N-terminal signals in the SNX-BAR paralogs Vps5 and Vin1 guide endosomal coat complex formation.

Endosomal coats incorporate membrane-binding subunits such as sorting nexin (SNX) proteins. The SNX-BAR paralogs Vin1 and Vps5 are respective subunits of the endosomal VINE and retromer complexes whose dimerizing BAR domains are required for complex assembly and membrane association. However, a degree of promiscuity is predicted for yeast BAR-BAR pairings, and recent work has implicated the unstructured N-terminal domains of Vin1 and Vps5 in coat formation.

The VINE complex is an endosomal VPS9-domain GEF and SNX-BAR coat.

Membrane trafficking pathways perform important roles in establishing and maintaining the endosomal network. Retrograde protein sorting from the endosome is promoted by conserved SNX-BAR-containing coat complexes including retromer which enrich cargo at tubular microdomains and generate transport carriers. In metazoans, retromer cooperates with VARP, a conserved VPS9-domain GEF, to direct an endosomal recycling pathway.

You can go your own way: SNX-BAR coat complexes direct traffic at late endosomes.

The endolysosomal network consists of highly dynamic membrane-bound compartments that control subcellular degradative and recycling processes. A conserved family of endosomal coat complexes known as SNX-BARs drive the formation of tubular membrane transport carriers for cargo retrieval. Whereas SNX1-related SNX-BARs were previously thought to rely on their association with the retromer complex to recognize cargo, recent work shows this class of SNX-BARs can directly bind and deliver cargo.

Dose mediates the protracted effects of adolescent THC exposure on reward and stress reactivity in males relevant to perturbation of the basolateral amygdala transcriptome.

Despite the belief that cannabis is relatively harmless, exposure during adolescence is associated with increased risk of developing several psychopathologies in adulthood. In addition to the high levels of use amongst teenagers, the potency of ∆-9-tetrahydrocannabinol (THC) has increased more than fourfold compared to even twenty years ago, and it is unclear whether potency influences the presentation of THC-induced behaviors. Expanded knowledge about the impact of adolescent THC exposure, especially high dose, is important to delineating neural networks and molecular mechanisms underlying psychiatric risk.

Temperature-sensitive recombinant subtilisin protease variants that efficiently degrade molecular biology enzymes.

We used error-prone PCR to generate mutations in a subtilisin protease-encoding gene, and screened for recombinants that expressed temperature-sensitive (TS) variants. From the dozens of mutations that we detected in the recombinant genes we found that those mutations that affected aspartate-75 had the most profound effect on temperature stability. We thus focused our analysis on two variants of subtilisin C, the more heat-sensitive variant 24 (V24), with amino acid changes D75G, L234M and Q274P; and variant 25 (V25), with a single amino acid change, D75A.

Cell-type-specific regulation of neuronal intrinsic excitability by macroautophagy.

The basal ganglia are a group of subcortical nuclei that contribute to action selection and reinforcement learning. The principal neurons of the striatum, spiny projection neurons of the direct (dSPN) and indirect (iSPN) pathways, maintain low intrinsic excitability, requiring convergent excitatory inputs to fire. Here, we examined the role of autophagy in mouse SPN physiology and animal behavior by generating conditional knockouts of Atg7 in either dSPNs or iSPNs.

Dopamine Triggers the Maturation of Striatal Spiny Projection Neuron Excitability during a Critical Period.

Neural circuits are formed and refined during childhood, including via critical changes in neuronal excitability. Here, we investigated the ontogeny of striatal intrinsic excitability. We found that dopamine neurotransmission increases from the first to the third postnatal week in mice and precedes the reduction in spiny projection neuron (SPN) intrinsic excitability during the fourth postnatal week.

α-Synuclein-Dependent Calcium Entry Underlies Differential Sensitivity of Cultured SN and VTA Dopaminergic Neurons to a Parkinsonian Neurotoxin.

Parkinson's disease (PD) is a debilitating neurodegenerative disease characterized by a loss of dopaminergic neurons in the substantia nigra (SN). Although mitochondrial dysfunction and dysregulated α-synuclein (aSyn) expression are postulated to play a role in PD pathogenesis, it is still debated why neurons of the SN are targeted while neighboring dopaminergic neurons of the ventral tegmental area (VTA) are spared. Using electrochemical and imaging approaches, we investigated metabolic changes in cultured primary mouse midbrain dopaminergic neurons exposed to a parkinsonian neurotoxin, 1-methyl-4-phenylpyridinium (MPP).

Lectin-directed enzyme activated prodrug therapy (LEAPT): Synthesis and evaluation of rhamnose-capped prodrugs.

The lectin-directed enzyme activated prodrug therapy (LEAPT) bipartite drug delivery system utilizes glycosylated enzyme, localized according to its sugar pattern, and capped prodrugs released by that enzyme. In this way, the sugar coat of a synthetic enzyme determines the site of release of a given drug. Here, prodrugs of doxorubicin and 5-fluorouracil capped by the nonmammalian l-rhamnosyl sugar unit have been efficiently synthesized and evaluated for use in the LEAPT system.

Esophageal transit of the weekly film-coated risedronate (Actonel) placebo tablet in subjects with Kyphosis.

Risedronate sodium is a pyridinyl bisphosphonate of proven effectiveness for the treatment and prevention of osteoporosis and Paget's disease of the bone. The aim of this study was to compare the esophageal transit and gastric emptying of the placebo film-coated risedronate tablet when taken with 50 or 120 mL of water in subjects with Kyphosis. A total of 23 patients with radiologically documented osteoporosis participated in a single-center, open-label, crossover gamma scintigraphy study.

Hydrophilic and lipophilic radiopharmaceuticals as tracers in pharmaceutical development: in vitro--in vivo studies.

Background: Scintigraphic studies have been performed to assess the release, both in vitro and in vivo, of radiotracers from tablet formulations. Four different tracers with differing physicochemical characteristics have been evaluated to assess their suitability as models for drug delivery.

Methods: In-vitro disintegration and dissolution studies have been performed at pH 1, 4 and 7.

Preparation of Tc-99m-macroaggregated albumin from recombinant human albumin for lung perfusion imaging.

Human serum albumin (HSA) extracted from pooled blood taken from human donors is used in the production of (99m)Tc-labelled macroaggregated albumin (MAA) for lung perfusion imaging. However, concerns for the safety of blood-derived products due to potential contamination by infective agents (e.g.

LEAPT: lectin-directed enzyme-activated prodrug therapy.

Targeted drug delivery to selected sites allows reduced toxicity, enhanced efficiency and interchangeable target potential [Langer, R. (2001) Science 293, 58-59 and Molema, G. & Meijer, D.

Radionuclide imaging in drug development.

Radioactive tracers have made an immense contribution to the understanding of human physiology and pathology. At the start of the 21st century nuclear imaging has emerged as the main metabolic imaging modality which is of growing importance in drug development and clinical pharmacology. Using techniques adapted from those undertaken in clinical radiopharmacy and nuclear medicine facilities drug molecules and carrier systems may be radiolabelled and their release, biodistribution and uptake may be visualized in human subjects.

Usage of radiopharmaceuticals in the development of pharmaceutical drug delivery systems: validation of [99mTc]DTPA and [99mTc]ECD.

Tablets containing drugs of different lipophilicity, ranitidine and cinarizine, and placebo were prepared and their in vitro behaviour was studied by dissolution and disintegration tests. [(99m)Tc]Diethylenetriamine-pentaacetic acid ([(99m)Tc]DTPA) and [(99m)Tc]ethyl cysteinate dimer ([(99m)Tc]ECD) were used as tracers of the process. Both of them were added to tablets during wet granulation.

Rhenium-188 and copper-67 radiopharmaceuticals for the treatment of bladder cancer.

The favourable nuclear properties of copper-67 and rhenium-188 for therapeutic application are described, together with methods for the chemical synthesis of a number of derivatives. Survival from invasive bladder cancer has changed little over the past 20 years. The intravesicular administration of Cu-67 or Re-188 radiopharmaceuticals in the treatment of bladder cancer offers some promise for improvement in this situation.

Evaluation of Pulsincap to provide regional delivery of dofetilide to the human GI tract.

Pulsincap formulations designed to deliver a dose of drug following a 5-h delay were prepared to evaluate the capability of the formulation to deliver dofetilide to the lower gastrointestinal (GI) tract. By the expected 5-h release time, the preparations were well dispersed throughout the GI tract, from stomach to colon. Plasma analysis permitted drug absorption to be determined as a function of GI tract site of release.

The use of scintigraphy to demonstrate the rapid esophageal transit of the oval film-coated placebo risedronate tablet compared to a round uncoated placebo tablet when administered with minimal volumes of water.

As our population ages, and the consumption of pharmaceutical products rises, the incidence of solid oral dosage forms lodging in the esophagus is likely to increase and may be formulation dependent. The aim of this study was to compare the esophageal transit of the commercial film-coated risedronate tablet and a round uncoated tablet resembling the alendronate 10 mg tablet which is reported to cause esophagitis if ingested with little to no water. Water volumes of 30 ml and 50 ml were selected as these volumes can detect formulations prone to esophageal adhesion and a habits and practice study showed that these volumes are within the range preferred by women (7-385 ml).

The effect of the nasal cycle on mucociliary clearance.

The nasal cycle is a well-recognised physiological phenomenon where each side of the nose alternates through phases of congestion and decongestion. Although many physiological properties of the nose alternate with the nasal cycle whether this has any effect on the nasal mucociliary clearance is less clear. As the nose is a potential site for the administration of pharmaceuticals, it is essential that any factors that could affect clearance (and hence absorption) are identified.

Oesophageal transit, disintegration and gastric emptying of a film-coated risedronate placebo tablet in gastro-oesophageal reflux disease and normal control subjects.

Background: Risedronate sodium is a pyridinyl bisphosphonate, proven effective for the treatment and prevention of postmenopausal osteoporosis and glucocorticoid-induced osteoporosis and Paget's disease of the bone.

Aim: To compare the oesophageal transit, disintegration and gastric emptying of the commercial film-coated risedronate tablet in subjects with gastro-oesophageal reflux disease (GERD) and normal control subjects.

Methods: A total of 30 subjects, 15 patients with GERD and 15 age- and sex-matched, normal control subjects, participated in a single-centre, open-label, comparative gamma scintigraphy study.

Imaging for staging bladder cancer: a clinical study of intravenous 111indium-labelled anti-MUC1 mucin monoclonal antibody C595.

Objective: To investigate the clinical application of an 111In-labelled anti-MUC1 mucin monoclonal antibody (mAb) imaging for staging invasive bladder cancer.

Patients And Methods: Indirect immunohistochemistry was used to confirm the expression of the MUC1 target antigen by metastatic tumours. Twelve patients with bladder cancer (two with superficial and 10 with locally invasive/metastatic disease) underwent planar gamma-scintigraphy 48 h after an intravenous injection with 111In-labelled anti-MUC1 mucin mAb C595.

Night-time quiescence and morning activation in the human colon: effect on transit of dispersed and large single unit formulations.

Objectives: Controlling the delivery of drugs to different regions of the colon remains an elusive goal. The aim of this study was to define the diurnal variation in colonic transit and show how this influences the colonic distribution and residence time of different formulations given either in the morning or evening.

Methods: Colonic transit of small particulates and a large capsule was measured during nocturnal sleep and daytime wakefulness.

Targeting superficial bladder cancer by the intravesical administration of copper-67-labeled anti-MUC1 mucin monoclonal antibody C595.

Purpose: More effective intravesical agents are required to limit the recurrence and progression of superficial bladder cancer. This study assessed the ability of copper-67 ((67)Cu)-C595 murine antimucin monoclonal antibody to bind selectively to superficial bladder tumors when administered intravesically, with a view to its development for therapy.

Patients And Methods: Approximately 20 MBq of (67)Cu-C595 monoclonal antibody was administered intravesically to 16 patients with a clinical indication of superficial bladder cancer.

Dry powder dosing in liquid vehicles: ocular tolerance and scintigraphic evaluation of a perfluorocarbon suspension.

The ocular tolerance and precorneal disposition of 99mTc-labelled sterile carbon-perfluorodecalin (PFD) and carbon-aqueous suspensions were examined in a cohort of healthy volunteers. Formulations were prepared in PFD or saline using charcoal particles, radiolabelled with [99mTc]diethylenetriaminepentaacetic acid (DTPA) under GMP conditions. Colloidal silicon dioxide was used as a suspending agent.