Abstracts

Rationale:
GABAB receptors are G-protein coupled receptors expressed in both presynaptic and postsynaptic neurons and contribute to GABAergic signaling by activation of postsynaptic K+ channels, reduction of presynaptic GABA release, and modulation of intracellular signaling by Gi/o. Although traumatic brain injury (TBI) is well-established to affect signaling by GABAA receptors in hippocampus, the effects of TBI on expression and function of GABAB receptors are incompletely understood.
Method:
Acute hippocampal brain slices were prepared from C57Bl/6 mice for patch-clamp recordings from dentate gyrus granule cells (DGGCs) and field potential recordings in the presence of 4-aminopyridine (4AP). Controlled cortical impact (CCI) model was used to generate experimental TBI (2 mm impactor tip, 1 mm deformation in cortical surface).
Results:
We measured membrane currents evoked by focal application of the GABAB receptor agonist baclofen from dentate gyrus granule cells (DGGCs); these currents were sensitive to the GABAB receptor antagonist CGP55845 (CGP), the K+ channel blocker Ba2+, and reversed direction near the equilibrium potential for K+ ions. Thus, baclofen was acting on GABAB receptors to activate inwardly-rectifying K+ channels (Kir). Two weeks after CCI, baclofen-induced currents in DGGCs were reduced by 55% compared to control/sham DGGCs (p< 0.05). This reduction was limited to DGGCs ipsilateral to CCI and was persistent at 2 months after CCI. To assess functional consequences of GABAB receptor downregulation after CCI, we measured spontaneous field potential discharges induced by 4AP (100 μM) to compare the inhibitory effect of baclofen on 4AP-induced activity between control and CCI slices (Siniscalchi and Avoli, 1992). We hypothesized that GABAB receptor downregulation after CCI would modify 4AP-induced activity and reduce the sensitivity to baclofen. Baclofen produced a dose-dependent reduction in frequency of spontaneous field potential discharges measured in area CA1, but did not affect the amplitude or duration of individual discharges. The baclofen effect was sensitive to CGP and the K+ channel blocker Ba2+, indicating the inhibitory effect of baclofen was mediated by GABAB receptor activation of Kir channels.  The GABAA receptor antagonist gabazine, at concentrations selective for synaptic receptors (i.e. 100 nM), did not modify the characteristics of 4AP-induced field potentials nor the inhibitory effect of baclofen. This result indicates that the inhibitory effect of baclofen was not caused by activation of presynaptic GABAB receptors and reduction of vesicular GABA release. CCI did not significantly affect the amplitude, duration, or frequency of spontaneous discharges induced by 4AP compared to control slices. In contrast to our hypothesis, the baclofen-sensitivity of field potential frequency was also not altered by CCI. Baclofen (1 μM) reduced frequency of 4AP-induced discharges by 67.5% and 68.2% in control and CCI-ipsilateral slices, respectively (p=0.92).
Conclusion:
Our results indicate GABAB receptor dysfunction in DGGCs after CCI. However, GABAB receptor dysfunction in DGGCs after CCI did not significantly impact 4AP-induced activity nor the inhibitory actions of baclofen. Our data indicate that Ba2+-sensitive K+ channels activated by GABAB receptors are essential for the inhibitory action of baclofen in this model. Ongoing work is examining the baclofen-sensitivity of 4AP-induced and electrically-evoked field potentials within the dentate gyrus itself.  
Funding:
:This work was supported by Merit Review I01BX002745-0 from the United States Department of Veterans Affairs (BLR&D) (CR).