Decreased Hyperpolarization-Activated Cyclic Nucleotide-Gated Channel 2 Activity in a Rat Model of Absence Epilepsy and the Effect of ZD7288, an Ih Inhibitor, on the Spike-and-Wave Discharges

Pharmacology ◽  
2022 ◽  
pp. 1-8
Author(s):  
Melis Yavuz ◽  
Banu Aydın ◽  
Nihan Çarçak ◽  
Filiz Onat
Keyword(s):  
Rat Model ◽  
Cyclic Nucleotide ◽  
Total Duration ◽  
Absence Epilepsy ◽  
Enzyme Linked Immunosorbent Assay ◽  
Control Group ◽  
Hcn Channels ◽  
Hcn Channel ◽  
Gated Channel ◽  
Channel Currents

<b><i>Introduction:</i></b> Hyperpolarization-activated cyclic nucleotide-gated (HCN) channel currents of <i>Ih</i> and absence epilepsy seizures are associated, but studies reveal differential results. <b><i>Objective:</i></b> In our study, we aimed to investigate the role of the HCN channels on the expression of spike-and-wave discharges (SWDs) using the Genetic Absence Epilepsy Rats from Strasbourg (GAERS) model. <b><i>Methods:</i></b> HCN isoform levels from isolated brains of both naïve nonepileptic Wistar and GAERS groups were evaluated by enzyme-linked immunosorbent assay. ZD7288, an <i>Ih</i> inhibitor as well as an HCN channel antagonist, was administered intracerebroventricularly to the adult GAERS groups, and to evaluate their SWD activities, electroencephalography was recorded. The effect of ZD7288 on the cumulative total duration and number of SWDs and the mean duration of each SWD complex was evaluated. <b><i>Results:</i></b> The HCN2 levels in the cortex and hippocampus of the GAERS group were lower compared to the naïve nonepileptic Wistar group (<i>p</i> &#x3c; 0.05). ZD7288 increased the number of SWDs at the 20th and 120th min with the highest administered dose of 7 μg (<i>p</i> &#x3c; 0.05). <b><i>Conclusion:</i></b> The <i>Ih</i> inhibitor ZD7288 increased the number of SWDs in a genetic absence epilepsy rat model, although this increase may not be significant due to the inconsistent time-dependent effects. In GAERS, the cortical and hippocampal HCN2 channel levels were significantly lower compared to the control group. Further studies are needed with higher doses of ZD7288 to determine if the effects will increase drastically.

scholarly journals The Role of HCN Channels on Membrane Excitability in the Nervous System

2012 ◽  
Vol 2012 ◽  
pp. 1-11 ◽  
Author(s):  
Daisuke Kase ◽  
Keiji Imoto
Keyword(s):  
Intracellular Signaling ◽  
Hcn Channels ◽  
Heart Cells ◽  
Hcn Channel ◽  
Membrane Excitability ◽  
Wide Range ◽  
Inward Currents ◽  
Range Of Functions ◽  
Channel Currents

Hyperpolarization-activated and cyclic nucleotide-gated (HCN) channels were first reported in heart cells and are recently known to be involved in a variety of neural functions in healthy and diseased brains. HCN channels generate inward currents when the membrane potential is hyperpolarized. Voltage dependence of HCN channels is regulated by intracellular signaling cascades, which contain cyclic AMP, PIP2, and TRIP8b. In addition, voltage-gated potassium channels have a strong influence on HCN channel activity. Because of these funny features, HCN channel currents, previously called funny currents, can have a wide range of functions that are determined by a delicate balance of modulatory factors. These multifaceted features also make it difficult to predict and elucidate the functional role of HCN channels in actual neurons. In this paper, we focus on the impacts of HCN channels on neural activity. The functions of HCN channels reported previously will be summarized, and their mechanisms will be explained by using numerical simulation of simplified model neurons.

scholarly journals Involvement of Hyperpolarization-Activated Cyclic Nucleotide-Gated Channel 3 in Oxytocin Neuronal Activity in Lactating Rats With Pup Deprivation

ASN NEURO ◽  
2020 ◽  
Vol 12 ◽  
pp. 175909142094465 ◽  
Author(s):  
Dongyang Li ◽  
Haitao Liu ◽  
Xiaoyu Liu ◽  
Hongyang Wang ◽  
Tong Li ◽  
...  
Keyword(s):  
Neuronal Activity ◽  
Cyclic Nucleotide ◽  
Brain Slices ◽  
Oxytocin Receptor ◽  
Molecular Association ◽  
Cyclooxygenase 2 ◽  
Enzyme Linked Immunosorbent Assay ◽  
Prostaglandin E ◽  
Western Blots ◽  
Gated Channel

Oxytocin, a hypothalamic neuropeptide essential for breastfeeding, is mainly produced in oxytocin neurons in the supraoptic nucleus (SON) and paraventricular nucleus. However, mechanisms underlying oxytocin secretion, specifically the involvement of hyperpolarization-activated cyclic nucleotide-gated channel 3 (HCN3) in oxytocin neuronal activity, remain unclear. Using a rat model of intermittent and continuous pup deprivation (PD) at the middle stage of lactation, we analyzed the contribution of HCN3 in oxytocin receptor (OTR)-associated signaling cascade to oxytocin neuronal activity in the SON. PD caused maternal depression, anxiety, milk shortage, involution of the mammary glands, and delays in uterine recovery, particularly in continuous PD. PD increased hypothalamic but not plasma oxytocin levels in enzyme-linked immunosorbent assay. In the SON, PD increased c-Fos expression but reduced expressions of cyclooxygenase-2 and HCN3 in Western blots and/or immunohistochemistry. Moreover, PD significantly increased the molecular association of OTR with HCN3 in coimmunoprecipitation. In brain slices, inhibition of HCN3 activity with DK-AH269 blocked prostaglandin E2-evoked increase in the firing activity and burst discharge in oxytocin neurons in patch-clamp recordings. In addition, oxytocin-evoked increase in the molecular association between OTR and HCN3 in brain slices of the SON was blocked by pretreatment with indomethacin, an inhibitor of cyclooxygenase-2. These results indicate that normal activity of oxytocin neurons is under the regulation of an oxytocin receptor–cyclooxygenase-2–HCN3 pathway and that PD disrupts maternal behavior through increasing intranuclear oxytocin secretion in the SON but likely reducing bolus oxytocin release into the blood through inhibition of HCN3 activity.

scholarly journals Preventive Effects of the Intestine Function Recovery Decoction, a Traditional Chinese Medicine, on Postoperative Intra-Abdominal Adhesion Formation in a Rat Model

2016 ◽  
Vol 2016 ◽  
pp. 1-10 ◽  
Author(s):  
Cancan Zhou ◽  
Pengbo Jia ◽  
Zhengdong Jiang ◽  
Ke Chen ◽  
Guanghui Wang ◽  
...  
Keyword(s):  
Chinese Medicine ◽  
Traditional Chinese Medicine ◽  
Rat Model ◽  
Adhesion Formation ◽  
Enzyme Linked Immunosorbent Assay ◽  
Control Group ◽  
High Dose ◽  
Abdominal Adhesion ◽  
Function Recovery ◽  
Model Control

The intestine function recovery decoction (IFRD) is a traditional Chinese medicine that has been used for the treatment of adhesive intestinal obstruction. In this study, the preventative effects and probable mechanism of the IFRD were investigated in a rat model. We randomly assigned rats to five groups: normal, model, control, low dose IFRD, and high dose IFRD. In the animal model, the caecum wall and parietal peritoneum were abraded to induce intra-abdominal adhesion formation. Seven days after surgery, adhesion scores were assessed using a visual scoring system, and histopathological samples were examined. The levels of serum interleukin-6 (IL-6) and transforming growth factor beta-1 (TGF-β1) were analysed by an enzyme-linked immunosorbent assay (ELISA). The results showed that a high dose of IFRD reduced the grade of intra-abdominal adhesion in rats. Furthermore, the grades of inflammation, fibrosis, and neovascularization in the high dose IFRD group were significantly lower than those in the control group. The results indicate that the IFRD can prevent intra-abdominal adhesion formation in a rat model. These data suggest that the IFRD may be an effective antiadhesion agent.

scholarly journals Inner activation gate in S6 contributes to the state-dependent binding of cAMP in full-length HCN2 channel

2012 ◽  
Vol 140 (1) ◽  
pp. 29-39 ◽  
Author(s):  
Shengjun Wu ◽  
Weihua Gao ◽  
Changan Xie ◽  
Xinping Xu ◽  
Christina Vorvis ◽  
...  
Keyword(s):  
Ligand Binding ◽  
Cyclic Nucleotide ◽  
Binding Domain ◽  
Hcn Channels ◽  
Channel Blockers ◽  
Structural Elements ◽  
Hcn Channel ◽  
Channel Interaction ◽  
State Dependent ◽  
Activation Gate

Recently, applications of the patch-clamp fluorometry (PCF) technique in studies of cyclic nucleotide–gated (CNG) and hyperpolarization-activated, cyclic nucleotide–regulated (HCN) channels have provided direct evidence for the long-held notion that ligands preferably bind to and stabilize these channels in an open state. This state-dependent ligand–channel interaction involves contributions from not only the ligand-binding domain but also other discrete structural elements within the channel protein. This insight led us to investigate whether the pore of the HCN channel plays a role in the ligand–whole channel interaction. We used three well-characterized HCN channel blockers to probe the ion-conducting passage. The PCF technique was used to simultaneously monitor channel activity and cAMP binding. Two ionic blockers, Cs+ and Mg2+, effectively block channel conductance but have no obvious effect on cAMP binding. Surprisingly, ZD7288, an open channel blocker specific for HCN channels, significantly reduces the activity-dependent increase in cAMP binding. Independent biochemical assays exclude any nonspecific interaction between ZD7288 and isolated cAMP-binding domain. Because ZD7228 interacts with the inner pore region, where the activation gate is presumably located, we did an alanine scanning of the intracellular end of S6, from T426 to A435. Mutations of three residues, T426, M430, and H434, which are located at regular intervals on the S6 α-helix, enhance cAMP binding. In contrast, mutations of two residues in close proximity, F431A and I432A, dampen the response. Our results demonstrate that movements of the structural elements near the activation gate directly affect ligand binding affinity, which is a simple mechanistic explanation that could be applied to the interpretation of ligand gating in general.

scholarly journals HCN domain is required for HCN channel expression and couples voltage- and cAMP-dependent gating mechanisms

2020 ◽  
Author(s):  
Ze-Jun Wang ◽  
Ismary Blanco ◽  
Sebastien Hayoz ◽  
Tinatin I. Brelidze
Keyword(s):  
Cyclic Nucleotide ◽  
Transmembrane Domain ◽  
Rhythmic Activity ◽  
Surface Expression ◽  
Hcn Channels ◽  
Hcn Channel ◽  
Structural Element ◽  
Functional Link ◽  
Membrane Hyperpolarization ◽  
Membrane Spanning

ABSTRACTHyperpolarization-activated cyclic nucleotide-gated (HCN) channels are major regulators of synaptic plasticity, and rhythmic activity in the heart and brain. Opening of HCN channels requires membrane hyperpolarization and is further facilitated by intracellular cyclic nucleotides (cNMPs). In HCN channels, membrane hyperpolarization is sensed by the membrane-spanning voltage sensor domain (VSD) and the cNMP-dependent gating is mediated by the intracellular cyclic nucleotide-binding domain (CNBD) connected to the pore-forming S6 transmembrane domain via the C-linker. Previous functional analysis of HCN channels suggested a direct or allosteric coupling between the voltage- and cNMP-dependent activation mechanisms. However, the specifics of the coupling were unclear. The first cryo-EM structure of an HCN1 channel revealed that a novel structural element, dubbed HCN domain (HCND), forms a direct structural link between the VSD and C-linker/CNBD. In this study, we investigated the functional significance of the HCND. Deletion of the HCND prevented surface expression of HCN2 channels. Based on the HCN1 structure analysis, we identified R237 and G239 residues on the S2 of the VSD that form direct interactions with I135 on the HCND. Disrupting these interactions abolished HCN2 currents. We then identified three residues on the C-linker/CNBD (E478, Q382 and H559) that form direct interactions with residues R154 and S158 on the HCND. Disrupting these interactions affected both voltage- and cAMP-dependent gating of HCN2 channels. These findings indicate that the HCND is necessary for the surface expression of HCN channels, and provides a functional link between the voltage- and cAMP-dependent mechanisms of HCN channel gating.

MMP-3-1612 polymorphism - a risk factor for deep venous thrombosis formation

VASA ◽  
2016 ◽  
Vol 45 (3) ◽  
pp. 233-239
Author(s):  
La-Mei Yu ◽  
Nai-Xuan Li ◽  
Yu-Guo Sheng
Keyword(s):  
Risk Factor ◽  
Venous Thrombosis ◽  
Deep Venous Thrombosis ◽  
Allele Frequency ◽  
Rat Model ◽  
Enzyme Linked Immunosorbent Assay ◽  
Control Group ◽  
Control Groups ◽  
And Control

Abstract. Background: We investigated the association of the 5A/6A polymorphism in the promoter region at -1612 of the matrix metalloproteinase-3 gene (MMP-3-1612) and deep venous thrombosis (DVT). Patients, materials and methods: The distribution of the MMP-3 (-1612 5A/6A) polymorphism in the case and control groups was detected by polymerase chain reaction-restriction fragment length polymorphism (PCR-RFLP). Serum MMP-3 level of two groups was detected using enzyme-linked immunosorbent assay (ELISA). HepG2 cells containing MMP-3-1612 recombinant plasmid were cultured in vitro and the MMP-3 level was defined by luminescence intensity of luciferase. A DVT rat model was built. Serum MMP-3 level in the rats’ wounded vein at different time points was detected by ELISA and recorded for investigation of the association between MMP-3 and DVT. Statistical data analysis was conducted with SPSS18.0. Results: On the basis of the observation of MMP-3-1612 genotype frequency and allele frequency in the case and control groups, we identified significantly higher MMP-3-1612 5A allele frequency and higher serum MMP-3 level in the case group than in the control group (both P < 0.05). According to in vitro luciferase measurements, the 5A allele had higher transcriptional activity than the 6A allele. As observed in the rat model, serum MMP-3 level increased with time passing and thrombosis formation after modelling. Conclusions: The MMP-3-1612 5A/6A polymorphism may effect serum MMP-3 level and over-expression of serum MMP-3 level may be a risk factor for DVT formation.

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