Publikationsserver der RWTH Aachen University
Genetische Inaktivierung des hyperpolarisations-aktivierten Ionenkanals HCN 4
Abstract
dc:descriptionThe present work describes the development of knockout models for the genetic inactivation of the hyperpolarization-activated channel HCN4. The HCN4-subunit is expressed in the sinoatrial node of the heart, where it is probably involved in the generation of cardiac pacemaking. As a consequence, HCN4-deficient mice (conventional knockout), might be lethal. Therefore, in addition to the conventional knockout model, a conditional knockout model was developed. A conditional knockout provides the possibility to inactivate the gene in a selected tissue or at a selected developmental stage. The Cre/loxP-based strategy was used to generate the conditional knockout model. As a prerequisite for the design of the targeting vectors the genomic structure of the mouse HCN4 gene was identified. The HCN4 gene is about 35 kbp and is located on chromosome 9. The locus contains eight exons. This information allowed to design targeting vectors for both conventional and conditional knockout of the HCN4-channel-subunit. These targeting vectors were transferred to embryonic stem cells (ES cells) by electroporation. Positive clones were selected for their antibiotics resistance. Conventional knockout: the DNA of the antibiotics resistent ES cell clones was analysed by southernblotting. In two of these clones, the vector was integrated in the HCN4 gene in the correct orientation. These clones were implantated into blastocysts, which were injected into pseudo-pregnant mice. Chimeras (+/-) were born that showed a normal phenotype. They were bred with wildtype mice to get pure heterozygote mice. By the time this thesis was submitted, the heterozygote mice had not been born. The heterozygote mice will have to be bred to get homozygote HCN4-deficient (-/-) mice. Conditional knockout: The analysis of the ES cell clones has not been completed yet. At present we can only speculate about the effect of the genetic inactivation of the HCN4-channel. The regulation of rhythmic activity in the thalamus and in the sinoatrial node of the heart is regulated by HCN-channels. In both tissues HCN4 is expressed together with other HCN-subunits. Especially in the thalamus, HCN-currents seem to be dominated by the HCN4-subunit. Thus in HCN4-deficient mice a strong disturbance in the generation of rhythmic activities is to be exspected.
Degree
thesis:*- Grantor dc:publisher
- Publikationsserver der RWTH Aachen University
- Year dc:date
- 2002
Author and committee
dc:creator, dc:contributor.*- Author dc:creator
-
- Köngeter, Astrid
- Contributors dc:contributor
-
- Kaupp, Ulrich Benjamin
Subjects
dc:subject × 3Rights
dc:rights- Statement dc:rights
-
- info:eu-repo/semantics/openAccess
- Language dc:language
- ger
Identifiers
dc:identifier.*- OAI identifier oai:identifier
- oai:publications.rwth-aachen.de:57205