Timothy E. Glass

Three-Input Logic Gates with Potential Applications for Neuronal Imaging

Convenient methods for the direct visualization of neurotransmitter trafficking would bolster investigations into the development of neurodegenerative diseases. Here, tunable fluorescent molecular logic gates with applications to neuronal imaging have been developed. The three-input AND molecular logic gates are based on the coumarin-3-aldehyde scaffold and designed to give a turn-on fluorescence response upon the corelease of glutamate and zinc from secretory vesicles via exocytosis. Spectroscopic studies reveal an 11-fold fluorescence enhancement under conditions mimicking exocytosis. Methylation of the scaffold was used to optimize the spectral profile of the sensors toward desired excitation wavelengths. A binding study that elucidates the sensor-analyte interactions is presented. These sensors serve as a proof-of-concept toward the direct imaging of neurotransmitters released upon exocytosis using fluorescent molecular logic gates.

Fluorescent sensors for diamines

A series of seven diamine sensors was prepared using dimers of a quinolone aldehyde chromophore. Binding to six different diamine guests was explored by a combination of NMR, absorption and fluorescence spectroscopy. It was shown that the dimeric sensors bound the diamine guests by formation of a bis-iminium ion which produced large changes in the fluorescence of the quinolone core. Issues of selectivity between guests are discussed.

Interpenetrated nano-capsule networks based on the alkali metal assisted assembly of p-carboxylatocalix[4]arene-O-methyl ether

Reaction of p-carboxylatocalix[4]arene-O-methyl ether with either rubidium or caesium hydroxide results in the formation of interpenetrated nano-capsule networks with the calixarene in the 1,3-alternate conformation.

Selective Catecholamine Recognition with NeuroSensor 521: A Fluorescent Sensor for the Visualization of Norepinephrine in Fixed and Live Cells

A method for the selective labeling and imaging of catecholamines in live and fixed secretory cells is reported. The method integrates a tailored approach using a novel fluorescence-based turn-on molecular sensor (NeuroSensor 521) that can exploit the high concentration of neurotransmitters and acidic environment within secretory vesicles for the selective recognition of norepinephrine and dopamine. The utility of the method was demonstrated by selectively labeling and imaging norepinephrine in secretory vesicles such that discrimination between norepinephrine- and epinephrine-enriched populations of chromaffin cells was observed. This method was validated in fixed cells by co-staining with an anti-PNMT antibody.

Molecular Tubes for Lipid Sensing: Tube Conformations Control Analyte Selectivity and Fluorescent Response

Two fluorescent sensors for lipids have been prepared and tested for detection of a number of hydrophobic compounds of varying shape and size. The data suggest that the two sensors have a different mode of fluorescent response. Yet, the two sensors are only different in the bridging group--one having a flexible amide and one having a rigid allyl bridge. The fluorescence data are explained based on a difference in conformation of the two sensors in aqueous solution.

ExoSensor 517: a dual-analyte fluorescent chemosensor for visualizing neurotransmitter exocytosis.

A dual-analyte fluorescent chemosensor (ExoSensor 517) for the direct visualization of neurotransmitters released upon exocytosis is presented. The sensor exploits the high concentration of neurotransmitters (e.g., glutamate, norepinephrine, and dopamine) and the pH gradient between the vesicle and synaptic cleft. The cooperative recognition elements require both binding and a change in environmental pH to afford a fluorescence response which makes ExoSensor 517 one of the first integrated molecular logic gates to be used for biological applications.

A novel method for the synthesis of 1,2-benzisoxazoline-3-one and its application to hypochlorite recognition

The reaction of salicylhydroxamic acid with hypochlorite produces 1,2-benzisoxazoline-3-one, a heterocycle that contains a fluorophore. As a result, this reaction was used as the basis for a new, selective and sensitive fluorescence system for the recognition of hypochlorite. The effectiveness of the method was demonstrated by its use to detect hypochlorite in a disinfectant solution as well as to image hypochlorite in cells.

Development of a Fluorescent Chemosensor for the Detection of Kynurenine

Kynurenine, a metabolite of tryptophan, is known to contribute to cancer progression when overproduced. A method for facile fluorescent sensing of kynurenine using sensor 1 has been developed. When bound at low pH, sensor 1 undergoes a very large bathochromic shift because kynurenine extends the conjugation of the fluorophore. This unusual mechanism of activation provides a 390-fold fluorescence enhancement that is very specific to kynurenine and a wavelength of fluorescence that extends into the red.

Synthesis of pyrimidine based metal ligands

Abstract A convenient synthesis of a pyrimidine based bis-tridentate metal ligand is presented. The pyrimidine core is constructed via cyclization of an amidine and a substituted propenone. A Stille coupling appends the terminal pyridyl units. The general methodology presented is amenable to functionality at several positions on the ligand framework.

General synthetic methods for the preparation of pinwheel receptors

Abstract The synthesis of bis-trityl diynes, used as cooperative fluorescent chemosensors, is described. The most convenient sequence for the preparation of the sensor framework was found to be intolerant of most functionality. Therefore, an unfunctionalized framework is constructed and recognition elements are selectively installed via electrophilic substitution on an electronically differentiated key intermediate. A divergent synthesis of a general class of chemical sensors emerges. This synthetic method is highlighted by its simplicity and modularity allowing for the introduction of various recognition elements and fluorophores.