The Rio Gavilan watershed, located in Mexico 's northern Sierra Madre Occidental , has significance in conservation history. Upon visiting the remote, largely un­developed watershed during two hunting trips in the 1930s, renowned conservationist Aldo Leopold thought it was the best picture of land health he had seen. His main indicators of healthy land were slow water runoff rates regulating erosion and historical predator-prey relationships. The visits confirmed Leopold's concept of land health, inspired many of his essays, and helped shape his land ethic. Leopold proposed the area as a control site to research healthy land throughout North America . The proposal never went forward and the area has since been more intensively logged and grazed. This dissertation research used extensive literature review, archives, oral histories, citizen surveys, and rapid assessment of forest, rangeland, riparian, and socioeconomic health to assess impacts of past cultures and update the area's land health status. Projects that could restore land health, such as linked eco-tourism, forest density reduction, and rotational grazing, were assessed for feasibility. Recent critiques of Leopold's land ethic were also reviewed. Results indicate most pre-1940s impacts were light, current land health status is moderate, and local interest exists in restoring land …

This study employed a new model of mild-to-moderate head trauma to specifically identify the role of dural mast cell (MC) histamine in trauma-induced increased permeability in the blood-brain barrier (BBB). A single line was scored partially through the left dorsal parietal skull. Immediately following the trauma, degranulation was seen in 39% of the MCs on the left and in 2% on the right. After a 20 min survival period, left duras showed 55% with MC degranulation (fewer with complete degranulation) compared to 34% on the right. In the other experiments two parallel lines were scored following the injection of Evan's blue. Histamine assay showed histamine increased in the left cortex to 154% at 5 min, 174% at 10 min, and 151% at 20 min. Fluorescent quantitation of extravasated Evan's blue at 20 min following the trauma gave an increase of 1385% over the value measured for the right cortex. Zolantidine, a selective histamine H2 receptor antagonist, administered at 10- and 20- mg/kg 30 min before the trauma blocked 65% of the Evan's blue extravasation compared with the control and 2.5 mg group.

Effects of episodic exposures have gained attention as the regulatory focus of the Clean Water Act has shifted away from continuous-flow effluents. Standardized laboratory toxicity tests require that exposure be held constant. However, this approach may not accurately predict organism responses in the field following episodic exposures such as those associated with rain-driven runoff events or accidental pollutant discharge. Using a modified version of the 7-day short-term chronic test recommended by the US Environmental Protection Agency, Ceriodaphnia dubia were exposed to copper or cadmium for durations ranging from 1 minute to 24 hours. In addition, adult reproductive recovery and effects on second generation individuals was assessed following select copper exposures. Finally, cadmium exposures were compared in reconstituted hard water (RHW) and municipal treated wastewater effluent (TWE). Following exposure, organisms were transferred to clean RHW or TWE and maintained for the remainder of the test. No- and lowest observed effect concentrations (NO- and LOECs) increased logarithmically with respect to logarithmic decreases in duration regardless of metal, endpoint or water type. Effective concentrations of cadmium however, were usually higher than those of copper, especially in TWE. LOECs for C. dubia survival following 24-hour and 5-minute exposures to copper were 116 and 417 …

Spontaneously active frontal cortex and spinal cord networks grown on microelectrode arrays were used to study effects of D-methylphenidate. These central nervous system tissues have relatively low concentrations of dopaminergic and noradrenergic neurons compared to the richly populated loci, yet exhibit similar neurophysiological responses to methylphenidate. The spontaneous spike activity of both tissues was inhibited in a concentration-dependent manner by serial additions of 1-500 µM methylphenidate. Methylphenidate is non-toxic as spike inhibition was recovered following washes. The average concentrations for 50% spike rate inhibition (IC50 ± SD) were 118 ± 52 (n= 6) and 57 ± 43 (n = 11) for frontal cortex and spinal cord networks, respectively. A 3 hour exposure of a network to 1 mM methylphenidate was nontoxic. The effective concentrations described in this study are within the therapeutic dosage range. Therefore, the platform may be used for further investigations of drug mechanisms.

Embryonic murine neuronal networks cultured on microelectrode arrays were used to quantify acute electrophysiological effects of fluoxetine and ethanol. Spontaneously active frontal cortex cultures showed highly repeatable, dose-dependent sensitivities to both compounds. Cultures began to respond to fluoxetine at 3 µM and were shut off at 10-16 µM. EC50s mean ± S.D. for spike and burst rates were 4.1 ± 1.5 µM and 4.5 ± 1.1 µM (n=14). The fluoxetine inhibition was reversible and without effect on action potential wave shapes. Ethanol showed initial inhibition at 20 mM, with spike and burst rate EC50s at 52.0 ± 17.4 mM and 56.0 ± 17.0 mM (n=15). Ethanol concentrations above 100 -140 mM led to cessation of activity. Although ethanol did not change the shape and amplitude of action potentials, unit specific effects were found. The combined application of ethanol and fluoxetine was additive. Ethanol did not potentiate the effect of fluoxetine.

Murine neuronal networks, derived from embryonic frontal cortex (FC) tissue grown on microelectrode arrays, were used to investigate zinc toxicity at concentrations ranging from 20 to 2000 mM total zinc acetate added to the culture medium. Continual multi-channel recording of spontaneous action potential generation allowed a quantitative analysis of the temporal evolution of network spike activity generation at specific zinc acetate concentrations. Cultures responded with immediate concentration-dependent excitation lasting from 5 to 50 min, consisting of increased spiking and enhanced, coordinated bursting. This was followed by irreversible activity decay. The time to 50% and 90% activity loss was concentration dependent, highly reproducible, and formed linear functions in log-log plots. Network activity loss generally preceded morphological changes. 20% cell swelling was correlated with 50% activity loss. Cultures pretreated with the GABAA receptor antagonists bicuculline (40 mM) and picrotoxin (1 mM) lacked the initial excitation phase. This suggests that zinc-induced excitation may be mediated by interfering with GABA inhibition. Partial network protection was achieved by stopping spontaneous activity with either tetrodotoxin (200 nM) or lidocaine (250 mM). However, recovery was not complete and slow deterioration of network activity continued over 6 hrs. Removal of zinc by early medium changes showed irreversible, catastrophic …

Spontaneously active monolayer neuronal networks cultured on photoetched multimicroelectrode plates (MMEPs) offer great potential for use in studying neuronal networks. However, there are many problems associated with frequent, long-term use of MMEPs. The major problems include (1) polysiloxane insulation deterioration and breakdown, (2) and loss of gold at the gold electroplated indium-tin oxide (ITO) electrodes. The objective of this investigation was to correct these major problems. Quality control measures were employed to monitor MMEP fabrication variables. The phenotypes of polysiloxane degradation were identified and classified. Factors that were found to contribute most to insulation deterioration were (1) moisture contamination during MMEP insulation, (2) loss of the quartz barrier layer from excessive exposure to basic solutions, and (3) repetitive use in culture. As a result, the insulation equipment and methods were modified to control moisture-dependent insulation deterioration, and the KOH reprocessing solution was replaced with tetramethylguanidine to prevent damage to the quartz. The problems associated with gold electroplating were solved via the addition of a pulsed-DC application of gold in a new citrate buffered electroplating solution.

N-Acylethanolamines (NAEs) are bioactive acylamides that are present in a wide range of organisms. Because NAE levels in seeds decline during imbibition similar to ABA, a physiological role was predicted for these metabolites in Arabidopsis thaliana seed germination and seedling development. There is also a corresponding increase of AtFAAH (fatty acid amide hydrolase), transcript levels and activity, which metabolizes NAE to ethanolamine and free fatty acids. Based on whole genome microarray studies it was determined that a number of up-regulated genes that were responsive to NAE were also ABA responsive. NAE induced gene expression in these ABA responsive genes without elevating endogenous levels of ABA. It was also determined that many of these NAE/ABA responsive genes were associated with an ABA induced secondary growth arrest, including ABI3. ABI3 is a transcription factor that regulates the transition from embryo to seedling growth, the analysis of transcript levels in NAE treated seedlings revealed a dose dependent, inverse relationship between ABI3 transcript levels and growth, high ABI3 transcript levels were associated with growth inhibition. Similar to ABA, NAE negatively regulated seedling growth within a narrow window of early seedling establishment. When seedlings are exposed to NAE or ABA within the window of sensitivity, …

Heavy metal bioaccumulation in aquatic organisms may occur through direct or indirect uptake routes. Research indicates that the significance of uptake route varies with contaminant and organism exposed. The relative importance of different metal sources in aquatic systems was investigated by exposing freshwater snails to dietary or dissolved sources of cadmium. Snails were exposed to control, contaminated food only, contaminated water only, and contaminated food and water treatments. During the 15-day exposure, samples were taken to determine Cd concentration in snail soft tissue, snail shell, algal food, and overlying water. Analyses of snail soft tissue and shells indicate that exposure route significantly affects Cd concentrations in the tissues. In both cases, dissolved Cd is the primary contributor to metal body burden.

Putative cotton cDNA clones and cognate genomic clones for a palmitoyl-acyl carrier protein (ACP) thioesterase (PATE) and an osmotin-like pathogenesis-related 5 (PR5) protein have been isolated and characterized. PATE is a class B fatty acid thioesterase with specificity for saturated long-chain fatty acids such as palmitate, and is implicated as a key enzyme to be targeted for regulation of fatty acid synthesis in order to alter cotton seed oil profiles. A nearly full-length 1.7-kb cDNA clone was isolated using a hybridization probe derived from an Arabidopsis PATE cDNA clone designated TE 3-2. A 17-kb genomic segment encompassing the PATE gene was also isolated, which has six exons and five introns with high sequence identity with other FatB cDNA/gene sequences. The deduced PATE preprotein amino acid sequence of 413 residues has putative signal sequences for targeting to the chloroplast stroma. PR5 proteins called osmotins are made in response to fungal pathogen stress or osmotic stress (water deprivation or salt exposure). Osmotins may actually form pores in fungal membranes, leading to osmotic rupture and destruction of the fungal cells. A cotton osmotin-like PR5 cDNA insert of 1,052 base-pairs was isolated and shown to encode a preprotein of 242 amino acids and is …

This research is designed to test how sublethal exposure to copper affects tadpole predator-escape behavior and how quickly tadpoles recover. After exposure, tadpoles were separated. Escape behavior was recorded for two-thirds of exposed tadpoles while one-third of the exposed population was measured weekly to determine growth and recovery. Control tadpoles were consumed within 15 minutes whereas those exposed to higher concentrations were consumed at a slower rate, which does not support the hypotheses. Although the rate of predation was lower, tadpoles exposed to higher Cu concentrations were on average, 1.47 cm in total body length. Those exposed to 0.93 mg/L averaged 0.86 cm. After being placed into clean water, treatment tadpoles recovered after 20 days.

An accurate fire model is a useful tool in predicting the behavior of a prescribed fire. Simulation of fire requires an extensive amount of data and can be accomplished best using GIS applications. This paper demonstrates integrative procedures of using of ArcGIS™, ERDAS Imagine™, GPS, and FARSITE© to predict prescribed fire behavior on the Kaibab-Paiute Reservation. ArcGIS was used to create a database incorporating all variables into a common spatial reference system and format for the FARSITE model. ArcGIS Spatial Analyst was then used to select optimal burn sites for simulation. Our predictions will be implemented in future interagency efforts towards vegetative restoration on the reservation.

I assessed fish community data collected by the Oklahoma Conservation Commission from 82 minimally impaired wadeable reference streams in eastern Oklahoma to determine whether existing aquatic ecoregions provide the best framework for spatial classification for the development of biological assessment methods and biocriteria. I used indirect ordination and classification to identify groups of sites that support similar fish communities. Although correspondence was observed between fish assemblages and three montane ecoregions, the classification system must be refined and expanded to include major drainage basins and physical habitat attributes for some areas to adequately partition variance in key measures of biological integrity. Results from canonical correspondence analysis indicated that substrate size and habitat type were the primary physical habitat variables that influenced the fish species composition and community structure.

Growth and element assimilation was investigated in post oak seedlings exposed to four different treatment combinations of fertilization and ectomycorrhizal inoculation. Element concentration in excised leaves was analyzed via particle induced X-ray emission spectrometry with a 1.8 MeV proton macrobeam. Mean growth was significantly different across the treatment groups as well as mean concentration of Mg, Al, S, K, Ca, Fe, Cu, and Zn. The data suggest that fertilization rather than mycorrhizal inoculation had a stronger influence on plant growth and nutrient uptake. A follow up study was conducted with a 3 MeV microbeam. A 850 μm2 scanned area of a post oak leaf produced topographical maps of 11 elements.

This thesis describes the development of an optimized method for culturing dissociated, monolayer neuronal networks from murine frontal cortex and midbrain. It is presented as a guidebook for use by cell culture specialists and laboratory personnel who require updated and complete procedures for use with microelectrode array (MEA) recording technology. Specific cell culture protocols, contamination prevention and control, as well common problems encountered within the cell culture facility, are discussed. This volume offers value and utility to the rapidly expanding fields of MEA recording and neuronal cell culture. Due to increasing interest in determining the mechanisms underlying Parkinson's disease, the newly developed procedures for mesencephalon isolation and culture on MEAs are an important research contribution.

Cowpea, Vigna unguiculata (L.) Walp is an excellent source of protein, vitamins and minerals and a major food crop many parts of Africa. Optimal production levels are hampered by insect pests and diseases. Biotechnological techniques such as tissue culture and genetic engineering can aid in the development of varieties with resistance to insect pests and diseases. The objective of this study was to investigate conditions necessary for the development of a reproducible tissue culture system that can be applied to regenerate transformed cells from culture. The in vitro manipulation of cowpea using Murashige and Skoog (MS) medium, auxins and cytokinins resulted in the formation of callus and rhizogenesis. Calli that were formed were separated into six classes based on color and texture. Yellowish friable callus, yellowish compact, soft yellowish callus and green and white were composed of largely vacuolated cells and were non-regenerative. Friable green callus was the most prevalent callus type and could form of roots in some hormone combinations. Green spots were formed on hard compact green callus. The green spots became nodular, forming root primordia and ultimately giving rise to roots. None of the six calli types gave rise to the formation of shoots. Embryogenic callus was …

Changes in spontaneous spike activities recorded from murine frontal cortex networks grown on substrate-integrated microelectrodes were used to determine the dissociation constant (KB) of three GABAA antagonists. Neuronal networks were treated with fixed concentrations of GABAA antagonists and titrated with muscimol, a GABAA receptor agonist. Muscimol decreased spike activity in a concentration dependent manner with full efficacy (100% spike inhibition) and a 50% inhibitory concentration (IC50) of 0.14 ± 0.05 µM (mean ± SD, n=6). At 10, 20, 40 and 80 µM bicuculline, the muscimol IC50 values were shifted to 4.3 ± 1.8 µM (n=6), 6.8 ± 1.7 µM (n=6), 19.3 ± 3.54 µM (n=10) and 43.5 µM (n=2), respectively (mean ± SD). Muscimol titration in the presence of 10, 20, 40 µM of gabazine resulted in IC50s values of 20.1 (n=2), 37.17 (n=4), and 120.45 (n=2), respectively. In the presence of 20, 80, and 160 µM of TMPP (trimethylolpropane phosphate) the IC50s were 0.86 (n=2), 3.07 (n=3), 6.67 (n=2) µM, respectively. Increasing concentrations of GABAA antagonists shifted agonist log concentration-response curves to the right with identical efficacies, indicating direct competition for the GABAA receptor. A Schild plot analysis with linear regression resulted in slopes of 1.18 ± 0.18, 1.29 …

Throughout history floods have been part of the human experience. They are recurring phenomena that form a necessary and enduring feature of all river basin and lowland coastal systems. In an average year, they benefit millions of people who depend on them. In the more developed countries, major floods can be the largest cause of economic losses from natural disasters, and are also a major cause of disaster-related deaths in the less developed countries. Flood disaster mitigation research was conducted to determine how remotely sensed data can effectively be used to produce accurate flood plain maps (FPMs), and to identify/quantify the sources of error associated with such data. Differences were analyzed between flood maps produced by an automated remote sensing analysis tailored to the available satellite remote sensing datasets (rFPM), the 100-year flooded areas "predicted" by the Flood Insurance Rate Maps, and FPMs based on DEM and hydrological data (aFPM). Landuse/landcover was also examined to determine its influence on rFPM errors. These errors were identified and the results were integrated in a GIS to minimize landuse / landcover effects. Two substantial flood events were analyzed. These events were selected because of their similar characteristics (i.e., the existence of FIRM or …

In order to understand the structural changes in myosin S1, fluorescence polarization and computational dynamics simulations were used. Dynamics simulations on the S1 motor domain indicated that significant flexibility was present throughout the molecular model. The constrained opening versus closing of the 50 kDa cleft appeared to induce opposite directions of movement in the lever arm. A sequence called the "strut" which traverses the 50 kDa cleft and may play an important role in positioning the actomyosin binding interface during actin binding is thought to be intimately linked to distant structural changes in the myosin's nucleotide cleft and neck regions. To study the dynamics of the strut region, a method of fluorescent labeling of the strut was discovered using the dye CY3. CY3 served as a hydrophobic tag for purification by hydrophobic interaction chromatography which enabled the separation of labeled and unlabeled species of S1 including a fraction labeled specifically at the strut sequence. The high specificity of labeling was verified by proteolytic digestions, gel electrophoresis, and mass spectroscopy. Analysis of the labeled S1 by collisional quenching, fluorescence polarization, and actin-activated ATPase activity were consistent with predictions from structural models of the probe's location. Although the fluorescent intensity of the …

Temperature effects on the development of the zebrafish embryos and larvae and adults were examined. It was found that the earlier in development a temperature change was performed on an embryo, the more significant the change in survival and/or subsequent development. Thus, viable temperature ranges for zebrafish widened significantly as development proceeded. Adults reared and bred at 25oC produced embryos that were significantly more successful at the lower range of rearing temperatures compared to embryos produced from adults reared at 28oC. The majority of this study focused on the physiological effects of swim training during development in the zebrafish. The earlier in development the zebrafish larvae were trained, the greater the mortality. Trained free swimming larvae had a significantly higher routine oxygen consumption after 11 days of training, and a higher mass specific routine metabolic rate after 8 and 11 days of training. Trained free swimming larvae consumed significantly less oxygen during swimming and were more efficient at locomotion, compared to control larvae. Training enhanced survival during exposure to extreme hypoxia in all age groups. Performance aspects of training were investigated in attempt to quantify training effects and in most cases, trained fish performed significantly better than controls. As blood …

Food waste in the waste stream is becoming an important aspect of integrated waste management systems. Current efforts are composting and animal feeding. However, these food waste disposal practices rely on slow thermodynamic processes of composting or finding farmers with domestic animals capable of consuming the food wastes. Bioconversion, a potential alternative, is a waste management practice that converts food waste to insect larval biomass and organic residue. This project uses a native and common non-pest insect in Texas, the black soldier fly, which processes large quantities of food wastes, as well as animal wastes and sewage in its larval stage. The goal of this research is to facilitate the identification and development of the practical parameters of bioconversion methods at a large cafeteria. Three major factors were selected to evaluate the practicality of a bioconversion system: (1) the biological constraints on the species; (2) the economic costs and benefits for the local community; (3) the perception of and interaction between the public and management agencies with respect to the bioconversion process. Results indicate that bioconversion is feasible on all levels. Larvae tolerate and consume food waste as well as used cooking grease, reducing the overall waste volume by 30-70% …

In this study, two different field experiments were designed to assess the relative influence of urbanization on benthic communities. During spring and summer, four urban and one reference sites from Denton County, Texas were selected for benthic macroinvertebrate evaluation. Statistically significant differences in colonized benthic macroinvertebrate taxa on artificial substrates were observed among the four urban sites and the reference site. Oligochaetes and chironomids were the dominant taxa at all sites. Identification of chironomid larvae at the subfamily and genus level to detect differences between sites had higher statistical power than the evaluation based on total chironomids. At the reference site, Caenis, Cladotanytarsus, Orthocladius, and Ceratopogonidae were the dominant taxa, while the urban sites were dominated by Dero, Physella, Ancylidae, Chironomus, Dicrotendipes, Glyptotendipes, Polypedilum, Pseudochironomus, Stenochironomus, and Tanytarsus. These differences may have been dependent upon differences in hydrologic regime and water quality between sites. Significant differences (ANOVA, p < 0.01) in water quality parameters (alkalinity, hardness, nitrates, phosphates, chlorides, sulfates, calcium, magnesium, potassium, and triazine) were found among water samples collected from the reference and urban sites. During the transfer period, most of the Ephemeroptera and Trichoptera taxa and a few other taxa disappeared from artificial substrates that were colonized …

A biological monitoring tool to assess water quality using bivalve gape behavior was developed and demonstrated. The purpose of this work was to develop methodologies for screening water quality appropriate to the goals of the watershed paradigm. A model of bivalve gape behavior based on prediction of behavior using autoregressive techniques was the foundation of the bivalve biomonitoring system. Current technology was used in developing the system to provide bivalve gape state data in a continuous real-time manner. A laboratory version of the system, including data collection and analysis hardware and software, was developed for use as a toxicological assay for determination of effective concentrations of toxicant(s) or other types of stress on bivalve gape behavior. Corbicula fluminea was monitored and challenged with copper, zinc, and chlorpyrifos using the system. Effective concentrations of 176±23µg/L copper, 768±412µg/L zinc, and 68µg/L chlorpyrifos were observed using a natural water with high dissolved organic carbon concentrations. A rugged field version of the bivalve biomonitoring system was developed and deployed in two locations. The field systems were fitted with a photovoltaic array, a single board computer, and a CDPD telemetry modem for robust remote operation. Data were telemetered at a time relevant rate of once …