The patch clamp recording technique in vivois a blind patch clamp recording methods to record the current of the spinal or cereral neurons of anaes：hesia （ or awake） animals. This technique can be used to study the...The patch clamp recording technique in vivois a blind patch clamp recording methods to record the current of the spinal or cereral neurons of anaes：hesia （ or awake） animals. This technique can be used to study the synaptic function and plasticity in central nervous system in vivoin order to understand the physiological properties of the ion channels from an integrated point of view. The advantage of this technique have already presented itself in the study of the synaptic transmission and nervous network. Nowadays, in vivo patch whole-cell recording technique in combination with other techniques is becoming a common method in the research fields.展开更多
Radiation therapy is a standard treatment for head and neck tumors.However,patients often exhibit cognitive impairments following radiation therapy.Previous studies have revealed that hippocampal dysfunction,specifica...Radiation therapy is a standard treatment for head and neck tumors.However,patients often exhibit cognitive impairments following radiation therapy.Previous studies have revealed that hippocampal dysfunction,specifically abnormal hippocampal neurogenesis or neuroinflammation,plays a key role in radiation-induced cognitive impairment.However,the long-term effects of radiation with respect to the electrophysiological adaptation of hippocampal neurons remain poorly characterized.We found that mice exhibited cognitive impairment 3 months after undergoing 10 minutes of cranial irradiation at a dose rate of 3 Gy/min.Furthermore,we observed a remarkable reduction in spike firing and excitatory synaptic input,as well as greatly enhanced inhibitory inputs,in hippocampal CA1 pyramidal neurons.Corresponding to the electrophysiological adaptation,we found reduced expression of synaptic plasticity marker VGLUT1 and increased expression of VGAT.Furthermore,in irradiated mice,long-term potentiation in the hippocampus was weakened and GluR1 expression was inhibited.These findings suggest that radiation can impair intrinsic excitability and synaptic plasticity in hippocampal CA1 pyramidal neurons.展开更多
BACKGROUND： Induced differentiation strategies and cytochemical properties of human embryonic stem ceils （hESCs） have been investigated. However, the electrophysiological functions of tyrosine hydroxylase （TH）-po...BACKGROUND： Induced differentiation strategies and cytochemical properties of human embryonic stem ceils （hESCs） have been investigated. However, the electrophysiological functions of tyrosine hydroxylase （TH）-positive cells dedved from hESCs remain unclear. OBJECTIVE： To investigate the differentiation efficiency of TH-positive cells from hESCs in vitro using modified four-step culture methods, including embryoid body formation, and to examine the functional characteristics of the differentiated TH-positive cells using electrophysiological techniques. DESIGN, TIME AND SETTING： Neuroelectrophysiology was performed at the Reproductive Medicine Center and Stem Cell Research Center, Peking University Third Hospital, and the Neuroscience Research Institute and Department of Neurobiology, Peking University, from September 2004 to August 2008. MATERIALS： The hESC line, PKU-1.1, a monoclonal cell line derived from a pre-implantation human blastocyst in the Reproductive Medical Center of Peking University Third Hospital. The patch clamp recording system was provided by the Neuroscience Research Institute and Department of Neurobiology, Peking University. METHODS： The hESC line was induced to differentiate into TH-positive cells in vitro using a modified four-step culture method, including the formation of embryoid body, as well as the presence of sonic hedgehog and fibroblast growth factor 8. The cell karyotype was assessed by G-banding karyotype analysis techniques and specific markers were detected immunocytochemically. Whole-cell configuration was obtained after obtaining a tight seal of over 1 GΩ. Ionic currents were detected by holding the cells at -70 mV and stepping to test voltages between -80 and 40 mV in 10-mV increments in voltage-clamp configuration. MAIN OUTCOME MEASURES： We measured the cell karyotype, specific cell markers, and the electrophysiological properties of the voltage-gated ion channels on the cell membrane of TH-positive dopaminergic cells differentiated from our hESCs line in vitro. RESULTS： The differentiated cells had a consistent appearance, and the majority of cells （〉 90%） expressed TH and β-tubulion, as well as the neural progenitor marker, nestino Cell karyotype analysis demonstrated that all of the hESCs had a stable and normal karyotype （46, XX） after differentiation. In addition, patch clamp recording showed that the 10 recorded TH-positive cells exhibited a fast inward current when the test voltage depolarized to -30 mV, and a delayed outward current when the test voltage depolarized to -10 mV. The peak of inward current was obtained at voltage between 10 mV and 0 mV, while the peak of outward current was obtained at 40 mV. The average peak of inward current density was （ -50.05 ± 15.50） pA/pF, and the average peak of outward current density was （41.98 ± 13.55） pA/pE CONCLUSION： More than 90% of the differentiated hESC-derived cells induced by the modified four-step culture method exhibit dopaminergic neuronal properties, including general electrophysiological functional properties, such as functional potassium and sodium channels.展开更多
文摘The patch clamp recording technique in vivois a blind patch clamp recording methods to record the current of the spinal or cereral neurons of anaes：hesia （ or awake） animals. This technique can be used to study the synaptic function and plasticity in central nervous system in vivoin order to understand the physiological properties of the ion channels from an integrated point of view. The advantage of this technique have already presented itself in the study of the synaptic transmission and nervous network. Nowadays, in vivo patch whole-cell recording technique in combination with other techniques is becoming a common method in the research fields.
基金supported by the National Natural Science Foundation of China,Nos.81925031(to YT),81820108026(to YT),81972967(to WJL),81872549(to YL)the Youth Program of National Natural Science Foundation of China,No.81801229(to YTX)+3 种基金a grant from Guangdong Science and Technology Department of China,Nos.2020B1212060018(to WJL),2020B1212030004(to WJL)the Natural Science Foundation of Guangdong Province,No.2019A1515011754(to WJL)the Science and Technology Program of Guangzhou of China,No.202007030001(to YT)the Science and Technology Planning Project of Guangzhou of China,No.201704030033(to YL).
文摘Radiation therapy is a standard treatment for head and neck tumors.However,patients often exhibit cognitive impairments following radiation therapy.Previous studies have revealed that hippocampal dysfunction,specifically abnormal hippocampal neurogenesis or neuroinflammation,plays a key role in radiation-induced cognitive impairment.However,the long-term effects of radiation with respect to the electrophysiological adaptation of hippocampal neurons remain poorly characterized.We found that mice exhibited cognitive impairment 3 months after undergoing 10 minutes of cranial irradiation at a dose rate of 3 Gy/min.Furthermore,we observed a remarkable reduction in spike firing and excitatory synaptic input,as well as greatly enhanced inhibitory inputs,in hippocampal CA1 pyramidal neurons.Corresponding to the electrophysiological adaptation,we found reduced expression of synaptic plasticity marker VGLUT1 and increased expression of VGAT.Furthermore,in irradiated mice,long-term potentiation in the hippocampus was weakened and GluR1 expression was inhibited.These findings suggest that radiation can impair intrinsic excitability and synaptic plasticity in hippocampal CA1 pyramidal neurons.
基金the National Natural Science Foundation of China, No. 30672239
文摘BACKGROUND： Induced differentiation strategies and cytochemical properties of human embryonic stem ceils （hESCs） have been investigated. However, the electrophysiological functions of tyrosine hydroxylase （TH）-positive cells dedved from hESCs remain unclear. OBJECTIVE： To investigate the differentiation efficiency of TH-positive cells from hESCs in vitro using modified four-step culture methods, including embryoid body formation, and to examine the functional characteristics of the differentiated TH-positive cells using electrophysiological techniques. DESIGN, TIME AND SETTING： Neuroelectrophysiology was performed at the Reproductive Medicine Center and Stem Cell Research Center, Peking University Third Hospital, and the Neuroscience Research Institute and Department of Neurobiology, Peking University, from September 2004 to August 2008. MATERIALS： The hESC line, PKU-1.1, a monoclonal cell line derived from a pre-implantation human blastocyst in the Reproductive Medical Center of Peking University Third Hospital. The patch clamp recording system was provided by the Neuroscience Research Institute and Department of Neurobiology, Peking University. METHODS： The hESC line was induced to differentiate into TH-positive cells in vitro using a modified four-step culture method, including the formation of embryoid body, as well as the presence of sonic hedgehog and fibroblast growth factor 8. The cell karyotype was assessed by G-banding karyotype analysis techniques and specific markers were detected immunocytochemically. Whole-cell configuration was obtained after obtaining a tight seal of over 1 GΩ. Ionic currents were detected by holding the cells at -70 mV and stepping to test voltages between -80 and 40 mV in 10-mV increments in voltage-clamp configuration. MAIN OUTCOME MEASURES： We measured the cell karyotype, specific cell markers, and the electrophysiological properties of the voltage-gated ion channels on the cell membrane of TH-positive dopaminergic cells differentiated from our hESCs line in vitro. RESULTS： The differentiated cells had a consistent appearance, and the majority of cells （〉 90%） expressed TH and β-tubulion, as well as the neural progenitor marker, nestino Cell karyotype analysis demonstrated that all of the hESCs had a stable and normal karyotype （46, XX） after differentiation. In addition, patch clamp recording showed that the 10 recorded TH-positive cells exhibited a fast inward current when the test voltage depolarized to -30 mV, and a delayed outward current when the test voltage depolarized to -10 mV. The peak of inward current was obtained at voltage between 10 mV and 0 mV, while the peak of outward current was obtained at 40 mV. The average peak of inward current density was （ -50.05 ± 15.50） pA/pF, and the average peak of outward current density was （41.98 ± 13.55） pA/pE CONCLUSION： More than 90% of the differentiated hESC-derived cells induced by the modified four-step culture method exhibit dopaminergic neuronal properties, including general electrophysiological functional properties, such as functional potassium and sodium channels.
