Reduction Of Vesicles Research Articles

Changes of synaptic vesicles in three-dimensional synapse models by treatment with umbelliferone in scopolamine-induced hippocampal injury model

The neuroprotective effects of umbelliferone (UMB) were visualized in three-dimensional (3D) images on vesicle density changes of organotypic hippocampal slice tissues (OHSCs) induced by scopolamine by high voltage electron microscopy. Observations revealed that the number of vesicles decreased in OHSCs induced by scopolamine, and UMB was found to inhibit scopolamine-induced reduction in vesicles, resulting in an increase in vesicle count. These 3D models provide valuable insight for understanding the increase of synapse vesicles in hippocampal tissues treated with UMB.

Bladder epithelial cell phosphate transporter inhibition protects mice against uropathogenic Escherichia coli infection.

Urinary tract infections are predominantly caused by uropathogenic Escherichia coli (UPEC). UPEC infects bladder epithelial cells (BECs) via fusiform vesicles, escapes into the cytosol to evade exocytosis, and establishes intracellular bacterial communities (IBCs) for the next round of infection. The UPEC vesicle escape mechanism remains unclear. Here we show that UPEC senses host immune responses and initiates escape by upregulating a key phospholipase. The UPEC phospholipase PldA disrupts the vesicle membrane, and pldA expression is activated by phosphate reduction in vesicles. The host phosphate transporter PIT1 is located on the fusiform vesicle membrane, transporting phosphate into the cytosol. UPEC infection upregulates PIT1 via nuclear factor κB (NF-κB), resulting in phosphate reduction. Silencing PIT1 blocks UPEC vesicle escape in BECs, inhibits IBC formation in mouse bladders, and protects mice from UPEC infection. Our results shed light on pathogenic bacteria responding to intracellular phosphate shortage and tackling host defense and provide insights for development of new therapeutic agents to treat UPEC infection.

Formulation and characterization of novel liposomes containing histidine for encapsulation of a poorly soluble vitamin

In this study, development of folic acid-loaded liposomes using a basic amino acid, histidine as a solubilizing agent for folic acid was presented, which tackled the poor solubility of this vitamin. The effect of the liposomal membrane modifiers, cholesterol and SPAN 20 on the characteristics of the final formulations was examined. Liposomes prepared from a commercially available purified soybean lecithin (Phospholipon 90G) by proliposome method were between 503 and 877 nm in average diameter, where cholesterol induced enlargement and SPAN.20 reduction of vesicles. High encapsulation efficiency of 84% and drug loading of 0.123 mg g−1 were achieved, irrespective to the composition. According to AFM images, folic acid-loaded liposomes of a fraction with a nano size were flattened compared to globular empty liposomes. FTIR analysis revealed possible interactions between phospholipids and histidine, while DSC study suggested interactions between folic acid and lipids during heating. Release study done by a Franz diffusion cell showed prolonged release of folic acid from liposomes and the release rate was determined by folic acid solubility.

A rice Serine/Threonine receptor-like kinase regulates arbuscular mycorrhizal symbiosis at the peri-arbuscular membrane

In terrestrial ecosystems most plant species live in mutualistic symbioses with nutrient-delivering arbuscular mycorrhizal (AM) fungi. Establishment of AM symbioses includes transient, intracellular formation of fungal feeding structures, the arbuscules. A plant-derived peri-arbuscular membrane (PAM) surrounds the arbuscules, mediating reciprocal nutrient exchange. Signaling at the PAM must be well coordinated to achieve this dynamic cellular intimacy. Here, we identify the PAM-specific Arbuscular Receptor-like Kinase 1 (ARK1) from maize and rice to condition sustained AM symbiosis. Mutation of rice ARK1 causes a significant reduction in vesicles, the fungal storage structures, and a concomitant reduction in overall root colonization by the AM fungus Rhizophagus irregularis. Arbuscules, although less frequent in the ark1 mutant, are morphologically normal. Co-cultivation with wild-type plants restores vesicle and spore formation, suggesting ARK1 function is required for the completion of the fungal life-cycle, thereby defining a functional stage, post arbuscule development.

Piccolo Promotes Vesicle Replenishment at a Fast Central Auditory Synapse.

Piccolo and Bassoon are the two largest cytomatrix of the active zone (CAZ) proteins involved in scaffolding and regulating neurotransmitter release at presynaptic active zones (AZs), but have long been discussed as being functionally redundant. We employed genetic manipulation to bring forth and segregate the role of Piccolo from that of Bassoon at central auditory synapses of the cochlear nucleus—the endbulbs of Held. These synapses specialize in high frequency synaptic transmission, ideally poised to reveal even subtle deficits in the regulation of neurotransmitter release upon molecular perturbation. Combining semi-quantitative immunohistochemistry, electron microscopy, and in vitro and in vivo electrophysiology we first studied signal transmission in Piccolo-deficient mice. Our analysis was not confounded by a cochlear deficit, as a short isoform of Piccolo (“Piccolino”) present at the upstream ribbon synapses of cochlear inner hair cells (IHC), is unaffected by the mutation. Disruption of Piccolo increased the abundance of Bassoon at the AZs of endbulbs, while that of RIM1 was reduced and other CAZ proteins remained unaltered. Presynaptic fiber stimulation revealed smaller amplitude of the evoked excitatory postsynaptic currents (eEPSC), while eEPSC kinetics as well as miniature EPSCs (mEPSCs) remained unchanged. Cumulative analysis of eEPSC trains indicated that the reduced eEPSC amplitude of Piccolo-deficient endbulb synapses is primarily due to a reduced readily releasable pool (RRP) of synaptic vesicles (SV), as was corroborated by a reduction of vesicles at the AZ found on an ultrastructural level. Release probability seemed largely unaltered. Recovery from short-term depression was slowed. We then performed a physiological analysis of endbulb synapses from mice which, in addition to Piccolo deficiency, lacked one functional allele of the Bassoon gene. Analysis of the double-mutant endbulbs revealed an increase in release probability, while the synapses still exhibited the reduced RRP, and the impairment in SV replenishment was exacerbated. We propose additive roles of Piccolo and Bassoon in SV replenishment which in turn influences the organization and size of the RRP, and an additional role of Bassoon in regulation of release probability.

UNC-41/Stonin Functions with AP2 to Recycle Synaptic Vesicles in Caenorhabditis elegans

The recycling of synaptic vesicles requires the recovery of vesicle proteins and membrane. Members of the stonin protein family (Drosophila Stoned B, mammalian stonin 2) have been shown to link the synaptic vesicle protein synaptotagmin to the endocytic machinery. Here we characterize the unc-41 gene, which encodes the stonin ortholog in the nematode Caenorhabditis elegans. Transgenic expression of Drosophila stonedB rescues unc-41 mutant phenotypes, demonstrating that UNC-41 is a bona fide member of the stonin family. In unc-41 mutants, synaptotagmin is present in axons, but is mislocalized and diffuse. In contrast, UNC-41 is localized normally in synaptotagmin mutants, demonstrating a unidirectional relationship for localization. The phenotype of snt-1 unc-41 double mutants is stronger than snt-1 mutants, suggesting that UNC-41 may have additional, synaptotagmin-independent functions. We also show that unc-41 mutants have defects in synaptic vesicle membrane endocytosis, including a ∼50% reduction of vesicles in both acetylcholine and GABA motor neurons. These endocytic defects are similar to those observed in apm-2 mutants, which lack the µ2 subunit of the AP2 adaptor complex. However, no further reduction in synaptic vesicles was observed in unc-41 apm-2 double mutants, suggesting that UNC-41 acts in the same endocytic pathway as µ2 adaptin.

Staying Connected: Synapses in Alzheimer Disease

Careful dissection is continuing to reveal the complexity and importance of synapse alterations in Alzheimer disease (AD). Synaptic alterations are the best biological correlate to the extent of cognitive loss1,2 and have become the gold standard of meaningful biomarkers of AD. Not neurofibrillary tangles (NFT), senile plaques, or even neuronal loss show such strong statistical correlation with dementia.2–4 Connections are the hallmark of neurons and without functional synapses, neurons are in “solitary.” Recent findings showing that amyloid-β (Aβ) affects and accumulates in synapses5 and that amyloid-β protein precursor (AβPP) over-expressing mice show synaptic alterations6,7 have provided additional evidence that synapses are a critical readout of the biology of AD. In this issue, Gylys and co-workers8 provide an elegant and precise dissection of synaptic alterations in AD using synaptosomal preparations. The results are both consistent with the synaptic alteration hypothesis proposed by Terry and colleagues2–4 and refine it. In previous studies synapses have been quantitated by synaptophysin immunoreactivity,1,9 a protein belonging to a subclass of synaptic vesicles. However, when synapses were quantitated by precise ultrastructural morphometric/stereological approaches, reduction in synapses was much less striking than the reduction in synaptophysin.10 Furthermore, synapse alterations included major remodeling of morphology11 as noted in some of the earliest studies of AD.12 These established findings lead one to consider whether the alterations in synapses in AD are limited to global reduction in vesicles rather than synapse changes. Gylys and collaborators8 used a broad range of pre- and postsynaptic markers to precisely analyze synaptosomes by flow cytometry. They found that the synaptosomes in AD show no reduction in synaptophysin and only a slight reduction in the overall number of synaptosomes. To account for the undoubted major reduction in synaptophysin while synapse number and synaptic vesicle number are only slightly reduced, brings to mind the seminal findings of Suzuki and Terry13 who reported substantial axonal transport abnormalities in AD. Consistently, one finds vesicle pile-up in cell bodies similar to transport blockage models.14 Recent findings that microtubules are greatly reduced in AD15 and that AβPP/Aβ play important roles in vesicular transport16–20 further link Terry’s transport and Gylys’ synaptic hypotheses. Not answered is where the extrasynaptic synaptophysin is located. One of the most attractive possibilities is that many of the synaptophysin vesicles are located in axons; for that reason, Terry’s initial insight to focus on vesicles was particularly fruitful because it marked the reduction most specific to AD. Additional work either by cell fractionation studies, as Gylys and collaborators8 have so elegantly performed, or by immunoelectron microscopy is called for to dissect this issue more clearly.

Synaptic vesicle alterations in rod photoreceptors of synaptophysin-deficient mice

The abundance of the integral membrane protein synaptophysin in synaptic vesicles and its multiple possible functional contributions to transmitter exocytosis and synaptic vesicle formation stand in sharp contrast to the observed lack of defects in synaptophysin knockout mice. Assuming that deficiencies are compensated by the often coexpressed synaptophysin isoform synaptoporin, we now show that retinal rod photoreceptors, which do not synthesize synaptoporin either in wild-type or in knockout mice, are affected by the loss of synaptophysin. Multiple pale-appearing photoreceptors, as seen by electron microscopy, possess reduced cytoplasmic electron density, swollen mitochondria, an enlarged cell surface area, and, most importantly, a significantly reduced number of synaptic vesicles with an unusually bright interior. Quantification of the number of synaptic vesicles per unit area, not only in these, but also in all other rod terminals of knockout animals, reveals a considerable reduction in vesicles that is even more pronounced during the dark period, i.e., at times of highest synaptic activity. Moreover, activity-dependent reduction in synaptic vesicle diameter, typically occurring in wild-type mice, is not detected in knockout animals. The large number of clathrin-coated pits and vesicles in dark-adapted synaptophysin knockout mice is taken as an indication of compensatory usage of synaptophysin-independent pathway(s), and, conversely, in view of the overall reduction in the number of synaptic vesicles, as an indication for the presence of another synaptophysin-dependent synaptic vesicle recycling pathway. Our results provide in vivo evidence for the importance of the integral membrane protein synaptophysin for synaptic vesicle recycling and formation.

Influence of antimicrotubular drugs on the Golgi apparatus of stomach secretory mucoid cells and small intestine absorptive cells

The effects of vinblastine and colchicine on the Golgi apparatus of stomach surface mucoid and absorptive intestinal cells were compared by cytochemical analysis. The two epithelial cells were chosen because of their different specific functions in the formation of secretory granules, the production of lysosomes and the intensity of membrane traffic in the cytoplasm. For the analysis, adult mice were injected with 1 mg/100 g b.w. of vinblastine and 1 mg/100 g b.w. of colchicine. For the demonstration of cis and trans cisternae of the Golgi apparatus, prolonged osmification, thiamine pyrophosphatase and acid phosphatase activity identification were applied. After treatment with vinblastine or colchicine, polarity of stacks in the Golgi apparatus of surface mucoid cells is preserved although the number of cisternae with thiamine pyrophosphatase or acid phosphatase activity decreases. However, the Golgi apparatus of intestinal absorptive cells completely disintegrates and only a few separated cis or trans cisternae can be identified. The main effect seems to be a reduction of vesicles which can be cytochemically identified as parts of the Golgi apparatus and an accumulation of vesicles which probably originate from budding ER. Communication between the ER and the Golgi apparatus seems to be interrupted.

釀造の諸條件と前年度試釀酒の火持性について

Three different kinds of sake were brewed by using the same tane koji (seeds of Aspergilli) consisting of a mixture of equal amount of the two kinds of Aspergilli strains C and D used inthe sake brewed in the previous year, with the moto (yeast culture) prepared by (1) the quick method with the addition of lactic acid, (2) the saccharification method at a higher temperature (55-58°C.), and (3) the “yamahai” method at a lower temperature without lactic acid. Thus thequalities of sake were compared according to the brewing conditions with the same tane koji. They were different not only by the brewing conditions, but also by the properties of Aspergilli. It seemed that particularly the aroma and the durabilities in storage of sake were more influenced by the Aspergilli.Most of the strains of Aspergilli isolated from the de-koji (koji made from the cultivation of mixture of two kinds of Aspergilli strains C and D) were different from the original strains C and D. Evident degeneration took place in these new strains such as decrease in the productive power of acids, kojic acid, pigments, and the aroma and the reduction of vesicles. Particularly the properties of the new strains were rather similar to those of the strain D and it seemed that also the three kinds of sake were commonly alike to the sake brewed with the strain D in the previousyear.