Reduced function of γ-aminobutyric acidA receptors in tottering mouse brain: Role of cAMP-dependent protein kinase

✍ Scribed by Mohammad H. Jalilian Tehrani; Eugene M. Barnes Jr.

• Publisher: Elsevier Science
• Year: 1995
• Tongue: English
• Weight: 711 KB
• Volume: 22
• Category: Article
• ISSN: 0920-1211
• DOI: 10.1016/0920-1211(95)00033-7

✦ Synopsis

The single-locus mutant mouse tottering (tg) displays spontaneous seizures that resemble those in human petit-mal epilepsy. In order to examine alterations in GABA A receptor function which could arise as a result of this mutation, the influx of 36C1 was determined using microsacs (membrane vesicles) isolated from the brain of tg/tg and coisogenic C57BL/6J (+/+ ) control mice. In microsacs from both tg/tg and +/+ strains, the maximum level of 36C1uptake induced by 50/xM GABA was observed during five seconds of incubation at 28°C. Compared to + / +, the GABA-dependent 36C1uptake in tg/tg microsacs was significantly lower and faded rapidly during longer incubations. The levels of gated 36C1uptake in tg/tg microsacs were 45 + 6.3%, 65 + 9.9%, and 33 ___ 6.1% of control (+/+) values for 3-, 5-, and 10-s incubations, respectively. GABA A receptor-specific agonists (30 /~M), muscimol, isoguvacine and THIP (4,5,6,7-tetrahydroisoazolo-[5,4-c]pyridin-3-ol) induced 36C1-influx in the order muscimol > GABA > isoguvacine > THIP.

This order was similar for both strains, but the agonist-dependent influx was always significantly lower in tg/tg compared to + / +. Treatment of the microsacs with 10 ~M H-89, a membrane-permeant inhibitor of the cAMP-dependent protein kinase (protein kinase A, PKA), was without effect on GABA-gated 36C1uptake in + / +, but increased the gated uptake in tg/tg microsacs by 44 + 16%. PKA was assayed using [3~-32]ATP and kemptide as the substrate. Triton X-100 (0.1%) increased both the basal and 8-Br-cAMP dependent PKA activity in microsacs by 3-4 four fold, showing that most of the enzyme was intravesicular. In the presence of Triton, the basal activity of PKA in the tg / tg preparations was twice that of +/+, while the strain difference was no longer apparent in assays containing 8-Br-cAMP. The data suggest that an abnormal elevation of protein kinase A activity in tottering mouse brain contributes to an impairment of GABA A receptor function. It is suggested that the resulting loss of inhibition could play a role in induction of the seizures which characterize the mutant phenotype.