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Application of optogenetic techniques to study beta-cell function

Safi, Fatemeh (2013) MOBN15 20122
Degree Projects in Molecular Biology
Popular Abstract
Insulin is the only blood glucose reducing hormone and is produced by beta-cells in pancreatic islets. Malfunctioning beta-cells are the main cause for developing type 2 diabetes. For proper insulin release to occur, the cell depends on intracellular calcium signals that can be modulated by adrenenaline receptors. The role of the various adrenergic receptor subtypes in pancreatic beta-cells is poorly understood as they are not only present in beta-cells but also in other pancreatic islet cell types. A specific targeting method for studying and isolating the effect of this type of receptor in beta-cells is needed. In other fields of biology like neurobiology, specific targeting has been achieved by using a new technique called optogenetics.... (More)
Insulin is the only blood glucose reducing hormone and is produced by beta-cells in pancreatic islets. Malfunctioning beta-cells are the main cause for developing type 2 diabetes. For proper insulin release to occur, the cell depends on intracellular calcium signals that can be modulated by adrenenaline receptors. The role of the various adrenergic receptor subtypes in pancreatic beta-cells is poorly understood as they are not only present in beta-cells but also in other pancreatic islet cell types. A specific targeting method for studying and isolating the effect of this type of receptor in beta-cells is needed. In other fields of biology like neurobiology, specific targeting has been achieved by using a new technique called optogenetics. Optogenetics makes use of light sensitive membrane proteins (including receptors and ion channel)s that can be specifically targeted to cells and activated by light.

Here we wanted to use this technique to stimulate beta-cell activity. We chose INS-1 832/13 cells, a rat beta-cell line that is commonly used as a model cell line to study glucose-induced insulin secretion. We used light-activatable light-sensitive receptor and ion channel constructs in these cells and tested different techniques to induce their expression and production in INS-1 832/13 cells.

We could successfully express light-sensitive constructs in INS-1 832/13 cells by lipofection but not by calcium precipitation or virus-mediated methods. The latter often caused cell aggregation and induced cell death. The method of choice, lipofection, induced prominent expression and production of our constructs, although in few cells. Using fluorescent microscopy and calcium imaging, we verified the functionality of these light-sensitive receptors and ion channels. By stimulating our cells with light, we observed increased intracellular calcium levels, although specificity has to be verified in additional experiments. If the observed increases in intracellular calcium also are able to induce insulin release, will be interesting to assess in future studies.

We conclude that optogenetics may be a promising approach to study beta-cell function.

Supervisor: Dr. Thomas Reinbothe, Department of Clinical Sciences Malmö, Medical Faculty, Lund University
Master´s Degree Project 45 credits in Cell and Molecular Biology, 2013
Department of Biology, Lund University (Less)
Please use this url to cite or link to this publication:
author
Safi, Fatemeh
supervisor
organization
course
MOBN15 20122
year
type
H2 - Master's Degree (Two Years)
subject
language
English
id
5432723
date added to LUP
2015-05-26 10:16:01
date last changed
2015-05-26 10:16:01
@misc{5432723,
  author       = {Safi, Fatemeh},
  language     = {eng},
  note         = {Student Paper},
  title        = {Application of optogenetic techniques to study beta-cell function},
  year         = {2013},
}