This dissertation research explored the capabilities of neuronal networks grown on substrate integrated microelectrode arrays in vitro with emphasis on utilizing such preparations in three specific application domains: pharmacology and drug development, biosensors and neurotoxicology, and the study of burst and synaptic mechanisms. Chapter 1 details the testing of seven novel AChE inhibitors, demonstrating that neuronal networks rapidly detect small molecular differences in closely related compounds, and reveal information about their probable physiological effects that are not attainable through biochemical characterization alone. Chapter 2 shows how neuronal networks may be used to classify and characterize an unknown compound. The compound, trimethylol propane phosphate (TMPP) elicited changes in network activity that resembled those induced by bicuculline, a known epileptogenic. Further work determined that TMPP produces its effects on network activity through a competitive inhibition of the GABAA receptor. This demonstrates that neuronal networks can provide rapid, reliable warning of the presence of toxic substances, and from the manner in which the spontaneous activity changes provide information on the class of compound present and its potential physiological effects. Additional simple pharmacological tests can provide valuable information on primary mechanisms involved in the altered neuronal network responses. Chapter 3 explores the effects produced …

This study examined central mechanisms of persistent pain using an autoradiographic technique to localize phosphoinositide hydrolysis (PI) in the rat spinal cord dorsal horn. The lateral half of laminae I-II showed the highest levels of baseline PI turnover and carbachol-stimulated PI turnover in normal animals as well as after inflammation. Inflammation resulted in increased baseline PI turnover in this region of the ipsilateral (76%) and contralateral (65%) dorsal horns. Carbachol increased PI turnover in this region in normal rats (55%) and following inflammation (ipsilateral: 46%, contralateral: 45%). The absolute magnitudes of these increases were 1.85, 2.71, and 2.51 nCi/mg, respectively. The results of this study demonstrate the involvement of PI turnover in neural mechanisms of persistent pain, and provide evidence for the involvement of cholinergic systems in this process. Because spinal cholinergic systems have been reported to be anti-nociceptive, the present results appear to reflect an upregulation of anti-nociceptive activity in response to inflammation. Thus, the spinal cholinergic system may be a regulatory site within the anti-nociceptive pathway, and may provide an attractive target for the development of new therapeutic agents.

Established interspecific metabolic allometric relationships do not adequately describe the complexity and variable physiological states of developing animals. Consequently, intraspecific allometric relationships of oxygen consumption and carbon dioxide production as a function of body mass; the respiratory quotient; the function of the silk cocoon; and body composition were investigated for each distinct developmental stage of the silkworm, Bombyx mori. Whole animal O2 consumption in Bombyx ranged from 0.00064 + 0.000047 ml O2 .hr-1 at larval instar I to 0.77 + 0.06 ml O2 .hr-1 in pre-pupal, falling to 0.21+ 0.01 ml O2 .hr-1 in the pupae. Those instars having a significant relationship between O2 consumption as a function of body mass, the slope of the line relating O2 consumption to body mass varied between 0.99 and 1.02, while across all instars the slope was 0.82. Developmental allometry should be presented for individual developmental stages because the individual allometric exponents of the stages can be significantly different from the overall allometric exponent throughout development and in some cases, the overall allometric exponent can be a statistical artifact. The first larval instar of Bombyx mori has the lowest cross sectional area of high metabolic tissue of the midgut (27%) and had one …

The black-tailed prairie dog (Cynomys ludovicianus) has experienced dramatic declines over much of its historical range due to habitat loss, plague, poisonings, and shootings. Many populations now occur as isolated genetic relicts. A multiple locus genetic profile was obtained using microsatellite analyses of six polymorphic nucleotide repeats from 319 black-tailed prairie dogs collected from 16 colonies throughout the state of Texas. This assessment revealed that existing populations have sufficient variation at all six loci to verify the usefulness of this approach as a primary genetic tool in conservation and preservation. The data reveals regional-dependent frequency patterns as well as support for founder/bottleneck effects for several of the 16 sites. Although the prairie dog population in Texas as a whole may appear genetically diverse, considerable genetic divergence has already occurred among the subpopulations (FST = 0.164). Isolation by distance is supported by genic differentiation analysis (P < 0.001) and pairwise correlation analysis between genetic distance and geographic distance (P < 0.001). Prairie dogs from six (COC, LUBA, LUBC, LUBD, LUBE, and TAR) of the original 16 sites have been relocated or exterminated, or were in the process of being relocated. Results indicated the following colonies (COT, DAL, HOW, and HUD) are …

Seventy-one aquatic species including the copepodids and nauplii were identified from Lake Texoma from August 1996 to September 1997. Zooplankton community structure, abundance and spatial and temporal distributions were compared among five lake zones delineated a priori based on chloride concentration. The zones, in order of decreasing chloride concentration, are the Red River zone (RRZ), Red river Transition zone (RRTZ), Main Lake zone (MLZ), Washita River Transition zone (WRTZ) and Washita River zone (WRZ). Bray Curtis Similarity Index showed community structure was most similar in the two Red River arm zones, the two Washita River arm zones and the MLZ. Zooplankton abundance was greatest in the Red River arm (312 org/L), intermediate in the Washita River arm (217 org/L) and least in the Main Lake body (103 org/L). A significant increase in the abundance of a deformed rotifer, Keratella cochlearis, was observed mainly in the Red River arm during a second study from March 1999 to June 1999. Seasonal dynamics, rather than spatial dynamics, were more important in structuring the zooplankton community, especially in the two river arms. Spatial variance was solely attributed to station and zone effects independent of time for a few crustacean species and many of the …

This dissertation comprises three parts and is presented in two chapters. Chapter 1 concerns Arthrobacter, a bacterium with an intriguing growth cycle. Whereas most bacteria exist as either a rod or coccus, this bacterium shares the rod/coccus lifestyle. It therefore seemed important to examine the growth regulatory pathways from the rod and coccus. The committed step, that catalyzed by aspartate transcarbamoylase (ATCase), in the pyrimidine biosynthetic pathway was chosen. The ATCase in Arthrobacter is like the well known Pseudomonas enzyme except that it has an active dihydroorotase (DHOase) associated. Included in Chapter 1 is the description of a microorganism, Burkholderia cepacia, whose ATCase has characteristics that are at once reminiscent of bacteria, mammals, and fungi. It differs in size or aggregation based on environmental conditions. In addition, it has an active DHOase associated with the ATCase, like Arthrobacter. B. cepacia is important both medically and for bioremediation. Since B. cepacia is resistant to most antibiotics, its unique ATCase is a prime target for inhibition. Whereas the first chapter deals with the de novo pathway to making pyrimidines, which is found mainly in the lag and log phase, Chapter 2 addresses the salvage pathway, which comes more into play during the …

GABAA receptor binding is transiently increased in rat whisker barrels during the second postnatal week, at a time when neurons in the developing rat cortex are vulnerable to excitotoxic effects. To test whether these GABAA receptors might serve to protect neurons from excessive excitatory input, polymer implants containing the GABAA receptor antagonist bicuculline were placed over barrel cortex for a 4-day period in young (postnatal days 8 - 12) and adult rats. In the cortex of young, but not adult rats, the chronic blockade of GABAA receptors resulted in substantial tissue loss and neuron loss. The greater loss of neurons in young rats supports the hypothesis that a high density of GABAA receptors protects neurons from excessive excitatory input during a sensitive period in development.

This study identifies the biogeographic factors that structure small mammal communities on mountains of the Colorado Plateau and adjacent regions. Forty six isolated ranges were characterized across a 5-state study area encompassing the Colorado Plateau, including the central high plateaus of Utah and the Basin and Range Province (i.e. the Great Basin and mountains of Arizona and New Mexico). Presence/absence data of 25 montane mammal species were used to explore the interactions between historical and ecological processes affecting local and regional diversity patterns. Multivariate analyses, such as non-metric dimensional scaling, were used to explore factors which influence community composition. Results of these analyses revealed the Colorado River as a significant biogeographic barrier that affects montane mammal community structure. MtDNA cytochrome b sequence variation was analyzed among populations of the long-tailed vole, Microtus longicaudus, sampled from five interior ranges of the Colorado Plateau- Abajo, LaSal, Henry, and Chuska Mts., and Boulder Mountain of the Aquarius Plateau-and analyzed using traditional phylogenetic approaches (parsimony and likelihood) as well as nested clade analysis. Results support previous documentation of a major east-west phylogeographic break occurring between populations southeast of the Colorado River (eastern Arizona, Colorado, Wyoming and New Mexico) and all other western populations, which …

The toxicity of methanol, atrazine, and copper to Pseudokirchneriella subcapitata (Korshikov) Hindák historically referred to as Selenastrum capricornutum Printz were determined following 96 hrs growth in a modified Goram's growth media. Methanol and atrazine inhibited fluorescence readings in the cultures by 50% (IC50) at concentrations of 2% and 82 µg/l respectively. These toxicity values compared favorably to other published reports. The IC50 for copper was 160 µg/l which is substantially higher than reported values. This is understandable because of the high chelating capacity of Goram's media. The use of stereologically derived relative volume in the chloroplasts, mitochondria, lipid bodies, phosphate bodies, and nucleus was investigated to determine if it could be used as a sensitive endpoint in toxicity tests. The volume fractions for the chloroplasts and mitochondria were normally distributed in control cells while the nuclei, phosphate bodies, and lipid bodies were not. The chloroplasts were the most dominate organelle occupying a mean relative volume of 46% and mitochondria occupied a mean relative volume of 3%. The nucleus and phosphate bodies occupied a median relative volume of 7% and 2% respectively. The lipid bodies were rare in section profile and no meaningful median relative volume could be calculated. Up to …

Networks of ground-based and satellite borne instruments to measure ultraviolet (UV) sunlight and total column ozone have greatly contributed to an understanding of increased amounts of UV reaching the surface of the Earth caused by stratospheric ozone depletion. Increased UV radiation has important potential effects on human health, and agricultural and ecological systems. Observations from these networks make it possible to monitor total ozone decreases and to predict ozone recovery trends due to global efforts to curb the use of products releasing chemicals harmful to the ozone layer. Thus, continued and expanded global monitoring of ozone and UV is needed. However, existing automatic stratospheric ozone monitors are complex and expensive instruments. The main objective of this research was the development of a low-cost fully automated total column ozone monitoring instrument which, because of its affordability, will increase the number of instruments available for ground-based observations. The new instrument is based on a high-resolution fiber optic spectrometer, coupled with fiber optics that are precisely aimed by a pan and tilt positioning mechanism and with controlling programs written in commonly available software platforms which run on a personal computer. This project makes use of novel low-cost fiber optic spectrometer technology. A cost …

The stonefly genus Isogenoides is revised following a holomorphological approach utilizing traditional morphology and behavioral lines of evidence. Species keys are provided for all life stages. One species, I. krumholzi (Ricker) is considered a synonym of I. doratus (Frison). Detailed species descriptions are provided for males, females, nymphs and ova. Distributions are updated utilizing all known published accounts and materials examined. The vibrational communication (drumming) behavior is reported for males and all but one species for females. The signals were species-specific and ranged in complexity from ancestral sequenced duets to derived grouped exchanges. I. olivaceus is least specialized, having mostly sequenced duets, and I. zionensis most specialized, displaying ancestral sequenced, derived grouped and complex derived exchanges containing both sequenced and grouped elements. Laboratory egg incubation experiments over a 2-4 year period show that Isogenoides has a great capacity for extended, sometimes-asynchronous diapause and hatching. The eggs of six species were incubated at a single, ca. simulated San Miguel River, Colorado, seasonal temperature regime. Direct hatch within two weeks occurred only for I. zionensis (Leopard Creek, Colorado) with small numbers hatching again after one, two, and four years. Eggs of I. doratus and I. varians hatched after an over summer, 3-5 …