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Synaptic transmission relies on presynaptic neurotransmitter (NT) release from synaptic vesicles (SVs) and on NT detection by postsynaptic receptors. Transmission exists in two principal modes: action-potential (AP) evoked and AP-independent, “spontaneous” transmission. AP-evoked neurotransmission is considered the primary mode of inter-neuronal communication, whereas spontaneous transmission is required for neuronal development, homeostasis, and plasticity. While some synapses appear dedicated to spontaneous transmission only, all AP-responsive synapses also engage spontaneously, but whether this encodes functional information regarding their excitability is unknown. Here we report on functional interdependence of both transmission modes at individual synaptic contacts of Drosophila larval neuromuscular junctions (NMJs) which were identified by the presynaptic scaffolding protein Bruchpilot (BRP) and whose activities were quantified using the genetically encoded Ca2+ indicator GCaMP. Consistent with the role of BRP in organizing the AP-dependent release machinery (voltage-dependent Ca2+ channels and SV fusion machinery), most active BRP-positive synapses (>85%) responded to APs. At these synapses, the level of spontaneous activity was a predictor for their responsiveness to AP-stimulation. AP-stimulation resulted in cross-depletion of spontaneous activity and both transmission modes were affected by the non-specific Ca2+ channel blocker cadmium and engaged overlapping postsynaptic receptors. Thus, by using overlapping machinery, spontaneous transmission is a continuous, stimulus independent predictor for the AP-responsiveness of individual synapses.

Synaptic transmission relies on presynaptic neurotransmitter (NT) release from synaptic vesicles (SVs) and on NT detection by postsynaptic receptors. Transmission exists in two principal modes: action-potential (AP) evoked and AP-independent, “spontaneous” transmission. AP-evoked neurotransmission is considered the primary mode of inter-neuronal communication, whereas spontaneous transmission is required for neuronal development, homeostasis, and plasticity. While some synapses appear dedicated to spontaneous transmission only, all AP-responsive synapses also engage spontaneously, but whether this encodes functional information regarding their excitability is unknown. Here we report on functional interdependence of both transmission modes at individual synaptic contacts of Drosophila larval neuromuscular junctions (NMJs) which were identified by the presynaptic scaffolding protein Bruchpilot (BRP) and whose activities were quantified using the genetically encoded Ca2+ indicator GCaMP. Consistent with the role of BRP in organizing the AP-dependent release machinery (voltage-dependent Ca2+ channels and SV fusion machinery), most active BRP-positive synapses (>85%) responded to APs. At these synapses, the level of spontaneous activity was a predictor for their responsiveness to AP-stimulation. AP-stimulation resulted in cross-depletion of spontaneous activity and both transmission modes were affected by the non-specific Ca2+ channel blocker cadmium and engaged overlapping postsynaptic receptors. Thus, by using overlapping machinery, spontaneous transmission is a continuous, stimulus independent predictor for the AP-responsiveness of individual synapses.

Both paralogs of the calcium-dependent activator protein for secretion (CAPS) are required for exocytosis of synaptic vesicles (SVs) and large dense core vesicles (LDCVs). Despite approximately 80% sequence identity, CAPS1 and CAPS2 have distinct functions in promoting exocytosis of SVs and LDCVs in dorsal root ganglion (DRG) neurons. However, the molecular mechanisms underlying these differences remain enigmatic. In this study, we applied high- and super-resolution imaging techniques to systematically assess the subcellular localization of CAPS paralogs in DRG neurons deficient in both CAPS1 and CAPS2. CAPS1 was found to be more enriched at the synapses. Using – in-depth sequence analysis, we identified a unique CAPS1 N-terminal sequence, which we introduced into CAPS2. This CAPS1/2 chimera reproduced the pre-synaptic localization of CAPS1 and partially rescued synaptic transmission in neurons devoid of CAPS1 and CAPS2. Using immunoprecipitation combined with mass spectrometry, we identified CAPS1-specific interaction partners that could be responsible for its pre-synaptic enrichment. Taken together, these data suggest an important role of the CAPS1-N terminus in the localization of the protein at pre-synapses.

Both paralogs of the calcium-dependent activator protein for secretion (CAPS) are required for exocytosis of synaptic vesicles (SVs) and large dense core vesicles (LDCVs). Despite approximately 80% sequence identity, CAPS1 and CAPS2 have distinct functions in promoting exocytosis of SVs and LDCVs in dorsal root ganglion (DRG) neurons. However, the molecular mechanisms underlying these differences remain enigmatic. In this study, we applied high- and super-resolution imaging techniques to systematically assess the subcellular localization of CAPS paralogs in DRG neurons deficient in both CAPS1 and CAPS2. CAPS1 was found to be more enriched at the synapses. Using – in-depth sequence analysis, we identified a unique CAPS1 N-terminal sequence, which we introduced into CAPS2. This CAPS1/2 chimera reproduced the pre-synaptic localization of CAPS1 and partially rescued synaptic transmission in neurons devoid of CAPS1 and CAPS2. Using immunoprecipitation combined with mass spectrometry, we identified CAPS1-specific interaction partners that could be responsible for its pre-synaptic enrichment. Taken together, these data suggest an important role of the CAPS1-N terminus in the localization of the protein at pre-synapses.

Neuronal synapses encompass three compartments: presynaptic axon terminal, synaptic cleft, and postsynaptic dendrite. Each compartment contains densely packed molecular machineries that are involved in synaptic transmission. In recent years, emerging evidence indicates that the assembly of these membraneless substructures or assemblies that are not enclosed by membranes are driven by liquid-liquid phase separation. We review here recent studies that suggest the phase separation-mediated organization of these synaptic compartments. We discuss how synaptic function may be linked to its organization as biomolecular condensates. We conclude with a discussion of areas of future interest in the field for better understanding of the structural architecture of neuronal synapses and its contribution to synaptic functions.

Neuronal synapses encompass three compartments: presynaptic axon terminal, synaptic cleft, and postsynaptic dendrite. Each compartment contains densely packed molecular machineries that are involved in synaptic transmission. In recent years, emerging evidence indicates that the assembly of these membraneless substructures or assemblies that are not enclosed by membranes are driven by liquid-liquid phase separation. We review here recent studies that suggest the phase separation-mediated organization of these synaptic compartments. We discuss how synaptic function may be linked to its organization as biomolecular condensates. We conclude with a discussion of areas of future interest in the field for better understanding of the structural architecture of neuronal synapses and its contribution to synaptic functions.

Synaptic transmission is mediated by neurotransmitters that are stored in synaptic vesicles (SV) and released at the synaptic active zone (AZ). While in recent years major progress has been made in unraveling the molecular machinery responsible for SV docking, fusion and exocytosis, the mechanisms governing AZ protein and SV trafficking through axons still remain unclear. Here, we performed stop flow nerve ligation to examine axonal trafficking of endogenous AZ and SV proteins. Rat sciatic nerves were collected 1 h, 3 h and 8 h post ligation and processed for immunohistochemistry and electron microscopy. First, we followed the transport of an integral synaptic vesicle protein, SV2A and a SVassociated protein involved in SV trafficking, Rab3a, and observed that while SV2A accumulated on both sides of ligation, Rab3a was only noticeable in the proximal segment of the ligated nerve indicating that only SV trans-membrane protein SV2A displayed a bi-directional axonal transport. We then demonstrate that multiple AZ proteins accumulate rapidly on either side of the ligation with a timescale similar to that of SV2A. Overall, our data uncovers an unexpected robust bi-directional, coordinated-trafficking of SV and AZ proteins in peripheral nerves. This implies that pathological disruption of axonal trafficking will not only impair trafficking of newly synthesized proteins to the synapse but will also affect retrograde transport, leading to neuronal dysfunction and likely neurodegeneration. (c) 2021 The Authors. Published by Elsevier Inc. This is an open access article under the CC BY-NC-ND license (http://creativecommons.org/licenses/by-nc-nd/4.0/).

Chemical neurotransmission typically occurs through synapses. Previous ultrastructural examinations of monoamine neuron axon terminals often failed to identify a pre- and postsynaptic coupling, leading to the concept of "volume" transmission. Whether this results from intrinsic properties of these neurons remains undefined. We find that dopaminergic neurons in vitro establish a distinctive axonal arbor compared to glutamatergic or GABAergic neurons in both size and propensity of terminals to avoid direct contact with target neurons. While most dopaminergic varicosities are active and contain exocytosis proteins like synaptotagmin 1, only similar to 20% of these are synaptic. The active zone protein bassoon was found to be enriched in dopaminergic terminals that are in proximity to a target cell. Finally, we found that the proteins neurexin-1 alpha(SS4-) and neuroligin-1(A+B) play a critical role in the formation of synapses by dopamine (DA) neurons. Our findings suggest that DA neurons are endowed with a distinctive developmental connectivity program.

Neuronal synapses encompass three compartments: presynaptic axon terminal, synaptic cleft, and postsynaptic dendrite. Each compartment contains densely packed molecular machineries that are involved in synaptic transmission. In recent years, emerging evidence indicates that the assembly of these membraneless substructures or assemblies that are not enclosed by membranes are driven by liquid-liquid phase separation. We review here recent studies that suggest the phase separation-mediated organization of these synaptic compartments. We discuss how synaptic function may be linked to its organization as biomolecular condensates. We conclude with a discussion of areas of future interest in the field for better understanding of the structural architecture of neuronal synapses and its contribution to synaptic functions.

Synaptic transmission is mediated by neurotransmitters that are stored in synaptic vesicles (SV) and released at the synaptic active zone (AZ). While in recent years major progress has been made in unraveling the molecular machinery responsible for SV docking, fusion and exocytosis, the mechanisms governing AZ protein and SV trafficking through axons still remain unclear. Here, we performed stop flow nerve ligation to examine axonal trafficking of endogenous AZ and SV proteins. Rat sciatic nerves were collected 1 h, 3 h and 8 h post ligation and processed for immunohistochemistry and electron microscopy. First, we followed the transport of an integral synaptic vesicle protein, SV2A and a SVassociated protein involved in SV trafficking, Rab3a, and observed that while SV2A accumulated on both sides of ligation, Rab3a was only noticeable in the proximal segment of the ligated nerve indicating that only SV trans-membrane protein SV2A displayed a bi-directional axonal transport. We then demonstrate that multiple AZ proteins accumulate rapidly on either side of the ligation with a timescale similar to that of SV2A. Overall, our data uncovers an unexpected robust bi-directional, coordinated-trafficking of SV and AZ proteins in peripheral nerves. This implies that pathological disruption of axonal trafficking will not only impair trafficking of newly synthesized proteins to the synapse but will also affect retrograde transport, leading to neuronal dysfunction and likely neurodegeneration. (c) 2021 The Authors. Published by Elsevier Inc. This is an open access article under the CC BY-NC-ND license (http://creativecommons.org/licenses/by-nc-nd/4.0/).

Chemical neurotransmission typically occurs through synapses. Previous ultrastructural examinations of monoamine neuron axon terminals often failed to identify a pre- and postsynaptic coupling, leading to the concept of "volume" transmission. Whether this results from intrinsic properties of these neurons remains undefined. We find that dopaminergic neurons in vitro establish a distinctive axonal arbor compared to glutamatergic or GABAergic neurons in both size and propensity of terminals to avoid direct contact with target neurons. While most dopaminergic varicosities are active and contain exocytosis proteins like synaptotagmin 1, only similar to 20% of these are synaptic. The active zone protein bassoon was found to be enriched in dopaminergic terminals that are in proximity to a target cell. Finally, we found that the proteins neurexin-1 alpha(SS4-) and neuroligin-1(A+B) play a critical role in the formation of synapses by dopamine (DA) neurons. Our findings suggest that DA neurons are endowed with a distinctive developmental connectivity program.

Téma diplomové práce poukazuje na provádění přechodových oblastí mostů, jež úzce souvisí i se zemními pláněmi dopravních staveb. Proto řeším problematiku zemních plání v mé diplomové práci a poukazuji na jejich provádění v praxi. Zabývám se prováděním zkoušek a řeším chyby a nedostatky v provádění zemních plání a přechodových oblastí v praxi. V diplomové práci se zabývám konkrétními případy realizovaných staveb. Cílem práce je posoudit provádění mnou realizovaných staveb s teoretickými modelovými situacemi, kde bude možné poukázat na klady a zápory v provádění. Součástí práce bude řešení konkrétní stavby, a to z hlediska projektové dokumentace, kontrolního a zkušebního plánu, chyb provádění, návrh sanace zemní pláně a vyčíslení finančních nároků na provedení opravných a sanačních prací., The topic of the diploma thesis points to the implementation of transition areas of bridges. The topic of transition areas is closely related to the land plains of transport structures. Therefore, I address the issue of land plains in my thesis and point out their implementation in practice. I deal with the implementation of tests and solve errors and shortcomings in the implementation of land plains and transition areas in practice. In my diploma thesis I deal with specific cases of realized constructions. The aim of the work is to assess the implementation of the constructions I have implemented with theoretical model situations, where it will be possible to point out the pros and cons in the implementation. Part of the work will be the solution of a specific construction, in terms of project documentation, control and test plan, implementation errors, the design of the remediation of the ground plain and quantification of financial claims for repair and remediation work., Dopravní fakulta Jana Pernera, Uchazeč prezentoval komisi hlavní výsledky a závěry své diplomové práce. Po prezentaci diplomové práce zodpověděl dotazy vedoucího a připomínky oponenta diplomové práce (uvedeny v posudcích práce). V diskusi o diplomové práci podrobně zodpověděl všechny dotazy členů komise., Dokončená práce s úspěšnou obhajobou

In article the short characteristic about modern state of development projects HTGR is given. The main fields of using of technological heat which obtained from HTGR and perspective it’s using in power-consuming industry of Ukraine are pointed. The main advantages of HTGR in comparison with the different types of reactors and also problems that stay on the way them creation are analyzed. The maim perspectives of safe working of reactors with gas heat carrier are underlined., В даній статті наведено коротку характеристику по сучасному стану розвитку проектів ВТГР. Вказано області застосування технологічної теплоти, отриманої при роботі ВТГР та перспективи її застосування в енергоємній промисловості України. Проаналізовані основні переваги ВТГР в порівняння з реакторами інших типів, а також проблеми, що стоять на шляху їх створення. Підкреслені основні аспекти безпечної роботи ректорів з газовим теплоносієм.

Presynaptic active zones (AZ) contain many molecules essential for neurotransmitter release and are assembled in a highly organized manner. A network of adaptor proteins known as Cytomatrix at the AZ (CAZ), is important for shaping the structural characteristics of AZ. RIM-binding protein (RBP) family are binding partners of the CAZ protein RIM and also bind the voltage-gated calcium channels (VGCC) in mice and flies. Here, we investigated the physiological roles of RIMB-1, the homolog of RBPs in the nematode RIMB-1 is expressed broadly in neurons and predominantly localized at presynaptic sites. Loss-of-function animals of displayed slight defects in motility and response to pharmacological inhibition of synaptic transmission, suggesting a modest involvement of in synapse function. We analyzed genetic interactions of by testing candidate genes and by an unbiased forward genetic screen for enhancer. Both analyses identified the RIM homolog UNC-10 that acts together with RIMB-1 to regulate presynaptic localization of the P/Q-type VGCC UNC-2/Ca2. We also find that the precise localization of RIMB-1 to presynaptic sites requires presynaptic UNC-2/Ca2. RIMB-1 has multiple FN3 and SH3 domains. Our transgenic rescue analysis with RIMB-1 deletion constructs revealed a functional requirement of a C-terminal SH3 in regulating UNC-2/Ca2 localization. Together, these findings suggest a redundant role of RIMB-1/RBP and UNC-10/RIM to regulate the abundance of UNC-2/Ca2 at the presynaptic AZ in , depending on the bidirectional interplay between CAZ adapter and channel proteins.Presynaptic active zones (AZ) are highly organized structures for synaptic transmission with characteristic networks of adapter proteins called Cytomatrix at the AZ (CAZ). In this study, we characterized a CAZ protein RIMB-1, named for RIM-binding protein (RBP), in the nematode Through systematic analyses of genetic interactions and an unbiased genetic enhancer screen of , we revealed a redundant role of tw

Presynaptic active zones (AZ) contain many molecules essential for neurotransmitter release and are assembled in a highly organized manner. A network of adaptor proteins known as Cytomatrix at the AZ (CAZ), is important for shaping the structural characteristics of AZ. RIM-binding protein (RBP) family are binding partners of the CAZ protein RIM and also bind the voltage-gated calcium channels (VGCC) in mice and flies. Here, we investigated the physiological roles of RIMB-1, the homolog of RBPs in the nematode RIMB-1 is expressed broadly in neurons and predominantly localized at presynaptic sites. Loss-of-function animals of displayed slight defects in motility and response to pharmacological inhibition of synaptic transmission, suggesting a modest involvement of in synapse function. We analyzed genetic interactions of by testing candidate genes and by an unbiased forward genetic screen for enhancer. Both analyses identified the RIM homolog UNC-10 that acts together with RIMB-1 to regulate presynaptic localization of the P/Q-type VGCC UNC-2/Ca2. We also find that the precise localization of RIMB-1 to presynaptic sites requires presynaptic UNC-2/Ca2. RIMB-1 has multiple FN3 and SH3 domains. Our transgenic rescue analysis with RIMB-1 deletion constructs revealed a functional requirement of a C-terminal SH3 in regulating UNC-2/Ca2 localization. Together, these findings suggest a redundant role of RIMB-1/RBP and UNC-10/RIM to regulate the abundance of UNC-2/Ca2 at the presynaptic AZ in , depending on the bidirectional interplay between CAZ adapter and channel proteins.Presynaptic active zones (AZ) are highly organized structures for synaptic transmission with characteristic networks of adapter proteins called Cytomatrix at the AZ (CAZ). In this study, we characterized a CAZ protein RIMB-1, named for RIM-binding protein (RBP), in the nematode Through systematic analyses of genetic interactions and an unbiased genetic enhancer screen of , we revealed a redundant role of tw

Presynaptic active zones (AZs) contain many molecules essential for neurotransmitter release and are assembled in a highly organized manner. A network of adaptor proteins known as cytomatrix at the AZ (CAZ) is important for shaping the structural characteristics of AZ. Rab3-interacting molecule (RIM)-binding protein (RBP) family are binding partners of the CAZ protein RIM and also bind the voltage-gated calcium channels (VGCCs) in mice and flies. Here, we investigated the physiological roles of RIMB-1, the homolog of RBPs in the nematode Caenorhabditis elegans RIMB-1 is expressed broadly in neurons and predominantly localized at presynaptic sites. Loss-of-function animals of rimb-1 displayed slight defects in motility and response to pharmacological inhibition of synaptic transmission, suggesting a modest involvement of rimb-1 in synapse function. We analyzed genetic interactions of rimb-1 by testing candidate genes and by an unbiased forward genetic screen for rimb-1 enhancer. Both analyses identified the RIM homolog UNC-10 that acts together with RIMB-1 to regulate presynaptic localization of the P/Q-type VGCC UNC-2/Cav2. We also find that the precise localization of RIMB-1 to presynaptic sites requires presynaptic UNC-2/Cav2. RIMB-1 has multiple FN3 and SH3 domains. Our transgenic rescue analysis with RIMB-1 deletion constructs revealed a functional requirement of a C-terminal SH3 in regulating UNC-2/Cav2 localization. Together, these findings suggest a redundant role of RIMB-1/RBP and UNC-10/RIM to regulate the abundance of UNC-2/Cav2 at the presynaptic AZ in C. elegans, depending on the bidirectional interplay between CAZ adaptor and channel proteins.SIGNIFICANCE STATEMENT Presynaptic active zones (AZs) are highly organized structures for synaptic transmission with characteristic networks of adaptor proteins called cytomatrix at the AZ (CAZ). In this study, we characterized a CAZ protein RIMB-1, named for RIM-binding protein (RBP), in the nematode Caenorhabditi

Presynaptic active zones (AZ) contain many molecules essential for neurotransmitter release and are assembled in a highly organized manner. A network of adaptor proteins known as Cytomatrix at the AZ (CAZ), is important for shaping the structural characteristics of AZ. RIM-binding protein (RBP) family are binding partners of the CAZ protein RIM and also bind the voltage-gated calcium channels (VGCC) in mice and flies. Here, we investigated the physiological roles of RIMB-1, the homolog of RBPs in the nematode RIMB-1 is expressed broadly in neurons and predominantly localized at presynaptic sites. Loss-of-function animals of displayed slight defects in motility and response to pharmacological inhibition of synaptic transmission, suggesting a modest involvement of in synapse function. We analyzed genetic interactions of by testing candidate genes and by an unbiased forward genetic screen for enhancer. Both analyses identified the RIM homolog UNC-10 that acts together with RIMB-1 to regulate presynaptic localization of the P/Q-type VGCC UNC-2/Ca2. We also find that the precise localization of RIMB-1 to presynaptic sites requires presynaptic UNC-2/Ca2. RIMB-1 has multiple FN3 and SH3 domains. Our transgenic rescue analysis with RIMB-1 deletion constructs revealed a functional requirement of a C-terminal SH3 in regulating UNC-2/Ca2 localization. Together, these findings suggest a redundant role of RIMB-1/RBP and UNC-10/RIM to regulate the abundance of UNC-2/Ca2 at the presynaptic AZ in , depending on the bidirectional interplay between CAZ adapter and channel proteins.Presynaptic active zones (AZ) are highly organized structures for synaptic transmission with characteristic networks of adapter proteins called Cytomatrix at the AZ (CAZ). In this study, we characterized a CAZ protein RIMB-1, named for RIM-binding protein (RBP), in the nematode Through systematic analyses of genetic interactions and an unbiased genetic enhancer screen of , we revealed a redundant role of tw

The arrival of an action potential (AP) at a synaptic terminal elicits highly synchronized quanta release. Repetitive APs produce successive synaptic vesicle (SV) fusions that require management of spent SV components in the presynaptic membrane with minimum disturbance of the secretory apparatus. To this end, the synaptic machinery is structured accordingly to the strength and the range of frequencies at which each particular synapse operates. This results in variations in the number and dimension of Active Zones (AZs), amount and distribution of SVs, and probably, in the primary endocytic mechanisms they use. Understanding better how these structural differences determine the functional response in each case has been a matter of long-term interest. Here we review the structural and functional properties of three distinct types of synapses: the neuromuscular junction (NMJ; a giant, highly reliable synapse that must exocytose a large number of quanta with each stimulus to guarantee excitation of the postsynaptic cell), the hippocampal excitatory small synapse (which most often has a single release site and a relatively small pool of vesicles), and the cerebellar mossy fiber-granule cell synapse (which possesses hundreds of release sites and is able to translocate, dock and prime vesicles at high speed). We will focus on how the release apparatus is organized in each case, the relative amount of vesicular membrane that needs to be accommodated within the periAZ upon stimulation, the different mechanisms for retrieving the excess of membrane and finally, how these factors may influence the functioning of the release sites.

The arrival of an action potential (AP) at a synaptic terminal elicits highly synchronized quanta release. Repetitive APs produce successive synaptic vesicle (SV) fusions that require management of spent SV components in the presynaptic membrane with minimum disturbance of the secretory apparatus. To this end, the synaptic machinery is structured accordingly to the strength and the range of frequencies at which each particular synapse operates. This results in variations in the number and dimension of Active Zones (AZs), amount and distribution of SVs, and probably, in the primary endocytic mechanisms they use. Understanding better how these structural differences determine the functional response in each case has been a matter of long-term interest. Here we review the structural and functional properties of three distinct types of synapses: the neuromuscular junction (NMJ; a giant, highly reliable synapse that must exocytose a large number of quanta with each stimulus to guarantee excitation of the postsynaptic cell), the hippocampal excitatory small synapse (which most often has a single release site and a relatively small pool of vesicles), and the cerebellar mossy fiber-granule cell synapse (which possesses hundreds of release sites and is able to translocate, dock and prime vesicles at high speed). We will focus on how the release apparatus is organized in each case, the relative amount of vesicular membrane that needs to be accommodated within the periAZ upon stimulation, the different mechanisms for retrieving the excess of membrane and finally, how these factors may influence the functioning of the release sites.

The arrival of an action potential (AP) at a synaptic terminal elicits highly synchronized quanta release. Repetitive APs produce successive synaptic vesicle (SV) fusions that require management of spent SV components in the presynaptic membrane with minimum disturbance of the secretory apparatus. To this end, the synaptic machinery is structured accordingly to the strength and the range of frequencies at which each particular synapse operates. This results in variations in the number and dimension of Active Zones (AZs), amount and distribution of SVs, and probably, in the primary endocytic mechanisms they use. Understanding better how these structural differences determine the functional response in each case has been a matter of long-term interest. Here we review the structural and functional properties of three distinct types of synapses: the neuromuscular junction (NMJ; a giant, highly reliable synapse that must exocytose a large number of quanta with each stimulus to guarantee excitation of the postsynaptic cell), the hippocampal excitatory small synapse (which most often has a single release site and a relatively small pool of vesicles), and the cerebellar mossy fiber-granule cell synapse (which possesses hundreds of release sites and is able to translocate, dock and prime vesicles at high speed). We will focus on how the release apparatus is organized in each case, the relative amount of vesicular membrane that needs to be accommodated within the periAZ upon stimulation, the different mechanisms for retrieving the excess of membrane and finally, how these factors may influence the functioning of the release sites.

The arrival of an action potential (AP) at a synaptic terminal elicits highly synchronized quanta release. Repetitive APs produce successive synaptic vesicle (SV) fusions that require management of spent SV components in the presynaptic membrane with minimum disturbance of the secretory apparatus. To this end, the synaptic machinery is structured accordingly to the strength and the range of frequencies at which each particular synapse operates. This results in variations in the number and dimension of Active Zones (AZs), amount and distribution of SVs, and probably, in the primary endocytic mechanisms they use. Understanding better how these structural differences determine the functional response in each case has been a matter of long-term interest. Here we review the structural and functional properties of three distinct types of synapses: the neuromuscular junction (NMJ; a giant, highly reliable synapse that must exocytose a large number of quanta with each stimulus to guarantee excitation of the postsynaptic cell), the hippocampal excitatory small synapse (which most often has a single release site and a relatively small pool of vesicles), and the cerebellar mossy fiber-granule cell synapse (which possesses hundreds of release sites and is able to translocate, dock and prime vesicles at high speed). We will focus on how the release apparatus is organized in each case, the relative amount of vesicular membrane that needs to be accommodated within the periAZ upon stimulation, the different mechanisms for retrieving the excess of membrane and finally, how these factors may influence the functioning of the release sites.

The arrival of an action potential (AP) at a synaptic terminal elicits highly synchronized quanta release. Repetitive APs produce successive synaptic vesicle (SV) fusions that require management of spent SV components in the presynaptic membrane with minimum disturbance of the secretory apparatus. To this end, the synaptic machinery is structured accordingly to the strength and the range of frequencies at which each particular synapse operates. This results in variations in the number and dimension of Active Zones (AZs), amount and distribution of SVs, and probably, in the primary endocytic mechanisms they use. Understanding better how these structural differences determine the functional response in each case has been a matter of long-term interest. Here we review the structural and functional properties of three distinct types of synapses: the neuromuscular junction (NMJ; a giant, highly reliable synapse that must exocytose a large number of quanta with each stimulus to guarantee excitation of the postsynaptic cell), the hippocampal excitatory small synapse (which most often has a single release site and a relatively small pool of vesicles), and the cerebellar mossy fiber-granule cell synapse (which possesses hundreds of release sites and is able to translocate, dock and prime vesicles at high speed). We will focus on how the release apparatus is organized in each case, the relative amount of vesicular membrane that needs to be accommodated within the periAZ upon stimulation, the different mechanisms for retrieving the excess of membrane and finally, how these factors may influence the functioning of the release sites.

The arrival of an action potential (AP) at a synaptic terminal elicits highly synchronized quanta release. Repetitive APs produce successive synaptic vesicle (SV) fusions that require management of spent SV components in the presynaptic membrane with minimum disturbance of the secretory apparatus. To this end, the synaptic machinery is structured accordingly to the strength and the range of frequencies at which each particular synapse operates. This results in variations in the number and dimension of Active Zones (AZs), amount and distribution of SVs, and probably, in the primary endocytic mechanisms they use. Understanding better how these structural differences determine the functional response in each case has been a matter of long-term interest. Here we review the structural and functional properties of three distinct types of synapses: the neuromuscular junction (NMJ; a giant, highly reliable synapse that must exocytose a large number of quanta with each stimulus to guarantee excitation of the postsynaptic cell), the hippocampal excitatory small synapse (which most often has a single release site and a relatively small pool of vesicles), and the cerebellar mossy fiber-granule cell synapse (which possesses hundreds of release sites and is able to translocate, dock and prime vesicles at high speed). We will focus on how the release apparatus is organized in each case, the relative amount of vesicular membrane that needs to be accommodated within the periAZ upon stimulation, the different mechanisms for retrieving the excess of membrane and finally, how these factors may influence the functioning of the release sites.

The arrival of an action potential (AP) at a synaptic terminal elicits highly synchronized quanta release. Repetitive APs produce successive synaptic vesicle (SV) fusions that require management of spent SV components in the presynaptic membrane with minimum disturbance of the secretory apparatus. To this end, the synaptic machinery is structured accordingly to the strength and the range of frequencies at which each particular synapse operates. This results in variations in the number and dimension of Active Zones (AZs), amount and distribution of SVs, and probably, in the primary endocytic mechanisms they use. Understanding better how these structural differences determine the functional response in each case has been a matter of long-term interest. Here we review the structural and functional properties of three distinct types of synapses: the neuromuscular junction (NMJ; a giant, highly reliable synapse that must exocytose a large number of quanta with each stimulus to guarantee excitation of the postsynaptic cell), the hippocampal excitatory small synapse (which most often has a single release site and a relatively small pool of vesicles), and the cerebellar mossy fiber-granule cell synapse (which possesses hundreds of release sites and is able to translocate, dock and prime vesicles at high speed). We will focus on how the release apparatus is organized in each case, the relative amount of vesicular membrane that needs to be accommodated within the periAZ upon stimulation, the different mechanisms for retrieving the excess of membrane and finally, how these factors may influence the functioning of the release sites.

The thermodynamic entropy production and the variational functional for diffusion of one species of particles from a flat membrane to an irregular membrane is studied in detail. The computations for the Laplacian field are realized with a special superconvergent overrelaxation algorithm which is of a very high precision. The possible irregular geometries examined in this work are all non-scaling geometries, all variants of an initial fractal generator. The calculations are realized for distances of the two membranes higher than the bump of the geometric irregularity, a case which we call here "the Far Field". The basic conclusion arising for this study is that the active zone approximation is always valid with a good precision. © 2010 American Institute of Physics., SCOPUS: cp.p, info:eu-repo/semantics/published

The thermodynamic entropy production and the variational functional for diffusion of one species of particles from a flat membrane to an irregular membrane is studied in detail. The computations for the Laplacian field are realized with a special superconvergent overrelaxation algorithm which is of a very high precision. The possible irregular geometries examined in this work are all non-scaling geometries, all variants of an initial fractal generator. The calculations are realized for distances of the two membranes higher than the bump of the geometric irregularity, a case which we call here "the Far Field". The basic conclusion arising for this study is that the active zone approximation is always valid with a good precision. © 2010 American Institute of Physics., SCOPUS: cp.p, info:eu-repo/semantics/published

Despite a voluminous knowledge concerning the composition of active zones, very little is known about how these presynaptic specializations are assembled. Furthermore, with a few exceptions little is known about the role that active zone molecules play in synaptic function. In this thesis, I describe the identification and characterization of Drosophila mutants for a newly-identified serine-arginine protein kinase, which has been termed serine-argining protein kinas at 79D (SRPK79D). In Chapter 2 I describe a series of experiments involving quantitative imaging, electron microscopy and electrophysiology. The results of these experiments indicate that SRPK79D is a negative regulator of T-bar assembly and have lead to a new model of active zone assembly. Specifically, the data suggest that in wild type animals active zone constituents are transported from the cell soma to the synapse where they are locally assembled. During transport assembly of these components must be constitutively repressed through the action of SRPK79D to prevent ectopic T-bar formation.In Chapter 3 I present the initial characterization of a synaptic function deficit at the neuromuscular junction of srpk79D mutants. In otherwise wild type animals, over-expression of the vesicular glutamate transporter (VGlut) leads to an increase in the amplitude of spontaneous fusion events (minis) and an inversely-proportional, compensatory decrease in the average number of vesicles that fuse in response to presynaptic stimulation. Thus, through this process, which we term reverse homeostatic compensation, the average postsynaptic response to presynaptic stimulation in VGlut over-expressing animals is identical to animals expressing normal levels of VGlut. In srpk79D mutants, VGlut over-expression causes an identical increase in the average mini amplitude. Reverse homeostatic compensation also ensues. However, compensation is excessive. Consequently, the average postsynaptic response to presynaptic stimulation

Despite a voluminous knowledge concerning the composition of active zones, very little is known about how these presynaptic specializations are assembled. Furthermore, with a few exceptions little is known about the role that active zone molecules play in synaptic function. In this thesis, I describe the identification and characterization of Drosophila mutants for a newly-identified serine-arginine protein kinase, which has been termed serine-argining protein kinas at 79D (SRPK79D). In Chapter 2 I describe a series of experiments involving quantitative imaging, electron microscopy and electrophysiology. The results of these experiments indicate that SRPK79D is a negative regulator of T-bar assembly and have lead to a new model of active zone assembly. Specifically, the data suggest that in wild type animals active zone constituents are transported from the cell soma to the synapse where they are locally assembled. During transport assembly of these components must be constitutively repressed through the action of SRPK79D to prevent ectopic T-bar formation.In Chapter 3 I present the initial characterization of a synaptic function deficit at the neuromuscular junction of srpk79D mutants. In otherwise wild type animals, over-expression of the vesicular glutamate transporter (VGlut) leads to an increase in the amplitude of spontaneous fusion events (minis) and an inversely-proportional, compensatory decrease in the average number of vesicles that fuse in response to presynaptic stimulation. Thus, through this process, which we term reverse homeostatic compensation, the average postsynaptic response to presynaptic stimulation in VGlut over-expressing animals is identical to animals expressing normal levels of VGlut. In srpk79D mutants, VGlut over-expression causes an identical increase in the average mini amplitude. Reverse homeostatic compensation also ensues. However, compensation is excessive. Consequently, the average postsynaptic response to presynaptic stimulation

This paper deals with the fine structure of the fibrillar centres of the nucleolus inAllium cepa cells in ultrathin, sections of in vivo fixed roots. The ultrastructural observations have allowed us to consider each nucleolar fibrillar centre as an active zone in the nucleolar chromatin loop, and to propose a possible model for the organization of the different components of the nucleolus within it.

Bakalářská práce se v teoretické části věnuje deformacím způsobených účinkem mrazu, především však mrazovým zdvihům, jejich postupným rozvojem a faktory ovlivňující jejich vznik. Dále rozdílnými způsoby měření míry namrzavosti v České republice a jejich porovnání. Seznámení s odlišnými metodami měření míry namrzavosti ve vybraných zemích v rámci Evropské Unie. Praktická část je zaměřena na průběh mrazového zdvihu vzorků s a bez použití tepelněizolační látky., The bachelor thesis deals with the theoretical part of the deformation caused by frost action, but primarily frost heave, their gradual development and factors affecting their final formation. Furthermore, different ways of measuring the frost susceptibility in the Czech Republic and their comparison. Introduction to the different methods of measuring frost susceptibility in selected countries within the European Union. The practical part is focused on the process of frost heave samples with and without thermal insulation material.

Bakalářská práce se v teoretické části věnuje deformacím způsobených účinkem mrazu, především však mrazovým zdvihům, jejich postupným rozvojem a faktory ovlivňující jejich vznik. Dále rozdílnými způsoby měření míry namrzavosti v České republice a jejich porovnání. Seznámení s odlišnými metodami měření míry namrzavosti ve vybraných zemích v rámci Evropské Unie. Praktická část je zaměřena na průběh mrazového zdvihu vzorků s a bez použití tepelněizolační látky., The bachelor thesis deals with the theoretical part of the deformation caused by frost action, but primarily frost heave, their gradual development and factors affecting their final formation. Furthermore, different ways of measuring the frost susceptibility in the Czech Republic and their comparison. Introduction to the different methods of measuring frost susceptibility in selected countries within the European Union. The practical part is focused on the process of frost heave samples with and without thermal insulation material.

Dizertační práce představuje výsledky numerické studie provedené s pomocí 3D diskrétního mezo–úrovňového mřížkově–částicového (lattice–particle) modelu betonu. Již existující model byl rozšířen o prostorovou variabilitu některých materiálových parametrů pomocí náhodného pole. Laboratorní experimenty z literatury posloužily jednak k identifikaci parametrů, zároveň také umožnily ověřit správné chování modelu s určenými parametry. Se základním i rozšířeným modelem byla provedena série numerických simulací zaměřená na popis vlivu míry proměnlivosti náhodného pole definované korelační délkou na celkovou únosnost prvku. Závěr práce je věnován popisu oblasti, ve které je během zatěžování uvolněna většina lomové energie, a to z hlediska její velikosti i tvaru. Tato zóna rozhoduje o celkové pevnosti prvku a jak je ukázáno, její tvar i velikost nejsou konstantní, ale závisí na typu zatěžování (přítomnost zářezu) a také na korelační délce náhodného pole., The thesis presents results of a numerical study on the performance of 3D discrete meso–scale lattice–particle model of concrete. The existing model was extended by introducing the spatial variability of chosen material parameter in form of random field. An experimental data from bending tests on notched and unnotched beams was exploited for the identification of model parameters as well as for the subsequent validation of its performance. With the basic and the extended randomized version of the model, numerical simulations were calculated so that the influence of the rate of fluctuation of the random field (governed by the correlation length) could be observed. The final part of the thesis describes the region in the beam active during the test in which the most of the fracture energy is released in terms of its size and shape. This region defines the strength of the whole member and as shown in the thesis, it does not have a constant size but it is influenced by the geometrical setup and the correlation length of the random field.

Bakalářská práce se v teoretické části věnuje deformacím způsobených účinkem mrazu, především však mrazovým zdvihům, jejich postupným rozvojem a faktory ovlivňující jejich vznik. Dále rozdílnými způsoby měření míry namrzavosti v České republice a jejich porovnání. Seznámení s odlišnými metodami měření míry namrzavosti ve vybraných zemích v rámci Evropské Unie. Praktická část je zaměřena na průběh mrazového zdvihu vzorků s a bez použití tepelněizolační látky., The bachelor thesis deals with the theoretical part of the deformation caused by frost action, but primarily frost heave, their gradual development and factors affecting their final formation. Furthermore, different ways of measuring the frost susceptibility in the Czech Republic and their comparison. Introduction to the different methods of measuring frost susceptibility in selected countries within the European Union. The practical part is focused on the process of frost heave samples with and without thermal insulation material.

Dizertační práce představuje výsledky numerické studie provedené s pomocí 3D diskrétního mezo–úrovňového mřížkově–částicového (lattice–particle) modelu betonu. Již existující model byl rozšířen o prostorovou variabilitu některých materiálových parametrů pomocí náhodného pole. Laboratorní experimenty z literatury posloužily jednak k identifikaci parametrů, zároveň také umožnily ověřit správné chování modelu s určenými parametry. Se základním i rozšířeným modelem byla provedena série numerických simulací zaměřená na popis vlivu míry proměnlivosti náhodného pole definované korelační délkou na celkovou únosnost prvku. Závěr práce je věnován popisu oblasti, ve které je během zatěžování uvolněna většina lomové energie, a to z hlediska její velikosti i tvaru. Tato zóna rozhoduje o celkové pevnosti prvku a jak je ukázáno, její tvar i velikost nejsou konstantní, ale závisí na typu zatěžování (přítomnost zářezu) a také na korelační délce náhodného pole., The thesis presents results of a numerical study on the performance of 3D discrete meso–scale lattice–particle model of concrete. The existing model was extended by introducing the spatial variability of chosen material parameter in form of random field. An experimental data from bending tests on notched and unnotched beams was exploited for the identification of model parameters as well as for the subsequent validation of its performance. With the basic and the extended randomized version of the model, numerical simulations were calculated so that the influence of the rate of fluctuation of the random field (governed by the correlation length) could be observed. The final part of the thesis describes the region in the beam active during the test in which the most of the fracture energy is released in terms of its size and shape. This region defines the strength of the whole member and as shown in the thesis, it does not have a constant size but it is influenced by the geometrical setup and the correlation length of the random field.

Dizertační práce představuje výsledky numerické studie provedené s pomocí 3D diskrétního mezo–úrovňového mřížkově–částicového (lattice–particle) modelu betonu. Již existující model byl rozšířen o prostorovou variabilitu některých materiálových parametrů pomocí náhodného pole. Laboratorní experimenty z literatury posloužily jednak k identifikaci parametrů, zároveň také umožnily ověřit správné chování modelu s určenými parametry. Se základním i rozšířeným modelem byla provedena série numerických simulací zaměřená na popis vlivu míry proměnlivosti náhodného pole definované korelační délkou na celkovou únosnost prvku. Závěr práce je věnován popisu oblasti, ve které je během zatěžování uvolněna většina lomové energie, a to z hlediska její velikosti i tvaru. Tato zóna rozhoduje o celkové pevnosti prvku a jak je ukázáno, její tvar i velikost nejsou konstantní, ale závisí na typu zatěžování (přítomnost zářezu) a také na korelační délce náhodného pole., The thesis presents results of a numerical study on the performance of 3D discrete meso–scale lattice–particle model of concrete. The existing model was extended by introducing the spatial variability of chosen material parameter in form of random field. An experimental data from bending tests on notched and unnotched beams was exploited for the identification of model parameters as well as for the subsequent validation of its performance. With the basic and the extended randomized version of the model, numerical simulations were calculated so that the influence of the rate of fluctuation of the random field (governed by the correlation length) could be observed. The final part of the thesis describes the region in the beam active during the test in which the most of the fracture energy is released in terms of its size and shape. This region defines the strength of the whole member and as shown in the thesis, it does not have a constant size but it is influenced by the geometrical setup and the correlation length of the random field.

Dizertační práce představuje výsledky numerické studie provedené s pomocí 3D diskrétního mezo–úrovňového mřížkově–částicového (lattice–particle) modelu betonu. Již existující model byl rozšířen o prostorovou variabilitu některých materiálových parametrů pomocí náhodného pole. Laboratorní experimenty z literatury posloužily jednak k identifikaci parametrů, zároveň také umožnily ověřit správné chování modelu s určenými parametry. Se základním i rozšířeným modelem byla provedena série numerických simulací zaměřená na popis vlivu míry proměnlivosti náhodného pole definované korelační délkou na celkovou únosnost prvku. Závěr práce je věnován popisu oblasti, ve které je během zatěžování uvolněna většina lomové energie, a to z hlediska její velikosti i tvaru. Tato zóna rozhoduje o celkové pevnosti prvku a jak je ukázáno, její tvar i velikost nejsou konstantní, ale závisí na typu zatěžování (přítomnost zářezu) a také na korelační délce náhodného pole., The thesis presents results of a numerical study on the performance of 3D discrete meso–scale lattice–particle model of concrete. The existing model was extended by introducing the spatial variability of chosen material parameter in form of random field. An experimental data from bending tests on notched and unnotched beams was exploited for the identification of model parameters as well as for the subsequent validation of its performance. With the basic and the extended randomized version of the model, numerical simulations were calculated so that the influence of the rate of fluctuation of the random field (governed by the correlation length) could be observed. The final part of the thesis describes the region in the beam active during the test in which the most of the fracture energy is released in terms of its size and shape. This region defines the strength of the whole member and as shown in the thesis, it does not have a constant size but it is influenced by the geometrical setup and the correlation length of the random field.

Bakalářská práce se v teoretické části věnuje deformacím způsobených účinkem mrazu, především však mrazovým zdvihům, jejich postupným rozvojem a faktory ovlivňující jejich vznik. Dále rozdílnými způsoby měření míry namrzavosti v České republice a jejich porovnání. Seznámení s odlišnými metodami měření míry namrzavosti ve vybraných zemích v rámci Evropské Unie. Praktická část je zaměřena na průběh mrazového zdvihu vzorků s a bez použití tepelněizolační látky., The bachelor thesis deals with the theoretical part of the deformation caused by frost action, but primarily frost heave, their gradual development and factors affecting their final formation. Furthermore, different ways of measuring the frost susceptibility in the Czech Republic and their comparison. Introduction to the different methods of measuring frost susceptibility in selected countries within the European Union. The practical part is focused on the process of frost heave samples with and without thermal insulation material.

Bakalářská práce se v teoretické části věnuje deformacím způsobených účinkem mrazu, především však mrazovým zdvihům, jejich postupným rozvojem a faktory ovlivňující jejich vznik. Dále rozdílnými způsoby měření míry namrzavosti v České republice a jejich porovnání. Seznámení s odlišnými metodami měření míry namrzavosti ve vybraných zemích v rámci Evropské Unie. Praktická část je zaměřena na průběh mrazového zdvihu vzorků s a bez použití tepelněizolační látky., The bachelor thesis deals with the theoretical part of the deformation caused by frost action, but primarily frost heave, their gradual development and factors affecting their final formation. Furthermore, different ways of measuring the frost susceptibility in the Czech Republic and their comparison. Introduction to the different methods of measuring frost susceptibility in selected countries within the European Union. The practical part is focused on the process of frost heave samples with and without thermal insulation material.