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Diss. (sammanfattning) Stockholm : Karolinska institutet, 2009.

Thesis (Ph. D.)--University of Chicago, Committee on Neurobiology, August 1999. / Includes bibliographical references. Also available on the Internet.

京都大学 / 新制・課程博士 / 博士(医学) / 甲第23095号 / 医博第4722号 / 新制||医||1050(附属図書館) / 京都大学大学院医学研究科医学専攻 / (主査)教授 伊佐 正, 教授 井上 治久, 教授 渡邉 大 / 学位規則第4条第1項該当 / Doctor of Medical Science / Kyoto University / DFAM

Oxidative stress has been implied in a wide variety of diseases, such as cancer, myocardial infarction, and neurodegenerative diseases including Parkinson's diseases (PD). PD is characterized by the degeneration of dopaminergic (DA) neurons; genetic studies have identified gene mutations causal to PD. Accumulating studies hypothesize that these genes protect DA neurons against oxidative stress. However, lack of experimental tools to target oxidative stress in specific cells has prevented direct evaluation of the hypothesis. We established a novel method to use KillerRed (KR), a genetically-encoded protein that generates radicals upon light activation. We showed its efficacy in live animals by cell-specific ablation of neurons in C. elegans. We applied KR to degenerate DA neurons. By controlling the level of stress via activation light, the protective role of PD-gene, LRRK2, against oxidative stress was confirmed. Thus, we established a method to address the role of oxidative stress in a cell-specific manner.

Oxidative stress has been implied in a wide variety of diseases, such as cancer, myocardial infarction, and neurodegenerative diseases including Parkinson's diseases (PD). PD is characterized by the degeneration of dopaminergic (DA) neurons; genetic studies have identified gene mutations causal to PD. Accumulating studies hypothesize that these genes protect DA neurons against oxidative stress. However, lack of experimental tools to target oxidative stress in specific cells has prevented direct evaluation of the hypothesis. We established a novel method to use KillerRed (KR), a genetically-encoded protein that generates radicals upon light activation. We showed its efficacy in live animals by cell-specific ablation of neurons in C. elegans. We applied KR to degenerate DA neurons. By controlling the level of stress via activation light, the protective role of PD-gene, LRRK2, against oxidative stress was confirmed. Thus, we established a method to address the role of oxidative stress in a cell-specific manner.

The involvement of dopamine (DA) in the emotional and psychiatric disturbances associated with schizophrenia and psychomotor stimulant abuse is well known; however, the mechanism by which DA mediates fear expression and anxiety is not well defined. Accordingly, the objective of the present thesis was to determine the fear-motivational functions of DA neurons in the ventral tegmental area (VTA) and to examine the role of DA in fear extinction using the potentiated startle paradigm. In Experiment 1, it was observed that electrical stimulation of the VTA produced a pronounced increase in the amplitude of the acoustic startle reflex. In subsequent experiments fear-potentiated startle was assessed following axon-sparing N-methyl-D-aspartic acid (NMDA) lesions of the VTA and after bilateral intra-VTA infusion of the DA D_2/3 receptor agonist quinpirole (Experiments 2-4). The NMDA lesions resulted in substantial cell loss in the medial ventral tegmentum and blocked fear-potentiated startle. Similarly, inhibition of DA neuronal activity associated with locally-administered quinpirole suppressed the expression of the conditioned fear-induced increase in startle amplitudes. The quinpirole results implicate DA neuronal functioning in fear motivation. To explore further the involvement of DA in aversive emotional behavior, pharmacological experiments were conducted in which the effects of peripherally-administered DA agonist drugs on fear extinction were assessed. Subjects in Experiment 5 received an acute injection of either cocaine hydrochloride (40.0 mg/kg), d-amphetamine sulphate (5.0 mg/kg), the D_2/3 agonist quinpirole hydrochloride (5.0 mg/kg), or the D₁-type agonist SKF 38393 (5.0 mg/kg) during a single extinction session following fear acquisition. Animals treated with cocaine, d-amphetamine, and SKF 38393 exhibited fear-potentiated startle, whereas quinpirole treatment failed to alter fear extinction to the nonreinforced conditioned stimulus (CS). Also, it was revealed using a within-subjects design in Experiment 6 that cocaine administration reinstated fear-potentiated startle following extinction. Taken together, the results of the present experiments suggests fundamental role for DA and DA D₁ receptors in fear expression. It was proposed that VTA DA neurons gate levels of aversive emotional arousal within the amygdala-based fear system.

Environmental experiences play a critical role in an individualʼs risk of becoming addicted. Positive experiences may mitigate addiction vulnerability, whereas adverse experiences, particularly during adolescence, have been shown to increase addiction risk. Social isolation in rodents is a model system used to study the effects of such experiences, yet its impact on the learning and memory processes that underlie addiction remains elusive. Although social isolation is known to alter the functioning of the dopaminergic system, as well as reward processing and learning, its effect on dopamine (DA) neurons of the ventral tegmental area (VTA) is unknown. The data presented in this dissertation demonstrate that social isolation of rats during a critical period in adolescence (postnatal days 21-42) enhances long-term potentiation (LTP) of N-methyl D-aspartate receptor (NMDAR)-mediated glutamatergic transmission in the VTA. Activation of NMDARs is critical to the generation of DA neuron bursts that encode rewards and reward-predictive cues, and NMDARs are necessary for associative reward learning. The isolation-induced enhancement of NMDAR LTP results from augmentation of metabotropic glutamate receptor (mGluR)-dependent calcium (Ca²⁺) signaling via an increase in inositol 1,4,5-trisphosphate(IP3) sensitivity. Isolation-mediated effects on Ca²⁺ signaling and NMDAR plasticity were not reversed by a subsequent period of resocialization. Furthermore, social isolation during this critical period occludes the effect of repeated amphetamine exposure on mGluR/IP₃-mediated Ca²⁺ signaling and synaptic plasticity. Although corticotropin releasing factor (CRF) further facilitates mGluR/IP3-mediated Ca²⁺ signaling in DA neurons, alterations in CRF receptors are not responsible for the effects of isolation on Ca²⁺ signaling and synaptic plasticity. In addition, the learning of associations between environmental stimuli and drug rewards is acquired more quickly and is more resistant to extinction in isolated animals. Data presented in this dissertation lend support to the theory that enhanced mGluR/IP₃-mediated Ca²⁺ signaling and NMDAR plasticity facilitate the learning and memory of drug-associated stimuli. This dissertation provides the first demonstration of a cellular basis for the critical time window of social isolation during adolescence. NMDAR plasticity in the VTA may thus represent a neural substrate by which early life experiences regulate addiction vulnerability. (Note: Behavioral data were acquired by Mickael Degoulet) / text

In this study we aimed to investigate the effects of 1-methyl-4-phenyl-1,2,3,6- tetrahydropyridine (MPTP) and rotenone neurotoxins on dopaminergic (DAergic) neuronal survival using ventral mesencephalic (VM) organotypic cell culture derived from postnatal rat pups (P4-5) and immunocytochemistry for tyrosine hydroxylase (TH) as a marker of DAergic cells. In addition, we examined the neuroprotective effects of glial cell line-derived neurotrophic factor (GDNF) on TH-ir cells exposed to MPTP and rotenone as a possible treatment for PD. The TH-ir cells in co-cultures with striatum (ST) as a target grew better then when VM was cultured alone and that TH-ir cells in co-cultures could be maintained without using conditioned and trophic media. We treated 7 day and 14 day co-cultures at different times with varying MPTP and rotenone concentrations and found 14 day old cultures were more vulnerable than 7 day old co-cultures to the effects of either neurotoxin with TH-ir cell numbers significantly lower in 14 day cultures compared to 7 day cultures. Both neurotoxins induced a dose-dependent TH-ir cell reduction in the co-cultures. In addition we compared the toxicity of MPTP and its active metabolite 1-methyl-4- phenylpyridinium (MPP+) as the neurotoxic effects of MPTP on DAergic cells depends on its conversion to MPP+ by astrocytes. We found no significant difference in TH-ir cell reduction in co-cultures treated with MPTP and MPP+. Rotenone was more toxic than MPTP with less TH-ir cell survival in the weeks post treatment. GDNF exposure produced increased cell size and significant increases in TH-ir cell branching in cocultures in a dose-dependent manner. Post treatment of GDNF against MPTP and rotenone provided significant neuroprotection as TH-ir cell survival was at the lower neurotoxin doses and not at the higher doses. / http://proxy.library.adelaide.edu.au/login?url= http://library.adelaide.edu.au/cgi-bin/Pwebrecon.cgi?BBID=1346931 / Thesis (Ph.D.) - University of Adelaide, School of Medical Sciences, 2008

Diss. (sammanfattning) Stockholm : Karol inst., 2004. / Härtill 5 uppsatser.