Upregulation of GABA neurotransmission suppresses hippocampal excitability and prevents long-term potentiation in transgenic superoxide dismutase-overexpressing mice

Abstract

Cu/Zn superoxide dismutase (SOD-1) is a key enzyme in oxygen metabolism in the brain. Overexpression of SOD-1 in transgenic (Tg) mice has been used to study the functional roles of this enzyme in oxidative stress, lipid peroxidation, and neurotoxicity. We found that Tg-SOD-1 mice are strikingly less sensitive to kainic acid-induced behavioral seizures than control mice. Furthermore, the hippocampus of Tg-SOD-1 mice was far less sensitive to local application of bicuculline, a GABA-A antagonist, than the hippocampus of control mice. GABAergic functions, expressed in extracellular paired-pulse depression, and in IPSCs recorded in dentate granular cells were enhanced in Tg-SOD-1 mice. Finally, long-term potentiation (LTP), not found in the dentate gyrus of Tg-SOD-1 mice, could be restored by local blockade of inhibition and could be blocked in control mice by injection of diazepam, which amplifies inhibition. These results indicate that constitutive elevation of SOD-1 activity exerts a major effect on neuronal excitability in the hippocampus, which, in turn, controls hippocampal ability to express LTP.

Fig. 1.

Behavioral seizures in response to kainic acid in control and Tg-SOD mice. KA (27.5 mg/kg, i.p.) was injected to the mouse, and its behavior was monitored and scored on a scale of 1–5 every 5 min. The results of two lines, SOD 51 and SOD 69, and their corresponding controls are presented separately. Most of the control mice died eventually, whereas all of the SOD mice survived the test.

Fig. 2.

Electrographic epilepsy in anesthetized mice.A, Illustration of seizure activity in the control mouse (left) and the lack of such activity in the Tg-SOD mouse, after an intraperitoneal injection of 20 mg/kg of KA. The identification of epileptic activity was clear-cut, and the mice were scored as having or not having seizures within 10–20 min after the injection. B, Summary of results for the presence of seizure activity in groups of six mice each after (fromleft to right) local application of 1 or 5 mm bicuculline and peripheral application of 10 and 20 mg/kg KA.

Fig. 3.

Reactivity to afferent stimulation is the same in control and Tg-SOD mice. A, Sample illustrations of population responses recorded in the granular layer to stimulation of the perforant path. Stimulus artifact, downward deflection. Population spikes are marked with an arrowhead. B, C, Input–output relations, depicting the changes in population spike (B) and EPSP slope (C) as a function of changes in stimulation intensity, are the same in the two groups of mice. Results of 20 mice in each group, including mice of the two lines, are summarized herein.

Fig. 4.

Differential effect of bicuculline in Tg-SOD and control mice. A, Sample illustration of population response to perforant path stimulation recorded with saline pipette and with 1 or 5 mm bicuculline-containing pipettes. A large increase in population spike is seen in the control mouse, but not in the Tg-SOD mouse. B, C, Summary of input–output relations in the two groups of mice, in the different drug conditions. The control mice were not tested with 5 mm bicuculline, because most of them produced epileptic activity under these recording conditions. A clear increase in population spikes, on the background of small, if any, effect on population EPSPs, is seen.

Fig. 5.

Paired-pulse depression is markedly enhanced in Tg-SOD mice. A, Illustrations of paired-pulse responses to stimulation applied with a 30 msec interpulse interval in control and Tg-SOD mice. Note that the control mouse produced about the same size population spike to the two stimuli, whereas in the Tg-SOD mouse, the second population spike is totally eliminated. B,Summary of the paired-pulse responses in the two groups, using three interpulse intervals, while recording with saline pipette, with bicuculline-containing pipette, or in the KA-injected mice. Note the marked difference between the two groups already in control conditions, and the persistence of the effect also after blockade of inhibition.

Fig. 6.

GABAergic action potential-dependent activity, but not mIPSPs, is enhanced in Tg-SOD mice. A,Examples of activity recorded in a slice obtained from a Tg-SOD mouse (a) and control (b). Traces are continuous recording from voltage-clamped cells in the presence of APV (20 μm) and DNQX (10 μm). In contrast to control, the activity recorded in Tg-SOD mice slices often appeared in clusters of many events. The histograms are the means of representative values of parameters measured in every cell (n = 6 in each group). The mean frequency was significantly higher than that detected in control mice slices (*p = 0.03), whereas the analysis of isolated events did not show any difference in amplitude, area, rise time, and decay time. B, Examples of miniature IPSP recorded in slices from Tg-SOD mice (a) and controls (b). Cells were clamped at −65 mV, and activity was recorded in the presence of TTX (1 mm), APV (20 μm), and DNQX (10 μm). In contrast to action potential-dependent activity, no difference was found in mIPSP frequency (c), as in other parameters such as amplitude, area, rise time, and decay time.

Fig. 7.

LTP is found in control but not Tg-SOD mice.A, Sample illustration of evoked responses before and after tetanic stimulation producing LTP in Tg-SOD and control mice. The numerals are taken at the time depicted in B andC. Tetanic stimulation is applied at thearrow, and a clear difference between control and Tg-SOD mice is seen both with the population spike and the EPSP slopes.

Fig. 8.

Bicuculline restored the ability to express LTP in Tg-SOD mice. Tetanic stimulation was first applied to the perforant path, and the response was recorded with a control pipette. Thereafter, the pipette was replaced with one containing 1 mmbicuculline, which by itself did not change the response characteristics of the hippocampus, but it did permit the expression of subsequent LTP to the same perforant path stimulation.

Fig. 9.

Diazepam (DZ; intraperitoneal injection) blocks the ability of normal control mice to express LTP. Control and DZ-treated mice are shown, before and after tetanic stimulation applied to the perforant path. A,Paired-pulse depression is enhanced in control mice after injection of diazepam. B, Summary of the results with the tetanic stimulation in the two groups of control mice.