

Biomolecules ◽  
2020 ◽  
Vol 10 (11) ◽  
pp. 1578 ◽  
Author(s):  
Laura Walrave ◽  
Mathieu Vinken ◽  
Luc Leybaert ◽  
Ilse Smolders
Keyword(s):  
Transmembrane Protein ◽  
Epileptiform Activity ◽  
Brain Slices ◽  
High Energy ◽  
Experimental Models ◽  
Functional Coupling ◽  
Seizure Outcome ◽  
Mrna Levels

In epilepsy research, emphasis is put on exploring non-neuronal targets such as astrocytic proteins, since many patients remain pharmacoresistant to current treatments, which almost all target neuronal mechanisms. This paper reviews available data on astrocytic connexin43 (Cx43) signaling in seizures and epilepsy. Cx43 is a widely expressed transmembrane protein and the constituent of gap junctions (GJs) and hemichannels (HCs), allowing intercellular and extracellular communication, respectively. A plethora of research papers show altered Cx43 mRNA levels, protein expression, phosphorylation state, distribution and/or functional coupling in human epileptic tissue and experimental models. Human Cx43 mutations are linked to seizures as well, as 30% of patients with oculodentodigital dysplasia (ODDD), a rare genetic condition caused by mutations in the GJA1 gene coding for Cx43 protein, exhibit neurological symptoms including seizures. Cx30/Cx43 double knock-out mice show increased susceptibility to evoked epileptiform events in brain slices due to impaired GJ-mediated redistribution of K+ and glutamate and display a higher frequency of spontaneous generalized chronic seizures in an epilepsy model. Contradictory, Cx30/Cx43 GJs can traffic nutrients to high-energy demanding neurons and initiate astrocytic Ca2+ waves and hyper synchronization, thereby supporting proconvulsant effects. The general connexin channel blocker carbenoxolone and blockers from the fenamate family diminish epileptiform activity in vitro and improve seizure outcome in vivo. In addition, interventions with more selective peptide inhibitors of HCs display anticonvulsant actions. To conclude, further studies aiming to disentangle distinct roles of HCs and GJs are necessary and tools specifically targeting Cx43 HCs may facilitate the search for novel epilepsy treatments.

Extracellular pH Responses in CA1 and the Dentate Gyrus During Electrical Stimulation, Seizure Discharges, and Spreading Depression

2000 ◽  
Vol 83 (6) ◽  
pp. 3519-3524 ◽  
Author(s):  
Zhi-Qi Xiong ◽  
Janet L. Stringer
Keyword(s):  
Dentate Gyrus ◽  
Spreading Depression ◽  
Seizure Activity ◽  
Epileptiform Activity ◽  
Extracellular Ph ◽  
Extracellular Acidification ◽  
Ca1 Region ◽  
Activity Dependent

Since neuronal excitability is sensitive to changes in extracellular pH and there is regional diversity in the changes in extracellular pH during neuronal activity, we examined the activity-dependent extracellular pH changes in the CA1 region and the dentate gyrus. In vivo, in the CA1 region, recurrent epileptiform activity induced by stimulus trains, bicuculline, and kainic acid resulted in biphasic pH shifts, consisting of an initial extracellular alkalinization followed by a slower acidification. In vitro, stimulus trains also evoked biphasic pH shifts in the CA1 region. However, in CA1, seizure activity in vitro induced in the absence of synaptic transmission, by perfusing with 0 Ca2+/5 mM K+medium, was only associated with extracellular acidification. In the dentate gyrus in vivo, seizure activity induced by stimulation to the angular bundle or by injection of either bicuculline or kainic acid was only associated with extracellular acidification. In vitro, stimulus trains evoked only acidification. In the dentate gyrus in vitro, recurrent epileptiform activity induced in the absence of synaptic transmission by perfusion with 0 Ca2+/8 mM K+medium was associated with extracellular acidification. To test whether glial cell depolarization plays a role in the regulation of the extracellular pH, slices were perfused with 1 mM barium. Barium increased the amplitude of the initial alkalinization in CA1 and caused the appearance of alkalinization in the dentate gyrus. In both CA1 and the dentate gyrus in vitro, spreading depression was associated with biphasic pH shifts. These results demonstrate that activity-dependent extracellular pH shifts differ between CA1 and dentate gyrus both in vivo and in vitro. The differences in pH fluctuations with neuronal activity might be a marker for the basis of the regional differences in seizure susceptibility between CA1 and the dentate gyrus.

scholarly journals Mouse model of ischemic acute kidney injury: technical notes and tricks

2012 ◽  
Vol 303 (11) ◽  
pp. F1487-F1494 ◽  
Author(s):  
Qingqing Wei ◽  
Zheng Dong
Keyword(s):  
Acute Kidney Injury ◽  
Mouse Model ◽  
Drug Testing ◽  
Ischemia Reperfusion ◽  
Small Animal ◽  
Kidney Injury ◽  
Experimental Models ◽  
Renal Ischemia

Renal ischemia-reperfusion leads to acute kidney injury (AKI), a major kidney disease associated with an increasing prevalence and high mortality rates. A variety of experimental models, both in vitro and in vivo, have been used to study the pathogenic mechanisms of ischemic AKI and to test renoprotective strategies. Among them, the mouse model of renal clamping is popular, mainly due to the availability of transgenic models and the relatively small animal size for drug testing. However, the mouse model is generally less stable, resulting in notable variations in results. Here, we describe a detailed protocol of the mouse model of bilateral renal ischemia-reperfusion. We share the lessons and experiences gained from our laboratory in the past decade. We further discuss the technical issues that account for the variability of this model and offer relevant solutions, which may help other investigators to establish a well-controlled, reliable animal model of ischemic AKI.

Role of Potassium and Calcium in the Generation of Cellular Bursts in the Dentate Gyrus

1997 ◽  
Vol 77 (5) ◽  
pp. 2293-2299 ◽  
Author(s):  
Enhui Pan ◽  
Janet L. Stringer
Keyword(s):  
Dentate Gyrus ◽  
Granule Cells ◽  
Seizure Activity ◽  
Epileptiform Activity ◽  
Hippocampal Slices ◽  
Extracellular Potassium ◽  
Sprague Dawley

Pan, Enhui and Janet L. Stringer. Role of potassium and calcium in the generation of cellular bursts in the dentate gyrus. J. Neurophysiol. 77: 2293–2299, 1997. Epileptiform activity, which appears to be endogenous, has been recorded in the granule cells of the dentate gyrus before the onset of synchronized seizure activity and has been termed cellular bursts. It has been postulated that an increase in input to the dentate gyrus causes a local increase in extracellular K+ concentration ([K+]o) and a decrease in [Ca2+]o that results in this cellular bursting. The first test of this hypothesis is to determine whether the cellular bursts appear in ionic conditions that occur in vivo before the onset of synchronized epileptic activity. This hypothesis was tested in vitro by varying the ionic concentrations in the perfusing solution and recording changes in the granule cells of the dentate gyrus. Intra- and extracellular recordings were made in the dentate gyri of hippocampal slices prepared from anesthetized adult Sprague-Dawley rats. Increasing the extracellular potassium or decreasing the extracellular calcium of the perfusing solution caused three forms of spontaneous activity to appear: depolarizing potentials, action potentials, and cellular bursts. Increasing potassium or decreasing calcium also caused the granule cells to depolarize and reduced their input resistance. No synchronized extracellular field activity was detected. Simultaneously increasing potassium and decreasing calcium caused cellular bursts to appear at concentrations recorded in vivo before the onset of synchronized reverberatory seizure activity.

Neutralization of a Low Molecular Weight Heparin (LHN-1) and Conventional Heparin by Protamine Sulfate in Rats

1986 ◽  
Vol 56 (03) ◽  
pp. 318-322 ◽  
Author(s):  
V Diness ◽  
P B Østergaard
Keyword(s):  
Molecular Weight ◽  
Low Molecular Weight Heparin ◽  
Thrombus Formation ◽  
Experimental Models ◽  
Protamine Sulfate ◽  
Low Molecular Weight ◽  
Antithrombotic Effect ◽  
Higher Doses

SummaryThe neutralization of a low molecular weight heparin (LHN-1) and conventional heparin (CH) by protamine sulfate has been studied in vitro and in vivo. In vitro, the APTT activity of CH was completely neutralized in parallel with the anti-Xa activity. The APTT activity of LHN-1 was almost completely neutralized in a way similar to the APTT activity of CH, whereas the anti-Xa activity of LHN-1 was only partially neutralized.In vivo, CH 3 mg/kg and LHN-1 7.2 mg/kg was given intravenously in rats. The APTT and anti-Xa activities, after neutralization by protamine sulfate in vivo, were similar to the results in vitro. In CH treated rats no haemorrhagic effect in the rat tail bleeding test and no antithrombotic effect in the rat stasis model was found at a protamine sulfate to heparin ratio of about 1, which neutralized APTT and anti-Xa activities. In LHN-1 treated rats the haemorrhagic effect was neutralized when APTT was close to normal whereas higher doses of protamine sulfate were required for neutralization of the antithrombotic effect. This probably reflects the fact that in most experimental models higher doses of heparin are needed to induce bleeding than to prevent thrombus formation. Our results demonstrate that even if complete neutralization of APTT and anti-Xa activities were not seen in LHN-1 treated rats, the in vivo effects of LHN-1 could be neutralized as efficiently as those of conventional heparin. The large fall in blood pressure caused by high doses of protamine sulfate alone was prevented by the prior injection of LHN-1.

Antimicrobial peptides for the treatment of pulmonary tuberculosis, allies or foes?

2018 ◽  
Vol 24 (10) ◽  
pp. 1138-1147
Author(s):  
Bruno Rivas-Santiago ◽  
Flor Torres-Juarez
Keyword(s):  
Pulmonary Tuberculosis ◽  
Antimicrobial Peptides ◽  
Experimental Models ◽  
New Drugs ◽  
World Health ◽  
Public Health Issue ◽  
Health Organization ◽  
Drug Resistant Strains

Tuberculosis is an ancient disease that has become a serious public health issue in recent years, although increasing incidence has been controlled, deaths caused by Mycobacterium tuberculosis have been accentuated due to the emerging of multi-drug resistant strains and the comorbidity with diabetes mellitus and HIV. This situation is threatening the goals of World Health Organization (WHO) to eradicate tuberculosis in 2035. WHO has called for the creation of new drugs as an alternative for the treatment of pulmonary tuberculosis, among the plausible molecules that can be used are the Antimicrobial Peptides (AMPs). These peptides have demonstrated remarkable efficacy to kill mycobacteria in vitro and in vivo in experimental models, nevertheless, these peptides not only have antimicrobial activity but also have a wide variety of functions such as angiogenesis, wound healing, immunomodulation and other well-described roles into the human physiology. Therapeutic strategies for tuberculosis using AMPs must be well thought prior to their clinical use; evaluating comorbidities, family history and risk factors to other diseases, since the wide function of AMPs, they could lead to collateral undesirable effects.

Epithelial Organotypic Cultures: A Viable Model to Address Mechanisms of Carcinogenesis by Epitheliotropic Viruses

2018 ◽  
Vol 18 (4) ◽  
pp. 246-255 ◽  
Author(s):  
Lara Termini ◽  
Enrique Boccardo
Keyword(s):  
Three Dimensional ◽  
Cell Types ◽  
Experimental Models ◽  
Biological Research ◽  
Specific Gene ◽  
Specific Cell ◽  
Organotypic Cultures ◽  
Skin Physiology

In vitro culture of primary or established cell lines is one of the leading techniques in many areas of basic biological research. The use of pure or highly enriched cultures of specific cell types obtained from different tissues and genetics backgrounds has greatly contributed to our current understanding of normal and pathological cellular processes. Cells in culture are easily propagated generating an almost endless source of material for experimentation. Besides, they can be manipulated to achieve gene silencing, gene overexpression and genome editing turning possible the dissection of specific gene functions and signaling pathways. However, monolayer and suspension cultures of cells do not reproduce the cell type diversity, cell-cell contacts, cell-matrix interactions and differentiation pathways typical of the three-dimensional environment of tissues and organs from where they were originated. Therefore, different experimental animal models have been developed and applied to address these and other complex issues in vivo. However, these systems are costly and time consuming. Most importantly the use of animals in scientific research poses moral and ethical concerns facing a steadily increasing opposition from different sectors of the society. Therefore, there is an urgent need for the development of alternative in vitro experimental models that accurately reproduce the events observed in vivo to reduce the use of animals. Organotypic cultures combine the flexibility of traditional culture systems with the possibility of culturing different cell types in a 3D environment that reproduces both the structure and the physiology of the parental organ. Here we present a summarized description of the use of epithelial organotypic for the study of skin physiology, human papillomavirus biology and associated tumorigenesis.

scholarly journals A novel phosphoramide compound, DCZ0805, shows potent anti-myeloma activity via the NF-κB pathway

2021 ◽  
Vol 21 (1) ◽  
Author(s):  
Xuejie Gao ◽  
Bo Li ◽  
Anqi Ye ◽  
Houcai Wang ◽  
Yongsheng Xie ◽  
...  
Keyword(s):  
Mouse Model ◽  
Tumor Burden ◽  
High Rate ◽  
Cell Counting ◽  
Tumor Xenograft ◽  
Luciferase Reporter ◽  
Therapeutic Effects ◽  
Xenograft Mouse Model

Abstract Background Multiple myeloma (MM) is a highly aggressive and incurable clonal plasma cell disease with a high rate of recurrence. Thus, the development of new therapies is urgently needed. DCZ0805, a novel compound synthesized from osalmide and pterostilbene, has few observed side effects. In the current study, we intend to investigate the therapeutic effects of DCZ0805 in MM cells and elucidate the molecular mechanism underlying its anti-myeloma activity. Methods We used the Cell Counting Kit-8 assay, immunofluorescence staining, cell cycle assessment, apoptosis assay, western blot analysis, dual-luciferase reporter assay and a tumor xenograft mouse model to investigate the effect of DCZ0805 treatment both in vivo and in vitro. Results The results showed that DCZ0805 treatment arrested the cell at the G0/G1 phase and suppressed MM cells survival by inducing apoptosis via extrinsic and intrinsic pathways. DCZ0805 suppressed the NF-κB signaling pathway activation, which may have contributed to the inhibition of cell proliferation. DCZ0805 treatment remarkably reduced the tumor burden in the immunocompromised xenograft mouse model, with no obvious toxicity observed. Conclusion The findings of this study indicate that DCZ0805 can serve as a novel therapeutic agent for the treatment of MM.

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