E-thesis / Viikki Campus

17.2. Mikko Käenmäki: Importance of COMT in the regulation of prefrontal dopamine

Käenmäki, Mikko

The prefrontal cortex (PFC), located in the anterior region of the frontal lobe, is considered to have several key roles in higher cognitive and executive functions. In general, the PFC can be seen as a coordinator of thought and action allowing subjects to behave in a goal-directed manner. Due to its anatomical connections with a variety of cortical and subcortical structures, several neurotransmitters, including dopamine, are involved in the regulation of PFC activity. In general, the majority of released dopamine is cleared by the dopamine transporter (DAT). In the PFC however, the number of presynaptic DAT is diminished, emphasizing the relative importance of catechol-O-methyltransferase (COMT) in dopamine metabolism. As a result, the role of COMT in the etiology of psychotic disorders is under constant debate.

The present study investigated the role of COMT in prefrontal cortical dopamine metabolism by different neurochemical methods in COMT knockout (COMT-KO) mice. Pharmacological tools to inhibit other dopamine clearing mechanisms were also used for a more comprehensive and collective picture. In addition, this study investigated how a lack of the soluble (S-) COMT isoform affects the total COMT activity as well as the pharmacokinetics of orally administered L-dopa using mutant mice expressing only the membrane-bound (MB-) COMT isoform. Also the role of COMT in striatal and accumbal dopamine turnover during Δ9-tetrahydrocannabinol (THC) challenge was studied.

We found markedly increased basal dopamine concentrations in the PFC, but not the striatum or nucleus accumbens (NAcc), of mice lacking COMT. Pharmacological inhibition of the noradrenaline transporter (NET) and monoamine oxidase (MAO) elevated prefrontal cortical dopamine levels several-fold, whereas inhibition of DAT did not. The lack of COMT doubled the dopamine raising effects of NET and MAO inhibition. No compensatory expression of either DAT or NET was found in the COMT-KO mice. The lack of S-COMT decreased the total COMT activity by 50-70 % and modified dopamine transmission and the pharmacokinetics of exogenous Ldopa in a sex and tissue specific manner. Finally, we found that subsequent tolcapone and THC increased dopamine levels in the NAcc, but not in the striatum.

Conclusively, this study presents neurochemical evidence for the important role of COMT in the PFC and shows that COMT is responsible for about half of prefrontal cortical dopamine metabolism. This study also highlights the previously underestimated proportional role of MB-COMT and supports the clinical evidence of a gene x environment interaction between COMT and cannabis.

23.2. Marjo Paavola: The Impact of Village Development Funds on Community Welfare in the Lao People's Democratic Republic

Paavola, Marjo

Micro credits have become an important tool in development efforts globally as well as at the national or local level. They are also increasingly linked to sustainable forest management and its role in poverty reduction in developing countries. The potential of micro credits in alleviating poverty is now recognized worldwide. Governments, donors, development agencies, banks, researchers and consultants are also increasingly interested in microfinance.

The objectives of the present study were, using Lao PDR as a case country, (1) to analyse the livelihood assets and activities of selected households with the emphasis on a comparison between poor and non-poor households and between those which had and had not received a micro credit; (2) to assess whether the well-being of the villages and households had improved as a consequence of the application and implementation of village development funds, that is, micro credits, and which were the factors affecting poverty; and (3) to evaluate the opinions of provincial level forestry and district leaders and other leaders on village development funds in contributing to well-being in Laos. The theories of sustainable livelihoods and entrepreneurship were used in analysing the impact of micro credits on the welfare of villagers and their livelihood assets. A basic assumption was also that access to various levels of assets and the combinations of assets both have an influence on peoples choice of livelihood strategies.

Village and household survey data were used. These data were collected in the field in the form of interviews with closed questions. Village heads were interviewed, as well as the heads and the members of the selected households. In total, 76 villages were studied and 1,518 households were interviewed. The village development financing system was assessed through provincial and district level semi-structured interviews, by focusing in particular on the adequacy and appropriateness of the development financing guidance, tools and training.

A quantitative analysis suggested that the effects of the SUFORD project and micro credits in the form of village development funds were small or non-existent. However, the quantitative analysis could define the specific factors related to poverty as follows: ethnic origin; glutinous rice production during the wet season; the farm area; the goods and chattel situation and the house type compared to the situation three years earlier; and the off-farm income of the household.

In contrast to the quantitative analysis, the major result of a qualitative analysis was that the SUFORD project had improved the welfare situation of the households and villages. The positive results of the qualitative analysis may be explained by the fact that the SUFORD project was perhaps regarded by the rural communities as much more powerful than it really was.

16.3. al-Hello Haider : Pathogenic and Molecular Characteristics of Two Isolates of HEV-B Species

Haider , al-Hello

Enteroviruses (EVs) are small non-enveloped RNA viruses forming a large group of different serotypes. EVs belong to the family Picornaviridae. The primary replication site of an enterovirus is typically the epithelium of the respiratory tract and the gastrointestinal mucosa. Virus replication in the gastrointestinal mucosa may continue, often asymptomatically, for several weeks occasionally causing viremia. During the viremia the virus spreads through the lymphatic system and circulation. Organ-specific symptoms rise after viral replication in the secondary target tissues. Occasionally, cellular adaptation is required for a virus to initiate replication in the secondary target tissue(s). Adaptation is linked to mutation(s) which may lead to alteration in cellular tropism, e.g., recognition of new surface receptor molecules or other host cell constituents essential for virus entry and replication. However, the critical step may also occur later during in the interaction of the host cell and the replicating virus. In the present study, genetic changes responsible for altered phenotypic features were sought using two strains of Human enterovirus B (HEV-B) species. Firstly, a laboratory isolate of coxsackievirus B5 (CV-B5), strain DS, was passaged 15 times in mouse pancreas in vivo, which resulted in a diabetogenic mouse pancreas passaged virus strain (MPP). The concept of diabetogenic means the ability of the MPP strain to replicate, cause insulitis and dysregulation of the glucose metabolism in the mouse pancreas in vivo. The interaction between the MPP virus strain and insulin producing β-cells was further studied in cell culture using a mouse-derived insulinoma cell line, MIN-6 cells, as an experimental model. The replication of the MPP virus strain was clearly slower in the MIN-6 cells compared to the other tested cell lines. After three days of incubation, extensive replication of MPP was evident in MIN-6 cells and resulted in a MIN-6 cell-adapted virus strain (MCA). Secondly, the ability of the D207 virus strain, isolated from a type 1 diabetic patient, to replicate in a primary human β-cell culture was tested. D207 was initially serotyped as coxsackievirus A9 (CV-A9) in a virus-specific neutralization assay. The D207 virus strain was found to cause cytolysis in the primary human β-cells and, simultaneously, severe functional damage of the surviving β-cells. The genomes of the four virus strains DS, MPP, MCA and D207 were cloned and sequenced. The sequence comparison of three CV-B5 strains (DS, MPP, and MCA) revealed only limited changes, three capsid and two non-structural (NS) amino acid substitutions between MPP and DS, and two capsid and six NS amino acid substitutions between MCA and MPP. In order to determine which of the amino acid substitutions were responsible for the changed phenotype in vivo and in vitro, full-length infectious clones were constructed from the MPP virus and its parental DS virus. By using reverse mutagenesis and chimeric viruses (MPP/DS and DS/MPP), it was shown that a change from MPP to the MCA phenotype in MIN-6 cells was mediated by only a single amino acid at position 94 in VP1, while the in vivo adaptation of the DS virus strain to the inflammation-inducing MPP virus strain may require multiple genetic determinants in the virus capsid and probably also in the NS proteins. Sequence analyses of D207 revealed that the virus belonged to a genogroup D of E-11, but was also neutralized with monotypic antisera to CV-A9. The isolate D207 was found to be closely related to a specific E-11 strains known to cause uveitis. Uveitis-causing E-11 strains were also found to be well neutralized with both CV-A9- and E-11-specific antisera. In a further study, a wide range of E-11 isolates were included to test the observed dual neutralizibility among isolates belonging to the D genogroup. Five of the six studied strains belonging to genogroup D were also neutralized with antisera against coxsackievirus A9 Griggs. The peptide scanning technique was utilized to identify antigenic regions of the capsid proteins of the D207 strain responsible for the observed dual neutralization. Several regions in the capsid of D207 were found to cross-react with an antiserum raised against CV-A9. However, epitopes responsible for the cross-neutralization remained unidentified. In conclusion, these studies indicate that the specific location of mutation may affect the phenotype of an enterovirus more than the overall quantity of changes. In the experimental settings, radical changes in the viral phenotypic features occurred only after a few amino acid substitutions. The majority of the studied viruses in the genogroup D of E-11 maintained exceptional phenotypic property, the cross-neutralization with CV-A9 specific antiserum, despite their genetic divergence.