Magister Pharmaceuticae - MPharm / Neurodegenerative disorders (NDs) are a range of chronic brain disorders that includes amongst others motor function loss. Parkinson’s disease (PD) is one of the common NDs that has an insidious onset and diagnosed when dopaminergic neurons in the substantia nigra are already lost. The loss creates a deficiency of the dopamine (neurotransmitter) thereby causing neurochemical imbalance resulting in the signs and symptoms of PD. NDs overlap at multiple levels so some of the symptoms overlap as well. NDs currently have no cure yet and current drug therapies only improve the quality of life of the patients by targeting the symptoms mainly. Treatment of PD currently involves different classes of drugs and depending on the stages of the disease, some drugs can be only used as an adjunct therapy. Anti-oxidants and monoamine oxidase inhibitors (MAO-I) are part of the treatment options.

The increase in extracellular dopamine (DA) following cocaine administration plays a major role in cocaine abuse. In vitro, cocaine binds to DA transporters (DAT) and blocks DA uptake. Moreover, cocaine can increase extracellular DA concentration as measured by in vivo neurochemical methods. The present study examined the effects of cocaine and other drugs on DA, NE and 5-HT uptake using an ex vivo assay. Rats were injected i.v. with saline or drug and sacrificed at various time points after injections. Brains were dissected for regional monoamine uptake studies ex vivo. In most brain regions, cocaine given in vivo blocked monoamine uptake as expected. [ H]DA uptake in nucleus accumbens was inhibited with an ED = 22.3 μmol/kg. Cocaine fully inhibited [ H]NE uptake (ED = 4.58 μmol/kg) in the occipital cortex and partially inhibited [ H]5-HT uptake (33% at 30 μmol/kg) in the midbrain. However, under the same conditions [ H]DA uptake in the striatum was not inhibited after injections of cocaine up to 56 μmol/kg. Although the mechanism for this discrepancy is unclear, DA binding and uptake sites may be distinct and/or there may be regional differences in DA transporters.

No description available.

Monoamine oxidase (MAO) is a mitochondrial membrane-bound enzyme that catalyzes the oxidative deamination of monoamines in a wide array of organisms. While the enzyme monoamine oxidase has been studied extensively in its role in moderating behavior in mammals, there is a paucity of research investigating this role in invertebrates, where the latter utilizes this enzyme in a major pathway to degrade monoamines. There is especially a dismal lack of information on how MAO influences activity in invertebrates, particularly in account of the circadian cycle. Previous studies revealed MAO degrades serotonin and norepinephrine in arachnids, but did not investigate other critically important compounds like octopamine. Larinioides cornutus is a species of orb-weaving spider that exhibits diel fluctuations in behavior, specifically levels of aggression. The monoamines octopamine and serotonin have been shown to influence aggressive behaviors in L. cornutus, thus this species was used to investigate if MAO is a potential site of regulation throughout the day. Not only did gene expression of MAO orthologs and MAO activity fluctuate at different times of day, but the enzymatic activity was substrate-specific producing a higher level of degradation of octopamine as compared to serotonin in vitro. This study further supports evidence that MAO has an active role in monoamine inactivation in invertebrates and provides a first look at how MAO ultimately may be regulating behavior in an invertebrate.

No description available.

<p> Serotonin positive (5HT+) and tyrosine hydroxylase positive (TH+) cells were identified using fluorescent immunocytochemical methods and quantified in the gill epithelium of six species of fish. 5HT+ cells were located in the filament epithelium in contact with the basal lamina on the efferent side, and in the lamellar epithelium where they were occasionally exposed to the external milieu. Thus, these cells appear to represent two populations of neuroepithelial cells (NEC) as proposed in other studies. In trout, bass and killi fish, NECs were revealed exclusively in the primary epithelium. In tilapia, NECs were located exclusively in the secondary epithelium, whereas in perch and zebrafish they occurred in both epithelial layers. The two types of NECs varied in number both within and among the species. Seasonal comparisons of NECs in perch revealed a decrease in cell density in the filament between July and November, though there was no significant difference in the density of NECs in the lamellae. TH+ cells were identified in perch, zebrafish and killi fish. In zebrafish TH+ cells occurred in similar numbers to 5HT+ cells, and were generally present in similar locations. It is likely that in this case, many of the labelled cells were positive for both markers. In two of the species, perch and killi fish, the density and distribution was such that the TH+ cells and 5HT+ cells were unlikely to be the same. A quantitative comparison of total catecholamine (CA) stores, using high performance liquid chromatography (HPLC), revealed that gill tissues in general contained higher levels of epinephrine (EPI) than norepinephrine (NOR) and dopamine (DOP). Finally, attempts were made to determine whether NECs would survive in 2-4 day old cultures of dispersed gill cells from perch, using immunocytochemical labelling for 5HT. A few successful cases are presented.</p> / Thesis / Master of Science (MSc)

The parkinsonian inducing drug 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) is bioactivated in a reaction catalyzed by the flavoenzyme monoamine oxidase B (MAO-B) to form the corresponding dihydropyridinium (MPDP+) subsequently pyridinium (MPP+) metabolites. As part of our ongoing studies to characterize the structural features responsible for this unexpected biotransformation, we have synthesized and examined the MAO-B substrate properties of a variety of MPTP analogs bearing various heteroaryl groups at the 4-position of the tetrahydropyridinyl ring. The results of these SAR studies indicate that electronic features, steric features and polar interactions can contribute to the substrate activities. Additionally, isotope effects have been examined to investigate the mechanism and stereoselectivity of the MAO-B catalytic pathway. The synthesis and characterization of regio and stereoselectively deuterated MPTP analogs have been achieved. The results indicate that the catalytic step occurs exclusively at the allylic C-6 position and is rate-determining for both good and poor substrates. The two enantiomers of MPTP bearing a deuterium atom at C-6 have been prepared via chiral aminooxazolinyl derivatives and have been characterized by 2H NMR in a chiral liquid crystal matrix. These enantiomers were used to determine the selectivity of the MAO-B catalyzed a C-H bond cleavage reaction leading to the dihydropyridinium metabolite MPDP+. Some of the cyclopropyl analogs of MPTP have also been synthesized as the potential inhibitors. / Ph. D.

The neuronal nitric oxide synthase (nNOS) inhibitor 7-nitroindazole (7-NI) protects against the neurotoxicity of MPTP in a mouse model of neurodegeneration. Since 7-NI also inhibits the monoamine oxidase-B (MAO-B) catalyzed bioactivation of MPTP, the role of nNOS inhibition as a mediator of 7-NI's neuroprotective properties have been challenged. In order to examine in greater detail the neuroprotective effects of indazolyl derivatives, the synthesis of water soluble indazolyltetrahydropyridinyl derivatives as potential "prodrugs" that may undergo MAO bioactivation in the brain was undertaken. During the course of the studies on the synthesis of indazolylpyridinium derivatives, precursors to these "prodrugs", an interesting reaction involving the rearrangement of 4-(2H-indazolyl)-1-methylpyridinium iodide to the corresponding 1H-isomer was encountered. A detailed investigation of this rearrangement reaction is reported in this thesis. The syntheses and interaction of nitroindazolyltetrahydropyridinyl "prodrugs" with MAO-B have been investigated previously. Molecular docking studies that attempt to explain the MAO-B substrate and inhibitor properties of members of this series of compounds are described. Finally, the MAO-A substrate properties of nitroindazolyltetrahydropyridinyl derivatives are reported. / Ph. D.

The flavin-containing monoamine oxidases (MAO) A and B catalyze the oxidative deamination of primary and secondary amines. The overall process involves a two electron oxidation of the amine to the iminium with concomitantreduction of the flavin. Based on extensive studies with a variety of chemical probes, Silverman and colleagues have proposed a catalytic pathway for the processing of amine substrates and inactivators by MAO-B that is initiated by a single electron transfer (SET) step from the nitrogen lone pair to the oxidized flavin followed by α-proton loss from the resulting amine radical cation that leads to a carbon radical. Subsequent transfer of the second electron leads to the reduced flavin and the iminium product. In the case of N-cyclopropylamines, the initially formed amine radical cation is proposed to undergo rapid ring opening to form a highly reactive primary carbon centered radical that is thought to be responsible for inactivation of the enzyme. In this thesis we have exploited the unique substrate and inactivator properties of 1,4-disubstituted tetrahydropyridine derivatives to probe the mechanism of MAO-B catalysis. Reports of the parkinsonian inducing neurotoxin 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine (MPTP) as a structurally unique substrate of MAO-B initiated these studies. Consistent with the SET pathway, the N-cyclopropyl analog of MPTP proved to be an efficient time and concentration dependent inactivator but not a substrate of MAO-B. On the other hand, the 4-benzyl-1-cyclopropyl analog is both a substrate and inactivator of MAO-B. These properties may not be consistent with the obligatory formation of a cyclopropylaminyl radicalcation intermediate. In an attempt to gain further insight into the mechanism associated with the MAO catalyzed oxidation of 1,4-disubstituted tetrahydropyridines, deuterium isotope effects studies on both the substrate and inactivation properties of the 4-benzyl-1- cyclopropyl derivative were undertaken. A series of 1-methyl- and 1-cyclopropyltetrahydropyridine derivatives bearing various heteroaro-matic groups at C-4 also have been examined. The MAO-B substrate properties, inactivator properties and partition ratios for these compounds together with preliminary results from chemical model studies are discussed in terms of the MAO-B catalytic pathway. / Ph. D.

Smoking is one of the leading causes of death in the world. About 80% of smokers start smoking before the age of 18. In the Appalachian area and the South in the United States, smoking percentages among adults and adolescents are higher than in other regions. Female smoking shows a variety of different trends from male smoking, and smoking brings particular health problems related to production to female smokers. These findings highlighted the importance of studying female teenage smokers in southwest Virginia. The initial project aimed to identify risk factors that might prevent smoking in an early stage. Dr. Helen Crawford led the Cognitive Neuroscience Lab at Virginia Tech in discovering the psychophysiological vulnerabilities of female teenage smokers. Toward this end, event-related potential (ERP), personality, and behavioral data were collected in teenage female smokers and non-smokers. These data were analyzed to examine possible psychophysiological vulnerabilities in female teenage smokers such as deficits in brain and cognitive function, personality traits, and environment influences. The purpose of this dissertation is to further analyze these data to elaborate and clarify the relationships among these vulnerabilities toward understanding teenage smoking behavior. Participants were 49 teenage girls (smokers and non-smokers) with age from 14 to 18. The measures included sensory gating, platelet MAO-B activity, attention, memory, temperament, schizotypal personality, recognition of facial expressions, taste and smell. The initial set of analyses compared smokers and non-smokers, including those classified as high and low dependent, on all dependent measures. The results suggested some psychophysiological vulnerabilities in female teenage smokers, which have been used as support for the self-medication and the orbito-frontal dysfunction models of why teenagers smoke (Crawford et al., 2004). Further examination of these factors may help teenagers to reduce the smoking dependency and possibly improve cognitive function. Specifically, this dissertation focused on the role of the variable of monoamine oxidase-B (MAO-B) in the correlations among sensory gating, MAO and other cognitive and personality measures. All smokers were divided into high and low MAO groups first. Comparison analyses were conducted between them. The high MAO group showed better sensory gating function than the low MAO group. Correlation analyses were conducted among all of the measures. The significant linear relationships between MAO and sensory gating, MAO and CO level and MAO and temperament were demonstrated. MAO activity positively correlated with the sensory gating function and negatively correlated with CO level and temperament characteristics. Finally, to explore the mechanisms of the relationship between MAO and sensory gating, the neurotransmitter systems related to MAO and sensory gating were discussed. / Ph. D.