The t-SNAREs syntaxin 1 and SNAP-25 are present on organelles that participate in synaptic vesicle recycling

Syntaxin 1 and synaptosome-associated protein of 25 kD (SNAP-25) are neuronal plasmalemma proteins that appear to be essential for exocytosis of synaptic vesicles (SVs). Both proteins form a complex with synaptobrevin, an intrinsic membrane protein of SVs. This binding is thought to be responsible for vesicle docking and apparently precedes membrane fusion. According to the current concept, syntaxin 1 and SNAP-25 are members of larger protein families, collectively designated as target-SNAP receptors (t-SNAREs), whose specific localization to subcellular membranes define where transport vesicles bind and fuse. Here we demonstrate that major pools of syntaxin 1 and SNAP-25 recycle with SVs. Both proteins cofractionate with SVs and clathrin-coated vesicles upon subcellular fractionation. Using recombinant proteins as standards for quantitation, we found that syntaxin 1 and SNAP-25 each comprise approximately 3% of the total protein in highly purified SVs. Thus, both proteins are significant components of SVs although less abundant than synaptobrevin (8.7% of the total protein). Immunoisolation of vesicles using synaptophysin and syntaxin specific antibodies revealed that most SVs contain syntaxin 1. The widespread distribution of both syntaxin 1 and SNAP-25 on SVs was further confirmed by immunogold electron microscopy. Botulinum neurotoxin C1, a toxin that blocks exocytosis by proteolyzing syntaxin 1, preferentially cleaves vesicular syntaxin 1. We conclude that t-SNAREs participate in SV recycling in what may be functionally distinct forms.

Imaging exocytosis and recycling of synaptic-like microvesicles in astrocytes.

Optical imaging techniques are well suited for following the dynamics of physiological processes in living cells. Total internal reflection fluorescence (TIRF) microscopy based on evanescent wave illumination (EWi) allows spectacular, real-time visualization of individual vesicle movements, fusions, and retrievals at the cell surface (i.e., within 100 nm of the plasma membrane). TIRF microscopy is an ideal approach for studying the properties of exocytosis and recycling in cultured astrocytes, particularly because these cells have a rather flat surface and contain secretory vesicles with sparse distribution. Among all populations of secretory vesicles, we focus here on synaptic-like microvesicles (SLMVs). We illustrate how TIRF microscopy using EWi is useful to study exocytosis and recycling of SLMVs at the single-vesicle level and, when combined with epifluorescence illumination (EPIi), can provide detailed information on the kinetics of exocytosis, endocytosis, and re-acidification at the whole-cell level.

Field Evaluation of Cold In-Place Recycling of Asphalt Concrete, HR-303, 1993

The average thickness of the existing asphalt cement concrete (ACC) along route E66 in Tama County was 156 mm (6.13 in.). The rehabilitation strategy called for widening the base using the top 75 mm (3 in.) of the existing ACC by a recycling process involving cold milling and mixing with additional emulsion/rejuvenator. The material was then placed into a widening trench and compacted to match the level of the milled surface. The project had the following results: (1) Cold recycled ACC pavement provided adequate pavement structure for a low volume road; (2) Premature cracking of the ACC in the widened pavement area was caused by compaction of the mix over a saturated subgrade; and (3) Considerably less transverse and longitudinal cracking was observed with 75 mm (3 in.) of cold recycled ACC and a 50 mm (2 in.) hot mix ACC overlay than with a conventional hot mix overlay with no cold recycling. More research should be done on efficient construction procedures and incorporating longer test sections for proper evaluation.

Asphalt Cement Concrete Pavement Recycling Cass and Montgomery Counties, HR-1018, 1986

This demonstration project consisted of three adjacent highway resurfacing projects using asphalt cement concrete removed from an Interstate highway which had become severely rutted. The salvaged asphaltic concrete was later crushed and hauled to a plant site where it was combined with virgin materials to resurface the three projects. Only two of the projects were used for performance evaluation as the third project was in an interchange area including ramps and was otherwise too short. It was concluded that recycling was cost effective and a high quality surface can be constructed using recycled asphalt cement concrete.

Recycling of Portland Cement Concrete Roads in Iowa, MLR-77-6, 1977

We are depleting the once seemingly endless supply of aggregate available for concrete paving in Iowa. At the present time, some parts of our state do not have locally available aggregates of acceptable quality for portland cement concrete paving. This necessitates lengthy truck and rail hauls which frequently more than doubles the price of aggregate. In some parts of the state, the only coarse aggregates available locally are "d-cracking" in nature. Iowa's recycling projects were devised to alleviate the shortage of aggregates wherever they were found to have an economic advantage. We completed our first recycling project in 1976 on a 1.4 project in Lyon county. The data collected in this project was used to schedule two additional projects in 1977. The larger of these two projects is located in Page and Taylor county on Highway #2 and is approximately 15 miles in length. This material is to be crushed and re-used in the concrete paving, it is to be reconstructed on approximately the same alignment. The second project is part of the construction of Interstate I-680 north of council Bluffs where an existing 24 foot portland cement concrete roadway is to be recycled and used as the aggregate in the slip form econocrete subbase and the portland cement concrete shoulders.

Evaluation of Air Pollution Control Devices for Asphalt Pavement Recycling Operations, HR-188, Progress Report, 1977

This report discusses the asphalt pavement recycling project designated Project HR-188 in Kossuth County, Iowa. Specific objectives were: (a) to determine the effectiveness of drum mixing plant modifications designed to control air pollution within limits specified by the Iowa Department of Environmental Quality; (b) to assess the impact of varying the proportions of recycled and virgin aggregates, (c) to assess the impact of varying the production rate of the plant, and (d) to assess the impact of varying the mixing temperature. The discussion includes information on the proposed use of research funds, project location and description, the project planning conference, plan development, bid letting, asphalt plant configuration, actual plant operation, why this method is successful, probable process limitations, pollution results, recycled pavement test results, and the cost of virgin vs. recycled asphalt pavements.

Iowa D.O.T's Experience with Recycling Portland Cement Pavement and Asphalt Cement Pavement, MLR-77-4, 1977

This report provides details of IADOT's experience removing and crushing asphaltic concrete and portland cement concrete for recycling. The recycled material was used on interstate highways for the subbase and shoulders. The major problem IADOT encountered on this project was the removal of reinforcing steel from the broken concrete. The contractor used hydraulic powered shears to clip off all protruding steel during the removal and loading of the concrete on the grade. This project took place in 1977.

Recycling of Cass County I-80 Asphalt Concrete, Progress Report, HR-1011, 1977

Approximately 40,000 tons of slightly damaged asphalt concrete has been removed from Interstate 80 in Cass and stockpiled. Laboratory tests had indicated that this material had considerable value when upgraded with new aggregate and asphalt cement. This report documents the procedures used and results obtained on an experimental recycling project. It was demonstrated that present drum mixing-recycling equipment and procedures can be used to utilize this material with satisfactory results. Laboratory analyses of material components and mixtures were performed; these analyses indicate mixture can be produced that is uniform, stable, and very closely resembles mixture produced with all new material. Follow~up evaluations will be made to determine the effects of traffic and environment. Preliminary data indicate that plans should be made to incorporate the stockpiled material in projects near the stockpile site.

Recycling of Asphalt Concrete From I-80 in Cass County, HR-1011, 1978

Approximately 40,000 tons of deteriorated asphalt concrete has been removed from Interstate 80 in Cass County and stockpiled. Laboratory tests indicate that this material has considerable value when upgraded with new aggregate and asphalt cement. This report documents the procedures used and results obtained on an experimental recycling project. It was demonstrated that present drum mixing-recycling equipment and procedures can be used to utilize this material with satisfactory results. Laboratory analyses of material components and mixtures were performed; these analyses indicate mixture can be produced that is uniform, stable, and very closely resembles mixture produced with all virgin material. A 1700 foot long test section was constructed on US 169 in Kossuth County wherein salvaged asphalt concrete from I-80 in Cass County was utilized. The salvaged mix was blended with virgin aggregate and recycled through a modified drum mixing plant, the reprocessed mixture was satisfactorily placed 1 1/2 inches thick as a resurfacing course on an old PCC pavement. An inspection of the test section was made in December of 1978 to evaluate the performance after one full year of service. There was no evidence of rutting or shoving from traffic. The surface does, however, have a very dry and somewhat ravelled appearance. This can be related to a low asphalt content in the mix and some temperature control problems which were difficult to get fully corrected on such a short project and with a short supply of readily available materials.

Manual of Laboratory Mix Design Procedure for Cold In-Place Recycling using Foamed Asphalt (CIR-foam), TR-474, 2007

The objective is to determine the optimum percentage of water needed to produce the best foam properties for a given asphalt binder. The optimum water content is determined by achieving the maximum expansion ratio and half-life of the foamed asphalt. Expansion ratio is defined as the maximum volume over its original volume and half-life is defined as the time in seconds for foam to become a half of its maximum volume.

Examination of Curing Criteria for Cold In-Place Recycling, TR-553, 2008

Cold In-Place Recycling (CIR) has been used widely in rehabilitating the rural highways because it improves a long-term pavement performance. A CIR layer is normally covered by a hot mix asphalt (HMA) overlay in order to protect it from water ingress and traffic abrasion and obtain the required pavement structure and texture. Curing is the term currently used for the period of time that a CIR layer should remain exposed to drying conditions before an HMA overlay is placed. The industry standard for curing time is 10 days to 14 days or a maximum moisture content of 1.5 percent, which appear to be very conservative. When the exposed CIR layer is required to carry traffic for many weeks before the wearing surface is placed, it increases the risk of a premature failure in both CIR layer and overlay. This study was performed to explore technically sound ways to identify minimum in-place CIR properties necessary to permit placement of the HMA overlay. To represent the curing process of CIR pavement in the field construction, three different laboratory curing procedures were examined: 1) uncovered, 2) semi-covered and 3) covered specimens. The indirect tensile strength of specimens in all three curing conditions did not increase during an early stage of curing but increased during a later stage of curing usually when the moisture content falls below 1.5%. Dynamic modulus and flow number increased as curing time increased and moisture contents decreased. For the same curing time, CIR-foam specimens exhibited the higher tensile strength and less moisture content than CIR-emulsion. The laboratory test results concluded that the method of curing temperature and length of the curing period significantly affect the properties of the CIR mixtures. The moisture loss index was developed to predict the moisture condition in the field and, in the future, this index be calibrated with the measurements of temperature and moisture of a CIR layer in the field.

Examination of Curing Criteria for Cold In-Place Recycling Phase 2: Measuring Temperature, Moisture, Deflection and Distress from CIR Test Section, TR-590, 2009

The previous research performed laboratory experiments to measure the impacts of the curing on the indirect tensile strength of both CIR-foam and CIR-emulsion mixtures. However, a fundamental question was raised during the previous research regarding a relationship between the field moisture content and the laboratory moisture content. Therefore, during this research, both temperature and moisture conditions were measured in the field by embedding the sensors at a midpoint and a bottom of the CIR layer. The main objectives of the research are to: (1) measure the moisture levels throughout a CIR layer and (2) develop a moisture loss index to determine the optimum curing time of CIR layer before HMA overlay. To develop a set of moisture loss indices, the moisture contents and temperatures of CIR-foam and CIR-emulsion layers were monitored for five months. Based on the limited field experiment, the following conclusions are derived: 1. The moisture content of the CIR layer can be monitored accurately using the capacitance type moisture sensor. 2. The moisture loss index for CIR layers is a viable tool in determining the optimum timing for an overlay without measuring actual moisture contents. 3. The modulus back-calculated based on the deflection measured by FWD seemed to be in a good agreement with the stiffness measured by geo-gauge. 4. The geo-gauge should be considered for measuring the stiffness of CIR layer that can be used to determine the timing of an overlay. 5. The stiffness of CIR-foam layer increased as a curing time increased and it seemed to be more influenced by a temperature than moisture content. The developed sets of moisture loss indices based on the field measurements will help pavement engineers determine an optimum timing of an overlay without continually measuring moisture conditions in the field using a nuclear gauge.