Skip to main content

LUP Student Papers

LUND UNIVERSITY LIBRARIES

GDNF and GFR⍺1 Overexpression via Viral Vector as a Gene Therapy Approach for Epilepsy

Suravarapu, Gnapika (2025) MOBN03 20242
Degree Projects in Molecular Biology
Abstract
Epilepsy affects a significant portion of people worldwide and still requires alternative treatments due to resistance to anti-seizure medications. Gene therapy offers a promising alternative with several targets already established. Of these, glial cell line-derived neurotrophic factor (GDNF) and its receptor GFR⍺1 are well recognised for their roles in enhancing inhibitory signalling and supporting neuronal survival. This study aimed to investigate whether an AAV viral vector encoding GDNF and GFRA1 could effectively overexpress these targets in the hippocampus of injected mice. Quantitative PCR and Western blot analysis confirmed increased mRNA and protein levels. Transfected HEK293 cells also revealed differences in expression due to... (More)
Epilepsy affects a significant portion of people worldwide and still requires alternative treatments due to resistance to anti-seizure medications. Gene therapy offers a promising alternative with several targets already established. Of these, glial cell line-derived neurotrophic factor (GDNF) and its receptor GFR⍺1 are well recognised for their roles in enhancing inhibitory signalling and supporting neuronal survival. This study aimed to investigate whether an AAV viral vector encoding GDNF and GFRA1 could effectively overexpress these targets in the hippocampus of injected mice. Quantitative PCR and Western blot analysis confirmed increased mRNA and protein levels. Transfected HEK293 cells also revealed differences in expression due to changes in plasmid orientation. Immunohistochemistry further revealed localised GDNF expression in targeted brain regions. Although we encountered some unexpected results in gene expression and localisation, we hope these findings can be used to optimise viral vector design for status epilepticus (SE) mouse models. (Less)
Popular Abstract
Gene Therapy for Epilepsy with GDNF and GFRA1

Epilepsy is a neurological disorder that affects approximately 1% of the global population. It is characterized by imbalances in the electrical activity of neuronal networks leading to seizure onset. Although several anti-seizure medications have been developed over the years there is still a need for a long term, effective treatments. Thus, gene therapy emerges as a promising mode of treatment.

Successful application of gene therapy requires identifying an appropriate target and an effective delivery system. Targets for epilepsy include neurotransmitters, ion channels, neurotropic factors and their receptors. Glial cell-derived neurotrophic factor (GDNF) is a vital protein heavily... (More)
Gene Therapy for Epilepsy with GDNF and GFRA1

Epilepsy is a neurological disorder that affects approximately 1% of the global population. It is characterized by imbalances in the electrical activity of neuronal networks leading to seizure onset. Although several anti-seizure medications have been developed over the years there is still a need for a long term, effective treatments. Thus, gene therapy emerges as a promising mode of treatment.

Successful application of gene therapy requires identifying an appropriate target and an effective delivery system. Targets for epilepsy include neurotransmitters, ion channels, neurotropic factors and their receptors. Glial cell-derived neurotrophic factor (GDNF) is a vital protein heavily involved in the central nervous system and is an important regulator of excitatory and inhibitory signals in neurons. Its receptor GFRA1 is also involved in signalling and downstream processes. We also know that overexpression of these targets leads to downregulation of excitatory signals in neurons leading to reduced seizures.

As a mode of delivery, we use an adeno-associated virus (AAV). The viral vector was created by inserting the target sequences and was then injected into mice hippocampi. RNA analysis was done with primers targeting GDNF and GFRA1. Protein analysis was done with GDNF and GFRA1 antibodies in Western Blot.

We used two viruses for injection in the mice, one vector with only GDNF and another with GFRA1 and GDNF in that order. Our RNA analysis with qPCR revealed unexpected results, we observed a few inconsistencies across the different primer pairs although we were able to confirm overexpression of the target mRNA in the injected mice hippocampus compared to the uninjected side. Our protein analysis with western blotting revealed a clear overexpression of GDNF and GFRA1 in the injected side of the mice hippocampus. We also performed immunohistochemistry with GDNF antibodies to visualise the localisation of GDNF in the hippocampus of virus injected mice, while it does require further optimisation, we were able to see differences in the viral vectors. Another experiment with mammalian HEK cells was done to determine whether a different orientation of GDNF and GFRA1 in the viral vector would affect the expression levels and GDNF release. We found that having GDNF before GFRA1 lead to better expression of GDNF and higher release.

Our results confirm GDNF and GFRA1 overexpression with this viral vector. The next step would be testing this vector in status epilepticus (SE) models and investigate its seizure mitigation ability.

Master’s Degree Project in Molecular Biology 60 credits 2025
Department of Biology, Lund University

Advisor: Marco Ledri
Epilepsy Center, Department of Clinical Sciences (Less)
Please use this url to cite or link to this publication:
author
Suravarapu, Gnapika
supervisor
organization
course
MOBN03 20242
year
type
H2 - Master's Degree (Two Years)
subject
language
English
id
9214829
date added to LUP
2025-11-03 15:34:08
date last changed
2025-11-03 15:34:08
@misc{9214829,
  abstract     = {{Epilepsy affects a significant portion of people worldwide and still requires alternative treatments due to resistance to anti-seizure medications. Gene therapy offers a promising alternative with several targets already established. Of these, glial cell line-derived neurotrophic factor (GDNF) and its receptor GFR⍺1 are well recognised for their roles in enhancing inhibitory signalling and supporting neuronal survival. This study aimed to investigate whether an AAV viral vector encoding GDNF and GFRA1 could effectively overexpress these targets in the hippocampus of injected mice. Quantitative PCR and Western blot analysis confirmed increased mRNA and protein levels. Transfected HEK293 cells also revealed differences in expression due to changes in plasmid orientation. Immunohistochemistry further revealed localised GDNF expression in targeted brain regions. Although we encountered some unexpected results in gene expression and localisation, we hope these findings can be used to optimise viral vector design for status epilepticus (SE) mouse models.}},
  author       = {{Suravarapu, Gnapika}},
  language     = {{eng}},
  note         = {{Student Paper}},
  title        = {{GDNF and GFR⍺1 Overexpression via Viral Vector as a Gene Therapy Approach for Epilepsy}},
  year         = {{2025}},
}