Synaptic vesicles can undergo several modes of exocytosis,endocytosis,and trafficking within individual synapses,and their fates may be linked to different vesicular protein compositions.Here,we mapped the intrasynapt...Synaptic vesicles can undergo several modes of exocytosis,endocytosis,and trafficking within individual synapses,and their fates may be linked to different vesicular protein compositions.Here,we mapped the intrasynaptic distribution of the synaptic vesicle proteins SV2B and SV2A in glutamatergic synapses of the hippocampus using three-dimensional electron microscopy.SV2B was almost completely absent from docked vesicles and a distinct cluster of vesicles found near the active zone.In contrast,SV2A was found in all domains of the synapse and was slightly enriched near the active zone.SV2B and SV2A were found on the membrane in the peri-active zone,suggesting the recycling from both clusters of vesicles.SV2B knockout mice displayed an increased seizure induction threshold only in a model employing high-frequency stimulation.Our data show that glutamatergic synapses generate molecularly distinct populations of synaptic vesicles and are able to maintain them at steep spatial gradients.The almost complete absence of SV2B from vesicles at the active zone of wildtype mice may explain why SV2A has been found more important for vesicle release.展开更多
Fibroblast growth factor(FGF)7,a member of FGF family,is initially found to be secreted from mesenchymal cells to repair epithelial tissues.However,its functions in the nervous system are largely unknown.The present s...Fibroblast growth factor(FGF)7,a member of FGF family,is initially found to be secreted from mesenchymal cells to repair epithelial tissues.However,its functions in the nervous system are largely unknown.The present study showed that FGF7 was a neuromodulator localized in the large dense-core vesicles(LDCVs)in nociceptive neurons.FGF7 was mainly expressed in small-diameter neurons of the dorsal root ganglion and could be transported to the dorsal spinal cord.Interestingly,FGF7 was mostly stored in LDCVs that did not contain neuropeptide substance P.Electrophysiological recordings in the spinal cord slice showed that buffer-applied FGF7 increased the amplitude of excitatory post-synaptic current evoked by stimulating the sensory afferent fibers.Behavior tests showed that intrathecally applied FGF7 potentiated the formalin-induced acute nociceptive response.Moreover,both acute and inflammatory nociceptive responses were significantly reduced in Fgf7-deficient mice.These results suggest that FGF7 exerts an excitatory modulation of nociceptive afferent transmission.展开更多
Atom transfer radical polymerization of protected 2 hydroxyethyl acrylate (HEA\|TMS) was conducted to prepare well\|defined bromo\|terminated macroinitiators,which were further allowed to react with large excess amoun...Atom transfer radical polymerization of protected 2 hydroxyethyl acrylate (HEA\|TMS) was conducted to prepare well\|defined bromo\|terminated macroinitiators,which were further allowed to react with large excess amount of C 60 under ATRP condition.A well defined hammer shaped C 60 PHEA was then obtained by hydrolysis of C 60 PHEA TMS under mild conditions.A TEM observation of the association behavior of these well defined hammer like C 60 PHEA macromolecules showed that they assemble into spherical vesicles in water.Their characteristic inner water phase was further confirmed by fluorescence probe method.Also,it was found that the size of vesicles decreased as the increase of molecular weight of PHEA.展开更多
Biological systems are the sum of their dynamic three-dimensional(3D)parts.Therefore,it is critical to study biological structures in 3D and at high resolution to gain insights into their physiological functions.Elect...Biological systems are the sum of their dynamic three-dimensional(3D)parts.Therefore,it is critical to study biological structures in 3D and at high resolution to gain insights into their physiological functions.Electron microscopy of metal replicas of unroofed cells and isolated organelles has been a key technique to visualize intracellular structures at nanometer resolution.However,many of these methods require specialized equipment and personnel to complete them.Here,we present novel accessible methods to analyze biological structures in unroofed cells and biochemically isolated organelles in 3D and at nanometer resolution,focusing on Arabidopsis clathrin-coated vesicles(CCVs).While CCVs are essential trafficking organelles,their detailed structural information is lacking due to their poor preservation when observed via classical electron microscopy protocols experiments.First,we establish a method to visualize CCVs in unroofed cells using scanning transmission electron microscopy tomography,providing sufficient resolution to define the clathrin coat arrangements.Critically,the samples are prepared directly on electron microscopy grids,removing the requirement to use extremely corrosive acids,thereby enabling the use of this method in any electron microscopy lab.Secondly,we demonstrate that this standardized sample preparation allows the direct comparison of isolated CCV samples with those visualized in cells.Finally,to facilitate the high-throughput and robust screening of metal replicated samples,we provide a deep learning analysis method to screen the“pseudo 3D”morphologies of CCVs imaged with 2D modalities.Collectively,our work establishes accessible ways to examine the 3D structure of biological samples and provide novel insights into the structure of plant CCVs.展开更多
Self-assemblies of amphiphiles in solutions were investigated by using freeze-fracture transmission electron microscopy(FF-TEM). Especially, vesicles were characterized by FF-TEM and the transition of self-assemblies ...Self-assemblies of amphiphiles in solutions were investigated by using freeze-fracture transmission electron microscopy(FF-TEM). Especially, vesicles were characterized by FF-TEM and the transition of self-assemblies was determined. The stacked lamellar Lα-phase was prepared without shear forces by a chemical reaction. The stacked lamellar Lα-phase can be transformed into multilamellar vesicles by the shearing forces that occur when the stacked lamellar Lα-phase sample is turned upside down a few times. The multilamellar vesicles can also be transformed into unilamellar vesicles by high shearing forces. These transitions were demonstrated by FF-TEM measurements. Zn2+-induced vesicle formation in the single-chain surfactant solutions was first achieved.展开更多
基金supported by grants from Deutsche Forschungsgemeinschaft(DFG)(SFB1089,SCHO 820/4-1,SCHO 820/6-1,SCHO 820/7-1,SCHO 820/5-2,and SPP1757 to S.S.,SFB1089,SPP1757,INST117215,DI853/3-5&7,and INST 217/785-1 to D.D.),the BONFOR program of the University of Bonn Medical Center(S.S.and D.D.),and UCB Pharma.
文摘Synaptic vesicles can undergo several modes of exocytosis,endocytosis,and trafficking within individual synapses,and their fates may be linked to different vesicular protein compositions.Here,we mapped the intrasynaptic distribution of the synaptic vesicle proteins SV2B and SV2A in glutamatergic synapses of the hippocampus using three-dimensional electron microscopy.SV2B was almost completely absent from docked vesicles and a distinct cluster of vesicles found near the active zone.In contrast,SV2A was found in all domains of the synapse and was slightly enriched near the active zone.SV2B and SV2A were found on the membrane in the peri-active zone,suggesting the recycling from both clusters of vesicles.SV2B knockout mice displayed an increased seizure induction threshold only in a model employing high-frequency stimulation.Our data show that glutamatergic synapses generate molecularly distinct populations of synaptic vesicles and are able to maintain them at steep spatial gradients.The almost complete absence of SV2B from vesicles at the active zone of wildtype mice may explain why SV2A has been found more important for vesicle release.
基金This work was supported by the National Natural Science Foundation of China(31130066)the Strategic Priority Research Program(B)of Chinese Academy of Sciences(XDB01020300).
文摘Fibroblast growth factor(FGF)7,a member of FGF family,is initially found to be secreted from mesenchymal cells to repair epithelial tissues.However,its functions in the nervous system are largely unknown.The present study showed that FGF7 was a neuromodulator localized in the large dense-core vesicles(LDCVs)in nociceptive neurons.FGF7 was mainly expressed in small-diameter neurons of the dorsal root ganglion and could be transported to the dorsal spinal cord.Interestingly,FGF7 was mostly stored in LDCVs that did not contain neuropeptide substance P.Electrophysiological recordings in the spinal cord slice showed that buffer-applied FGF7 increased the amplitude of excitatory post-synaptic current evoked by stimulating the sensory afferent fibers.Behavior tests showed that intrathecally applied FGF7 potentiated the formalin-induced acute nociceptive response.Moreover,both acute and inflammatory nociceptive responses were significantly reduced in Fgf7-deficient mice.These results suggest that FGF7 exerts an excitatory modulation of nociceptive afferent transmission.
文摘Atom transfer radical polymerization of protected 2 hydroxyethyl acrylate (HEA\|TMS) was conducted to prepare well\|defined bromo\|terminated macroinitiators,which were further allowed to react with large excess amount of C 60 under ATRP condition.A well defined hammer shaped C 60 PHEA was then obtained by hydrolysis of C 60 PHEA TMS under mild conditions.A TEM observation of the association behavior of these well defined hammer like C 60 PHEA macromolecules showed that they assemble into spherical vesicles in water.Their characteristic inner water phase was further confirmed by fluorescence probe method.Also,it was found that the size of vesicles decreased as the increase of molecular weight of PHEA.
文摘Biological systems are the sum of their dynamic three-dimensional(3D)parts.Therefore,it is critical to study biological structures in 3D and at high resolution to gain insights into their physiological functions.Electron microscopy of metal replicas of unroofed cells and isolated organelles has been a key technique to visualize intracellular structures at nanometer resolution.However,many of these methods require specialized equipment and personnel to complete them.Here,we present novel accessible methods to analyze biological structures in unroofed cells and biochemically isolated organelles in 3D and at nanometer resolution,focusing on Arabidopsis clathrin-coated vesicles(CCVs).While CCVs are essential trafficking organelles,their detailed structural information is lacking due to their poor preservation when observed via classical electron microscopy protocols experiments.First,we establish a method to visualize CCVs in unroofed cells using scanning transmission electron microscopy tomography,providing sufficient resolution to define the clathrin coat arrangements.Critically,the samples are prepared directly on electron microscopy grids,removing the requirement to use extremely corrosive acids,thereby enabling the use of this method in any electron microscopy lab.Secondly,we demonstrate that this standardized sample preparation allows the direct comparison of isolated CCV samples with those visualized in cells.Finally,to facilitate the high-throughput and robust screening of metal replicated samples,we provide a deep learning analysis method to screen the“pseudo 3D”morphologies of CCVs imaged with 2D modalities.Collectively,our work establishes accessible ways to examine the 3D structure of biological samples and provide novel insights into the structure of plant CCVs.
基金This work was supported by the National Natural Science Foundation of China(Grant No.20243007)the Alexander von Humboldt Foundation+4 种基金the Program of Hundreds Talent of the Chinese Academy of SciencesShandong University(Hao Jingcheng) by the National Natural Science Foundation of China(Grant No.50275142) the Minis try of ScienceTechnology ofChina(Grant No.2202AA302609)(LiuWeimin).
文摘Self-assemblies of amphiphiles in solutions were investigated by using freeze-fracture transmission electron microscopy(FF-TEM). Especially, vesicles were characterized by FF-TEM and the transition of self-assemblies was determined. The stacked lamellar Lα-phase was prepared without shear forces by a chemical reaction. The stacked lamellar Lα-phase can be transformed into multilamellar vesicles by the shearing forces that occur when the stacked lamellar Lα-phase sample is turned upside down a few times. The multilamellar vesicles can also be transformed into unilamellar vesicles by high shearing forces. These transitions were demonstrated by FF-TEM measurements. Zn2+-induced vesicle formation in the single-chain surfactant solutions was first achieved.
