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Substances addiction is one of the important factors that deeply affect human health.At present, there is still lack of effective treatment drugs in the clinic.Exploring mechanisms of substances addiction, finding new therapeutic targets and developing effective therapeutic drugs are important issues to be solved.Hyperpolarization-activated cyclic nucleotide gated cation channels (HCN channels) participate in many advanced brain activities and are closely related to the occurrence and progression of various brain diseases.Among them, the researches on the role and mechanism of HCN channels in substances addiction are gradually gaining attention.Reviewing the researches regarding substances addiction, abnormal function and abnormal expression of HCN channels were observed in many brain regions under the condition of psychoactive substances addiction.However, it has not yet been able to fully understand the mechanism and the behavioral consequences of this change.Therefore, we review the neurobiological mechanisms of HCN channels in substances addiction induced by opioids, cocaine, cannabis, amphetamines, alcohol and tobacco, in order to provide new ideas for the mechanism researches and treatment of substance addiction.
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Hyperpolarization activated cyclic nucleotide-gated cation (HCN) channels are involved in neuronal rhythm regulation, synaptic activity, membrane resistance and dendritic integration due to their voltage-gated physiological properties. Recent studies have shown that HCN channels play an important role in central nervous system diseases, such as temporal lobe epilepsy, neuropathic pain, learning and memory disorder, substance abuse addiction and other related diseases. In this paper, we summarize the research progress of HCN channels in central nervous system diseases in recent years.
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Objective To explore the effect of dronedaronel on hyperpolarization-activated cyclic-nucleotide-gated(HCN) channel expression by detecting the change of HCN channel mRNA and protein level before and after giving dronedarone in neonatal rat ventricular myocytes.Methods Neonatal rat ventricular myocytes were separated and digested by type Ⅱ collagenase,and then single ventricular myocytes were collected through differential sticking wall separation method.According to the concentrations (0.1,0.5,1.0,5.0,10.0,20.0 μmol/L of dronedaronel for treating myocytes for 48 h) and time(10 μmol/L of dronedaronel for treating myocytes for 1,6,12,24,48 h)the gradient grouping was conducted.The levels of HCN2 and HCN4 channel mRNA and protein level were determined by real-time PCR and Western blot.Results The HCN2 mRNA and HCN4 mRNA expression levels in concentration gradient group and time gradient group were lower than those in the control group(P＜0.05);compared with the control group,the protein level in the 10 umol/L dronedaronel treatment for 12 h group was significantly down-regulated(P＜ 0.01).Conclusion Dronedaronel could inhibit the expression of HCN2/HCN4 channel mRNA and protein,moreover its action shows the concentration dependency and reaches the maximum at 12 h after medication.
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In this study, we determined mode of action of a novel carbamoyloxy arylalkanoyl arylpiperazine compound (SKL-NP) on hyperpolarization-activated cyclic nucleotide-gated (HCN) channel currents (Ih) that plays important roles in neuropathic pain. In small or medium-sized dorsal root ganglion (DRG) neurons (<40 microm in diameter) exhibiting tonic firing and prominent Ih, SKL-NP inhibited Ih and spike firings in a concentration dependent manner (IC50=7.85 microM). SKL-NP-induced inhibition of Ih was blocked by pretreatment of pertussis toxin (PTX) and N-ethylmaleimide (NEM) as well as 8-Br-cAMP, a membrane permeable cAMP analogue. These results suggest that SKL-NP modulates Ih in indirect manner by the activation of a Gi-protein coupled receptor that decreases intracellular cAMP concentration. Taken together, SKL-NP has the inhibitory effect on HCN channel currents (I h) in DRG neurons of rats.
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Animals , Rats , Diagnosis-Related Groups , Ethylmaleimide , Fires , Ganglia, Spinal , Membranes , Neuralgia , Neurons , Pertussis Toxin , Spinal Nerve RootsABSTRACT
Objective: To transplant the autologous mesenchymal stem cells (MSCs) carrying human hyperpolarization activated cyclic nucleoside gated channel 2 (hHCN2) gene into the His-bundle in porcine model of complete heart block (CHB), so as to assess the possibility of establishing autologous biological pacing. Methods: We constructed the recombinant adenovirus containing hHCN2 gene to transfect the porcine MSCs. After autotransplantation into the Hie-bundle in CHB model, the genetically-altered MSCs were tested for their ability to provide a stable and catecholamine-responsive heart rhythm. Histological and immunofluorescence analyses were also performed for the myocardium of the injection site. Results: Recombinant adenovirus pAd. hHCN2 was successfully constructed. Porcine MSCs were transfected by the adenovirus. After autotransplantation, transgenic MSCs significantly enhanced the heart rates in porcine CHB model compared with the control group (P < 0.01), and the cardiac rhythms in the transgenic MSC group were catecholamine responsive. Tissues obtained from the transplanted heart sites showed that the MSCs survived in the myocardium and overexpressed hHCN2 protein. Conclusion: Transplantation of autologous genetically-altered MSCs into the His-bundle in porcine CHB model can serve as a short-term catecholamine-responsive biological pacemaker.
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The possibility of rats mesenchymal stem cells (MSCs) modified with murine hyperpolarization-activated cyclic nucleotide-gated 2 (mHCN2) gene as biological pacemakers in vitro was studied.The cultured MSCs were transfected with pIRES2-EGFP plasmid carrying enhanced green fluorescent protein (EGFP) gene and mHCN2 gene.The identification using restriction enzyme and sequencing indicated that the mHCN2 gene was inserted to the pIRES2-EGFP.Green fluorescence could be seen in MSCs after transfection for 24-48 h.The expression of mHCN2 mRNA and protein in the transfected cells was detected by RT-PCR and Western blot,and the quantity of mHCN2 mRNA and protein expression in transfected MSCs was 5.31 times and 7.55 times higher than that of the non-transfected MSCs respectively (P＜0.05,P＜0.05).IHCN2 was recorded by whole-cell patch clamp method.The effect of Cs+,a specific blocker of pacemaker current,was measured after perfusion by patch clamp.The results of inward current indicated that there was no inward current recording in non-transfected MSCs and a large voltage-dependent inward and Cs+-sensitive current activated on hyperpolarizations presented in the transfected MSCs.IHCN2 was fully activated around -140 mV with an activation threshold of-60 mV.The midpoint (V50) was -95.1±0.9 mV (n=9).The study demonstrates that mHCN2 mRNA and protein can be expressed and the currents of HCN2 channels can be detected in genetically modified MSCs.The gene-modified MSCs present a novel method for pacemaker genes into the heart or other electrical syncytia.
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Aim To study the effect of ZD7288 on synaptic transmission in the pathway from perforant pathway (PP) fibers to CA3 region in rat hippocampus. Methods The extracellular recording technique in vivo was used to record the CA3 region field potentials. High-performance liquid chromatography (HPLC) with fluorescence detection was applied to measure the content of amino acids in hippocampal tissues. The effect of ZD7288 and CsCl on the amplitudes of population spike (PS) in CA3 region evoked by stimulation (0.5 Hz) of the perforant pathway (PP) fibers, and the content of amino acids in hippocampal tissue were observed. Results Microinjection of ZD7288 (20, 100 and 200 nmol)and CsCl (1,5 and 10 μmol) into CA3 region decreased the population spike (PS) amplitudes in a dosedependent manner. The inhibitory effects appeared at 5 min after microinjection and lasted at least 90 min.In those rats treated with ZD7288 (100 nmol), the contents of glutamate, aspartate, glycine and GABA decreased significantly as compared to those of saline control ( all P＜0.01, except P＜0.05 for that of glycine). A similar decrease in the contents of amino acids was observed when the rats were microinjected with CsCl (5 μmol). Conclusion ZD7288 could obviously inhibit synaptic transmission in the pathway from PP fibers to CA3 region in rat hippocampus, and this action of ZD7288 may be associated with altered contents of amino acids.
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0.05),but there were statistically difference within E15,E19,P2,P10(Pa
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Objective To find the way of inducing the bone marrow mesynchymal stem cells(MSCs)into cardiac cells with pacemaking function in vitro.Methods Dissociate the rat MSCs and induce them with 5AZA,bFGF+EGF,HGF,SCF and lysate of the sinoatrial cells respectively.The morphological changes were observed,and the expressing of protein cTnT,connexin 43 and HCN2/4 were analyse by immunohistologic and flowcytometry techniques.The pacmaking current If were evaluted by patch clamp techniques.Results All the methods can induce the bone marrow MSCs to differentiate into cardiac cells,which expressing cardiac cell specific protein and HCN2.Cells induced by 5AZA,bFGF+EGF and SAN CMs show higher rate of HCN2 expressing(22.9%,22.3%,11%).The cells of these groups have the pacmaking current If.Conclusion Lysate of the sinoatrial cells are ideal methods of inducing the bone marrow MSCs to differentiate into cardiac cells with pacemaking function in vitro.HCN is a promising marker protein to select pacmaking cells out of the differentiated cells.
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Objective To investigate the modulation of electrophysiological properties of hyperpolarization-activated cyclic nucleotide-gated channels(HCN) HCN1 and HCN2 by transfecting MinK-related peptide 1(MiRP1) in Chinese hamster ovarian(CHO) cells.Methods CHO cells were co-transfected with plasmid DNA encoding either of the cardiac HCN isoforms(HCN1,HCN2) with MiRP1 to observe the effect of MiRP1 on HCN whole-cell currents.Whole-cell patch clamp technique was used to record the 2 channel currents of the transfected cells.Results MiRP1 significantly increased whole-cell current density of HCN2 [(37.8?4.8) pA/pF(n=11) vs control(25.9?1.7) pA/pF(n=10);at-140 mV,P0.05].Moreover,MiRP1 accelerated the activation kinetic of HCN1 [tau(180.9?8.6) ms vs control(306.1?45.6) ms;at-80 mV,P