Self-Assembly of Stimuli-Responsive Water-Soluble [60]Fullerene End-Capped Ampholytic Block Copolymer

Well-defined, water-soluble, pH and temperature stimuli-responsive [60]fullerene (C₆₀) containing ampholytic block copolymer of poly((methacrylic acid)-block-(2-(dimethylamino)ethyl methacrylate))-block–C₆₀ (P(MAA-b-DMAEMA)-b-C₆₀) was synthesized by the atom transfer radical polymerization (ATRP) technique. The self-assembly behaviour of the C₆₀ containing polyampholyte in aqueous solution was characterized by dynamic light scattering (DLS), and transmission electron microscopy. This amphiphilic mono-C₆₀ end-capped block copolymer shows enhanced solubility in aqueous medium at room and elevated temperatures and at low and high pH but phase-separates at intermediate pH of between 5.4 and 8.8. The self assembly of the copolymer is different from that of P(MAA-b-DMAEMA). Examination of the association behavior using DLS revealed the co-existence of unimers and aggregates at low pH at all temperatures studied, with the association being driven by the balance of hydrophobic and electrostatic interactions. Unimers and aggregates of different microstructures are also observed at high pH and at temperatures below the lower critical solution temperature (LCST) of PDMAEMA. At high pH and at temperatures above the LCST of PDMAEMA, the formation of micelles and aggregates co-existing in solution is driven by the combination of hydrophobic, electrostatic, and charge-transfer interactions.

Stimuli-responsive Novel Amphiphilic Polymers for Chemical and Biomedical Applications

Amphiphilic polymers are a class of polymers that self-assemble into different types of microstructure, depending on the solvent environment and external stimuli. Self assembly structures can exist in many different forms, such as spherical micelles, rod-like micelles, bi-layers, vesicles, bi-continuous structure etc. Most biological systems are basically comprised of many of these organised structures arranged in an intelligent manner, which impart functions and life to the system. We have adopted the atom transfer radical polymerization (ATRP) technique to synthesize various types of block copolymer systems that self-assemble into different microstructure when subject to an external stimuli, such as pH or temperature. The systems that we have studied are: (1) pH responsive fullerene (C60) containing poly(methacrylic acid) (PMAA-b-C60); (2) pH and temperature responsive fullerene containing poly[2-(dimethylamino)ethyl methacrylate] (C₆₀-b-PDMAEMA); (3) other responsive water-soluble fullerene systems. By varying temperature, pH and salt concentration, different types microstructure can be produced. In the presence of inorganic salts, fractal patterns at nano- to microscopic dimension were observed for negatively charged PMAA-b-C60, while such structure was not observed for positively charged PDMAEMA-b-C60. We demonstrated that negatively charged fullerene containing polymeric systems can serve as excellent nano-templates for the controlled growth of inorganic crystals at the nano- to micrometer length scale and the possible mechanism was proposed. The physical properties and the characteristics of their self-assembly properties will be discussed, and their implications to chemical and biomedical applications will be highlighted.

Targeted Stimuli-Responsive Dextran Conjugates for Doxorubicin Delivery to Hepatocytes

A targeted, stimuli-responsive, polymeric drug delivery vehicle is being developed in our lab to help alleviate severe side-effects caused by narrow therapeutic window drugs. Targeting specific cell types or organs via proteins, specifically, lectin-mediated targeting holds potential due to the high specificity and affinity of receptor-ligand interactions, rapid internalization, and relative ease of processing. Dextran, a commercially available, biodegradable polymer has been conjugated to doxorubicin and galactosamine to target hepatocytes in a three-step, one-pot synthesis. The loading of doxorubicin and galactose on the conjugates was determined by absorbance at 485 nm and elemental analysis, respectively. Conjugation efficiency based on the amount loaded of each reactant varies from 20% to 50% for doxorubicin and from 2% to 20% for galactosamine. Doxorubicin has also been attached to dextran through an acid-labile hydrazide bond. Doxorubicin acts by intercalating with DNA in the nuclei of cells. The fluorescence of doxorubicin is quenched when it binds to DNA. This allows a fluorescence-based cell-free assay to evaluate the efficacy of the polymer conjugates where we measure the fluorescence of doxorubicin and the conjugates in increasing concentrations of calf thymus DNA. Fluorescence quenching indicates that our conjugates can bind to DNA. The degree of binding increases with polymer molecular weight and substitution of doxorubicin. In cell culture experiments with hepatocytes, the relative uptake of polymer conjugates was evaluated using flow cytometry, and the killing efficiency was determined using the MTT cell proliferation assay. We have found that conjugate uptake is much lower than that of free doxorubicin. Lower uptake of conjugates may increase the maximum dose of drug tolerated by the body. Also, non-galactosylated conjugate uptake is lower than that of the galactosylated conjugate. Microscopy indicates that doxorubicin localizes almost exclusively at the nucleus, whereas the conjugates are present throughout the cell. Doxorubicin linked to dextran through a hydrazide bond was used to achieve improved killing efficiency. Following uptake, the doxorubicin dissociates from the polymer in an endosomal compartment and diffuses to the nucleus. The LC₅₀ of covalently linked doxorubicin is 7.4 μg/mL, whereas that of hydrazide linked doxorubicin is 4.4 μg/mL.

Disparity Gradients and Depth Scaling

The binocular perception of shape and depth relations between objects can change considerably if the viewing direction is changed only by a small angle. We explored this effect psychophysically and found a strong depth reduction effect for large disparity gradients. The effect is found to be strongest for horizontally oriented stimuli, and stronger for line stimuli than for points. This depth scaling effect is discussed in a computational framework of stereo based on a Baysian approach which allows integration of information from different types of matching primitives weighted according to their robustness.

Parallel Computation of Vernier Offsets, Curvature and Chevrons in Humans

A vernier offset is detected at once among straight lines, and reaction times are almost independent of the number of simultaneously presented stimuli (distractors), indicating parallel processing of vernier offsets. Reaction times for identifying a vernier offset to one side among verniers offset to the opposite side increase with the number of distractors, indicating serial processing. Even deviations below a photoreceptor diameter can be detected at once. The visual system thus attains positional accuracy below the photoreceptor diameter simultaneously at different positions. I conclude that deviation from straightness, or change of orientation, is detected in parallel over the visual field. Discontinuities or gradients in orientation may represent an elementary feature of vision.

Viewpoint-Specific Representations in Three-Dimensional Object Recognition

We report a series of psychophysical experiments that explore different aspects of the problem of object representation and recognition in human vision. Contrary to the paradigmatic view which holds that the representations are three-dimensional and object-centered, the results consistently support the notion of view-specific representations that include at most partial depth information. In simulated experiments that involved the same stimuli shown to the human subjects, computational models built around two-dimensional multiple-view representations replicated our main psychophysical results, including patterns of generalization errors and the time course of perceptual learning.

Spatial Reference Frames for Object Recognition: Tuning for Rotations in Depth

The inferior temporal cortex (IT) of monkeys is thought to play an essential role in visual object recognition. Inferotemporal neurons are known to respond to complex visual stimuli, including patterns like faces, hands, or other body parts. What is the role of such neurons in object recognition? The present study examines this question in combined psychophysical and electrophysiological experiments, in which monkeys learned to classify and recognize novel visual 3D objects. A population of neurons in IT were found to respond selectively to such objects that the monkeys had recently learned to recognize. A large majority of these cells discharged maximally for one view of the object, while their response fell off gradually as the object was rotated away from the neuron"s preferred view. Most neurons exhibited orientation-dependent responses also during view-plane rotations. Some neurons were found tuned around two views of the same object, while a very small number of cells responded in a view- invariant manner. For five different objects that were extensively used during the training of the animals, and for which behavioral performance became view-independent, multiple cells were found that were tuned around different views of the same object. No selective responses were ever encountered for views that the animal systematically failed to recognize. The results of our experiments suggest that neurons in this area can develop a complex receptive field organization as a consequence of extensive training in the discrimination and recognition of objects. Simple geometric features did not appear to account for the neurons" selective responses. These findings support the idea that a population of neurons -- each tuned to a different object aspect, and each showing a certain degree of invariance to image transformations -- may, as an assembly, encode complex 3D objects. In such a system, several neurons may be active for any given vantage point, with a single unit acting like a blurred template for a limited neighborhood of a single view.

The Role of Attention in Binocular Rivalry as Revealed Through Optokinetic Nystagmus

When stimuli presented to the two eyes differ considerably, stable binocular fusion fails, and the subjective percept alternates between the two monocular images, a phenomenon known as binocular rivalry. The influence of attention over this perceptual switching has long been studied, and although there is evidence that attention can affect the alternation rate, its role in the overall dynamics of the rivalry process remains unclear. The present study investigated the relationship between the attention paid to the rivalry stimulus, and the dynamics of the perceptual alternations. Specifically, the temporal course of binocular rivalry was studied as the subjects performed difficult nonvisual and visual concurrent tasks, directing their attention away from the rivalry stimulus. Periods of complete perceptual dominance were compared for the attended condition, where the subjects reported perceptual changes, and the unattended condition, where one of the simultaneous tasks was performed. During both the attended and unattended conditions, phases of rivalry dominance were obtained by analyzing the subject"s optokinetic nystagmus recorded by an electrooculogram, where the polarity of the nystagmus served as an objective indicator of the perceived direction of motion. In all cases, the presence of a difficult concurrent task had little or no effect on the statistics of the alternations, as judged by two classic tests of rivalry, although the overall alternation rate showed a small but significant increase with the concurrent task. It is concluded that the statistical patterns of rivalry alternations are not governed by attentional shifts or decision-making on the part of the subject.

How do Humans Determine Reflectance Properties under Unknown Illumination?

Under normal viewing conditions, humans find it easy to distinguish between objects made out of different materials such as plastic, metal, or paper. Untextured materials such as these have different surface reflectance properties, including lightness and gloss. With single isolated images and unknown illumination conditions, the task of estimating surface reflectance is highly underconstrained, because many combinations of reflection and illumination are consistent with a given image. In order to work out how humans estimate surface reflectance properties, we asked subjects to match the appearance of isolated spheres taken out of their original contexts. We found that subjects were able to perform the task accurately and reliably without contextual information to specify the illumination. The spheres were rendered under a variety of artificial illuminations, such as a single point light source, and a number of photographically-captured real-world illuminations from both indoor and outdoor scenes. Subjects performed more accurately for stimuli viewed under real-world patterns of illumination than under artificial illuminations, suggesting that subjects use stored assumptions about the regularities of real-world illuminations to solve the ill-posed problem.