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Attempt to trace midbrain dopaminergic neurons

Yang, Fan (2012) MOBT29 20112
Degree Projects in Molecular Biology
Abstract
Abstract

Dopamine as a neurotransmitter was discovered in the late 1950s. The dopaminergic system is one of the most well studied neurotransmitter systems in the brain. It is involved in many important functions, such as motor function control; reward, reinforcement, motivation, associative learning, salience information encoding; cognition etc. Disturbances of the dopaminergic system may result in motor disorders such as hypokinesia, dyskinesia, chorea etc; behavioural or emotional abnormalities such as drug addiction, hallucination, delusion, inappropriate affect, heightened distractibility etc; cognitive impairments etc. One of the most straightforward ways to further understand the functions of the dopaminergic system is to look in... (More)
Abstract

Dopamine as a neurotransmitter was discovered in the late 1950s. The dopaminergic system is one of the most well studied neurotransmitter systems in the brain. It is involved in many important functions, such as motor function control; reward, reinforcement, motivation, associative learning, salience information encoding; cognition etc. Disturbances of the dopaminergic system may result in motor disorders such as hypokinesia, dyskinesia, chorea etc; behavioural or emotional abnormalities such as drug addiction, hallucination, delusion, inappropriate affect, heightened distractibility etc; cognitive impairments etc. One of the most straightforward ways to further understand the functions of the dopaminergic system is to look in even greater detail on the anatomy with the help of the rapidly advancing techniques. In this study, we try to label large amount of dopaminergic neurons with omputerdistinguishable colours by delivering three primary colours of randomly determined intensities to the these neurons, trace their axonal projections and thus further illustrate the possible patterns of interaction among these dopaminergic neurons or the fibres arising from them. We generated new Brainbow transgenic mice and examined the expression levels and patterns of the fluorescent proteins coding the three primary colours. We found there was only very weak or no fluorescence expression at either the substantia nigra/ventral tegmental area, or the other parts of the brain or the other parts of the whole mouse body. In addition to the Brainbow transgenic method, we have also adopted a viral gene delivery approach. We prepared different pAAV-stop-XFP plasmids and tested them in HEK 293 cells. We found all the XFPs could function properly and the transcriptional stop cassette would be strict enough to restrain the expression of XFPs. From the preliminary results, we conclude that our new Brainbow mice are not applicable for the purpose of labelling and tracing dopaminergic neurons. The viral gene delivery approach is still under development, with a promising outlook.

Popular science summary:

Dopamine as a neurotransmitter was discovered in the late 1950s. The dopaminergic system is one of the most well studied neurotransmitter systems in the brain. It is involved in many important functions, such as motor function control, reward, motivation and cognition etc. Disturbances of the dopaminergic system may result in motor disorders such as those related to Parkinson disease; behavioural or emotional abnormalities such as drug addiction, hallucination, delusion, inappropriate affect, heightened distractibility; cognitive impairments etc.

One of the most straightforward ways to analyse the health state of the dopaminergic system is to look in greater detail on the anatomy with the help of the rapidly advancing techniques. In this study, we try to label dopaminergic neurons and their fibres with computer-distinguishable colours. The idea is to deliver three primary colours of randomly determined intensities to the dopaminergic neurons. By the same principle that three primary colours make a television screen colourful, if our approach is successful, then the dopaminergic system in the midbrain could also be illuminated colourful. Consequently, this would allow us to trace the fibre projections arising from different dopaminergic neurons.

We generated transgenic mice to express different fluorescent proteins as the three primary colours. We examined the expression levels and patterns of these fluorescent proteins. We found there was only very weak or no fluorescence expression in the brain or at the other parts of the mouse body. In addition to the transgenic method, we have also adopted a viral gene delivery approach. In this approach, the three primary colours would still be expressed by different fluorescent proteins. The difference from the transgenic approach is that we try to bring the three primary colours into the brain by engineered viruses. These viruses are safe to use because they have lost their pathogenic ability. From the preliminary results, we conclude that our transgenic mice are not applicable for the purpose of labelling and tracing midbrain dopaminergic neurons. The viral gene delivery approach is still under development, with a promising outlook.

If the basic tracing is successful, then we would further trace dopaminergic neurons under different conditions, e.g. during development or during ageing in normal (wild type) or abnormal (genetically engineered) mice. We could also label other types of neurons so that we can study cell-cell interactions. The potential findings in these studies may possibly indicate new functions of certain brain regions, certain cell types, and/or certain biochemical molecules. More, such as pharmacological, behavioural etc studies could be conducted even further based on these potential findings.



Advisor: Edgar Kramer, Center for Molecular Neurobiology Hamburg, Germany
Master´s Degree Project 60 credits in Neurobiology, 2011-2012
Department of Biology, Lund University (Less)
Please use this url to cite or link to this publication:
author
Yang, Fan
supervisor
organization
course
MOBT29 20112
year
type
H2 - Master's Degree (Two Years)
subject
language
English
id
3799768
date added to LUP
2013-05-23 15:26:48
date last changed
2013-05-23 15:26:48
@misc{3799768,
  abstract     = {Abstract

Dopamine as a neurotransmitter was discovered in the late 1950s. The dopaminergic system is one of the most well studied neurotransmitter systems in the brain. It is involved in many important functions, such as motor function control; reward, reinforcement, motivation, associative learning, salience information encoding; cognition etc. Disturbances of the dopaminergic system may result in motor disorders such as hypokinesia, dyskinesia, chorea etc; behavioural or emotional abnormalities such as drug addiction, hallucination, delusion, inappropriate affect, heightened distractibility etc; cognitive impairments etc. One of the most straightforward ways to further understand the functions of the dopaminergic system is to look in even greater detail on the anatomy with the help of the rapidly advancing techniques. In this study, we try to label large amount of dopaminergic neurons with omputerdistinguishable colours by delivering three primary colours of randomly determined intensities to the these neurons, trace their axonal projections and thus further illustrate the possible patterns of interaction among these dopaminergic neurons or the fibres arising from them. We generated new Brainbow transgenic mice and examined the expression levels and patterns of the fluorescent proteins coding the three primary colours. We found there was only very weak or no fluorescence expression at either the substantia nigra/ventral tegmental area, or the other parts of the brain or the other parts of the whole mouse body. In addition to the Brainbow transgenic method, we have also adopted a viral gene delivery approach. We prepared different pAAV-stop-XFP plasmids and tested them in HEK 293 cells. We found all the XFPs could function properly and the transcriptional stop cassette would be strict enough to restrain the expression of XFPs. From the preliminary results, we conclude that our new Brainbow mice are not applicable for the purpose of labelling and tracing dopaminergic neurons. The viral gene delivery approach is still under development, with a promising outlook.

Popular science summary:

Dopamine as a neurotransmitter was discovered in the late 1950s. The dopaminergic system is one of the most well studied neurotransmitter systems in the brain. It is involved in many important functions, such as motor function control, reward, motivation and cognition etc. Disturbances of the dopaminergic system may result in motor disorders such as those related to Parkinson disease; behavioural or emotional abnormalities such as drug addiction, hallucination, delusion, inappropriate affect, heightened distractibility; cognitive impairments etc.

One of the most straightforward ways to analyse the health state of the dopaminergic system is to look in greater detail on the anatomy with the help of the rapidly advancing techniques. In this study, we try to label dopaminergic neurons and their fibres with computer-distinguishable colours. The idea is to deliver three primary colours of randomly determined intensities to the dopaminergic neurons. By the same principle that three primary colours make a television screen colourful, if our approach is successful, then the dopaminergic system in the midbrain could also be illuminated colourful. Consequently, this would allow us to trace the fibre projections arising from different dopaminergic neurons. 

We generated transgenic mice to express different fluorescent proteins as the three primary colours. We examined the expression levels and patterns of these fluorescent proteins. We found there was only very weak or no fluorescence expression in the brain or at the other parts of the mouse body. In addition to the transgenic method, we have also adopted a viral gene delivery approach. In this approach, the three primary colours would still be expressed by different fluorescent proteins. The difference from the transgenic approach is that we try to bring the three primary colours into the brain by engineered viruses. These viruses are safe to use because they have lost their pathogenic ability. From the preliminary results, we conclude that our transgenic mice are not applicable for the purpose of labelling and tracing midbrain dopaminergic neurons. The viral gene delivery approach is still under development, with a promising outlook. 

If the basic tracing is successful, then we would further trace dopaminergic neurons under different conditions, e.g. during development or during ageing in normal (wild type) or abnormal (genetically engineered) mice. We could also label other types of neurons so that we can study cell-cell interactions. The potential findings in these studies may possibly indicate new functions of certain brain regions, certain cell types, and/or certain biochemical molecules. More, such as pharmacological, behavioural etc studies could be conducted even further based on these potential findings.
	


Advisor: Edgar Kramer, Center for Molecular Neurobiology Hamburg, Germany
Master´s Degree Project 60 credits in Neurobiology, 2011-2012
Department of Biology, Lund University},
  author       = {Yang, Fan},
  language     = {eng},
  note         = {Student Paper},
  title        = {Attempt to trace midbrain dopaminergic neurons},
  year         = {2012},
}