Dibutyryl-cAMP Attenuates the Actions of Topiramate on Recombinant [alpha][sub]2[/sub][beta][sub]3[/sub][gamma][sub]2S[/sub] GABA[sub]A[/sub] Receptors Expressed in [italic]Xenopus[/italic] Oocytes.
Abstract number :
1.036
Submission category :
Year :
2001
Submission ID :
678
Source :
www.aesnet.org
Presentation date :
12/1/2001 12:00:00 AM
Published date :
Dec 1, 2001, 06:00 AM
Authors :
T.A. Simeone, Pharmacology and Toxicology, University of Utah, Salt Lake City, UT; H.S. White, PhD, Pharmacology and Toxicology, University of Utah, Salt Lake City, UT
RATIONALE: Topiramate (TPM) is a novel anticonvulsant that affects the responses of several ion channels including GABA[sub]A[/sub] receptors. TPM has exhibited varying results in several different experimental GABA[sub]A[/sub] receptor model systems. Since function may be altered by the phosphorylation state of the receptor, we hypothesized that kinase activity may modulate the effect of TPM on GABA[sub]A[/sub] receptor currents.
METHODS: [italic]Xenopus laevis[/italic] oocytes were nuclear injected with cDNAs encoding the [alpha][sub]2[/sub][beta][sub]3[/sub][gamma][sub]2S[/sub] GABA[sub]A[/sub] receptor subunits. Experiments were performed 48-96 hours after injection. Whole-cell currents were recorded using two-electrode voltage clamp techniques and data was acquired with pClamp6 software. After establishing control responses, 100 [mu]M dibutyryl-cAMP, a cell permeable cyclic AMP analog that preferentially activates cAMP-dependent protein kinase, was bath applied over the oocytes for 30 minutes. Current responses were elicited every three minutes by bath application of GABA (EC[sub]1[/sub]), TPM (1 mM), GABA+TPM (100 [mu]M), or GABA+TPM (1 mM). The oocytes were subsequently washed with normal Ringer[ssquote]s solution for 20 minutes to assess the reversibility of the observed effects. Although 1 mM TPM is above the therapeutic range, it was used to examine dibutyryl-cAMP modulation of TPM direct activation of the receptor.
RESULTS: Exposure to dibutyryl-cAMP attenuated GABA- and TPM (1 mM)-mediated currents by 19% and 28%, respectively. Enhancement of GABA-mediated currents by 100 [mu]M and 1 mM TPM was reduced by 71% and 37%, respectively. These effects of dibutyryl-cAMP were rapid in onset (within 3 minutes of initial application) and were rapidly reversed upon washout of dibutyryl cAMP.
CONCLUSIONS: These results suggest that direct activation of the [alpha][sub]2[/sub][beta][sub]3[/sub][gamma][sub]2S[/sub] GABA[sub]A[/sub] receptor by GABA and TPM and TPM enhancement of the GABA-mediated current may be modulated by PKA phosphorylation of the receptor. Future studies will assess the effect of kinase (PKA and PKC) activators and inhibitors on the GABA and TPM responses of the GABA[sub]A[/sub] receptor.
Disclosure: Grant - R.W. Johnson (H.S. White); Consulting - R.w. Johnson (H.S. White).