A simulation of the neural action potential under the influence of acetylcholinesterase inhibitors modeled in the neuromuscular junction

The rise of terrorism has created an interest in better ways to detect when humans are exposed to neurotoxins, especially nerve gases developed for military use, most of which are acetylcholinesterase inhibitors. Many current methods of detection are based on mass spectrometry, a method that is cumbersome and not particularly robust when used as an early warning method. The detection of acetylcholinesterase inhibitors would benefit from a combined model of the processes occurring in the neuromuscular junction between the presynaptic action potential and the motor end-plate action potential that includes the kinetics of acetylcholine and acetylcholinesterase in the synaptic cleft. The ability to simulate the impact of different amounts of neurotoxin on the physiological processes needed for the generation of an action potential and subsequent muscle contraction would allow better estimates on the physiological toxicity of a nerve agent and its impact on an organism. The goal of this research was to assist the future development of a unified model and simulation of the chemical kinetics and electrical dynamics occurring in the synaptic cleft during acetylcholinesterase inhibition by neurotoxins. The first objective towards the goal of this research was to develop an accurate and useful model of the kinetics of acetylcholinesterase inhibition that can be simulated and coupled to the voltage and current signals generated by a neuron. A one dimensional diffusion model was used which took advantage of geometric symmetry to focus on the dominant transport effects. It will be shown that the simulation herein can reproduce the work of earlier research in depicting the time and spatial course of a normal action potential, and the time and spatial course of action potentials influenced by different degrees of acetylcholinesterase inhibition. This is the first simulation to achieve a model of acetylcholinesterase inhibition during the diffusion of a neuro-toxic inhibitor into the neuromuscular junction, and show the altered subsequent action potentials. Also illustrated will be how this simulation could detect the time and space dynamics of moving concentration gradients in the neuromuscular junction under suitable conditions. In addition, an in vivo simulation of inhibited acetylcholinesterase being returned to the active state through the kinetics of pralidoxime therapy will be shown. The mathematical method used in these simulations easily generalizes to a complete three dimensional transport model of the diffusion-reaction processes occurring in the neuromuscular junction

Gene Expression of Candidate Chemoreceptor Protein Families in Transcriptomes of Two Major Chemosensory Organs and Brain in Decapod Crustaceans

Chemoreceptor proteins are necessary for animals to detect chemical signals and cues in their environment in a process known as chemical sensing. The diversity and number of chemoreceptor proteins have been characterized in many groups of animals, but few have studied the repertoire of chemoreceptor proteins expressed by decapod crustaceans. Crustaceans express at least three classes of putative chemoreceptor proteins. These are: Variant Ionotropic Receptors (IRs), derived from the ionotropic glutamate receptors (iGluRs); Transient Receptor Potential (TRP) channels, a diverse set of sensor-channels; and Gustatory Receptor Like receptors (GRLs), a family of ionotropic receptor proteins that are ancestral to Gustatory Receptors (GRs) of insects. IRs are typically the most numerically dominant of these receptor proteins in crustaceans. In order to identify families of candidate chemoreceptor proteins that are expressed by decapod crustaceans, I examined and compared transcriptomes from four decapod crustaceans that are established models of chemoreception: the Caribbean spiny lobster Panulirus argus, the clawed lobster Homarus americanus, the red swamp crayfish Procambarus clarkii, and the blue crab Callinectes sapidus. Transcriptomes were generated from: a) two major chemosensory organs, the lateral flagella of the antennules (LF) and dactyls of the walking legs (dactyl), of all four decapod crustaceans; and b) the supraesophageal ganglion (brain) of only three decapod crustaceans, P. argus, H. americanus, and P. clarkii. Each species expressed genes for at least ca. 100 to 250 IRs, ca. 15 TRP channels including those shown to be chemoreceptors in other species, and 1 to 4 GRLs. The IRs show different degrees of phylogenetic conservation: protostome-conserved, arthropod-conserved, pancrustacean-conserved, crustacean-conserved, and species-specific. Many IRs appear to be more highly expressed in the LF than dactyl. In the brain transcriptomes, few IRs, almost all TRP channels, and GRLs (in the case of H. americanus) were also detected. Immunocytochemistry in LF and dactyl of P. argus and H. americanus, revealed protein expression of co-receptor IR, IR25a, in olfactory sensory neurons and chemosensory neurons. This research lays the foundation for future functional studies by showing that decapod crustaceans have an abundance of gene expression for chemoreceptor proteins of different types, phylogenetic conservation, and expression patterns

Characterization of multiphase flows integrating X-ray imaging and virtual reality

Multiphase flows are used in a wide variety of industries, from energy production to pharmaceutical manufacturing. However, because of the complexity of the flows and difficulty measuring them, it is challenging to characterize the phenomena inside a multiphase flow. To help overcome this challenge, researchers have used numerous types of noninvasive measurement techniques to record the phenomena that occur inside the flow. One technique that has shown much success is X-ray imaging. While capable of high spatial resolutions, X-ray imaging generally has poor temporal resolution. This research improves the characterization of multiphase flows in three ways. First, an X-ray image intensifier is modified to use a high-speed camera to push the temporal limits of what is possible with current tube source X-ray imaging technology. Using this system, sample flows were imaged at 1000 frames per second without a reduction in spatial resolution. Next, the sensitivity of X-ray computed tomography (CT) measurements to changes in acquisition parameters is analyzed. While in theory CT measurements should be stable over a range of acquisition parameters, previous research has indicated otherwise. The analysis of this sensitivity shows that, while raw CT values are strongly affected by changes to acquisition parameters, if proper calibration techniques are used, acquisition parameters do not significantly influence the results for multiphase flow imaging. Finally, two algorithms are analyzed for their suitability to reconstruct an approximate tomographic slice from only two X-ray projections. These algorithms increase the spatial error in the measurement, as compared to traditional CT; however, they allow for very high temporal resolutions for 3D imaging. The only limit on the speed of this measurement technique is the image intensifier-camera setup, which was shown to be capable of imaging at a rate of at least 1000 FPS. While advances in measurement techniques for multiphase flows are one part of improving multiphase flow characterization, the challenge extends beyond measurement techniques. For improved measurement techniques to be useful, the data must be accessible to scientists in a way that maximizes the comprehension of the phenomena. To this end, this work also presents a system for using the Microsoft Kinect sensor to provide natural, non-contact interaction with multiphase flow data. Furthermore, this system is constructed so that it is trivial to add natural, non-contact interaction to immersive visualization applications. Therefore, multiple visualization applications can be built that are optimized to specific types of data, but all leverage the same natural interaction. Finally, the research is concluded by proposing a system that integrates the improved X-ray measurements, with the Kinect interaction system, and a CAVE automatic virtual environment (CAVE) to present scientists with the multiphase flow measurements in an intuitive and inherently three-dimensional manner

Biomaterial-Mediated Reprogramming of the Wound Interface to Enhance Meniscal Repair

Endogenous repair of fibrous connective tissues is limited, and there exist few successful strategies to improve healing after injury. As such, new methods that advance repair by enhancing cell migration to the wound interface, extracellular matrix (ECM) production, and tissue integration would represent a marked clinical advance. Using the adult meniscus as a test platform, we hypothesized that ECM density and stiffness increase throughout tissue maturation, and that these age-related changes present biophysical barriers to interstitial cell migration during wound healing. We further posited that modulating the matrix could remove these impediments, enabling endogenous cells to reach the injury site. To test our hypotheses, we compared the microenvironment of fetal and adult meniscal ECM via atomic force microscopy (AFM) indentation and second harmonic generation (SHG) imaging of the collagenous matrix. We also explored interstitial cell mobility through fetal and adult native tissue environments using a three-dimensional ex vivo system. We further investigated strategies that might expedite cell migration, including enzymatic degradation of the ECM with collagenase to reduce matrix stiffness and increase porosity. To restrict these biological manipulations to the wound interface, we fabricated a delivery system in which selected biofactors were stored inside composite electrospun nanofibrous scaffolds and released upon hydration. The ability for bioactive scaffolds to enhance the cellularity and integration of meniscal injuries was evaluated in vivo using tissue explants in a subcutaneous implantation model, as well as an orthotopic meniscal injury model. Our findings suggest that matrix stiffness, density, and organization increase with meniscal development at the expense of cell mobility. Our results also indicate that partial digestion of the wound interface with collagenase improves repair by creating a more compliant and porous microenvironment that facilitates cell migration. Furthermore, when scaffolds containing collagenase-releasing fibers were placed inside meniscal defects, enzymatic digestion was localized and resulted in improved cellular colonization and closure of the wound site, similar to treatment with aqueous collagenase. This innovative approach of targeted delivery may aid the many patients that exhibit meniscal tears by promoting integrative repair, thereby circumventing the pathologic consequences of partial meniscus removal, and may find widespread application in the treatment of injuries to a variety of dense connective tissues