The goals of the proposed research are to determine the role of GABAA/benzodiazepine receptors in the development of seizure activity in animal models of inherited epilepsy and to determine whether changes in specific amino acid residues of the receptors or in regulation of expression of unchanged receptors are responsible for any such role. The major approach will involve cloning the receptors from brains of inbred rodent strains chosen on the basis of their susceptibility (SP, seizure prone) or resistance (SR, seizure resistant) to various forms of seizure activity. Gammaaminobutyric acid (GABA) is the major inhibitory neurotransmitter of the brain. Its receptor, which includes a chloride channel as part of its structure, is thought to be the site of action of a number of important anticonvulsant drugs, including benzodiazepines and barbiturates, and certain convulsants. Blockade of GABA receptor function causes seizures and its enhancement prevents them. Recent evidence for changes in GABA-related synaptic markers, including receptor binding, in several forms of inherited epilepsy in rodents suggests that seizure susceptibility may result, at least in part, from changes in GABA receptor structure or expression. To test this hypothesis, we will prepare cDNA libraries in lambda gt10 vector from brains of SP and SR rodents (initially audiogenic seizure prone mice, DBA/2J, vs. 2 controls, one C57BL/6J and one congenic), screen them with probes representing pieces of the cloned bovine GABA receptor, sequence positive clones, look for linkage of receptor differences, as detected with specific oligonucleotide probes, with seizure susceptibility and other markers in recombinant inbred strains, and examine levels of GABA receptor mRNA in different brain regions by in situ hybridization. Crude mRNA from SP and SR brains, prepared as the first step in making cDNA libraries, will also be injected into Xenopus oocytes for electrophysiological studies by patch clamp and intracellular microelectrode techniques. This should reveal any differences in GABA receptor function between SP and SR mice and also give data for later comparison with that obtained using artificial mRNAs prepared from cloned cDNAs, verifying the cloning. These studies may eventually lead to gene therapy of inherited epilepsy.

Agency
National Institute of Health (NIH)
Institute
National Institute of Neurological Disorders and Stroke (NINDS)
Type
Research Project (R01)
Project #
5R01NS025525-02
Application #
3410754
Study Section
Neurological Sciences Subcommittee 1 (NLS)
Project Start
1988-07-01
Project End
1991-06-30
Budget Start
1989-07-01
Budget End
1990-06-30
Support Year
2
Fiscal Year
1989
Total Cost
Indirect Cost
Name
University of Maryland Baltimore
Department
Type
Schools of Medicine
DUNS #
003255213
City
Baltimore
State
MD
Country
United States
Zip Code
21201
Cestari, I N; Liu, Z F; Mu, W et al. (1998) GABA(A) receptor alpha4 subunit in DBA/2J and C57BL/6J mice. Brain Res Bull 47:643-7
Wang, J B; Liu, Z F; Kofuji, P et al. (1998) The GABA(A) receptor gamma1-subunit in seizure prone (DBA/2) and resistant (C57BL/6) mice. Brain Res Bull 45:421-5
Liu, Z F; Kamatchi, G L; Moreira, T et al. (1998) The alpha5 subunit of the murine type A GABA receptor. Brain Res Mol Brain Res 59:84-9
Kamatchi, G L; Kofuji, P; Wang, J B et al. (1995) GABAA receptor beta 1, beta 2, and beta 3 subunits: comparisons in DBA/2J and C57BL/6J mice. Biochim Biophys Acta 1261:134-42
Uchida, I; Kamatchi, G; Burt, D et al. (1995) Etomidate potentiation of GABAA receptor gated current depends on the subunit composition. Neurosci Lett 185:203-6
Kubek, M J; Knoblach, S M; Sharif, N A et al. (1993) Thyrotropin-releasing hormone gene expression and receptors are differentially modified in limbic foci by seizures. Ann Neurol 33:70-6
Cestari, I N; Albuquerque, E X; Burt, D R (1993) A PCR shortcut to oocyte expression. Biotechniques 14:404-7
Wang, J B; Kofuji, P; Burt, D R (1992) Strain comparisons and developmental profile of the delta subunit of the murine GABAA receptor. Brain Res Bull 29:119-23
Wang, J B; Kofuji, P; Fernando, J C et al. (1992) The alpha 1, alpha 2, and alpha 3 subunits of GABAA receptors: comparison in seizure-prone and -resistant mice and during development. J Mol Neurosci 3:177-84
Wang, J B; Burt, D R (1991) Differential expression of two forms of GABAA receptor gamma 2-subunit in mice. Brain Res Bull 27:731-5

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