Gabapentin inhibits presynaptic Ca2+ influx and synaptic transmission in rat hippocampus and neocortex

https://doi.org/10.1016/S0014-2999(02)02044-7Get rights and content

Abstract

Gabapentin is a widely used drug with anticonvulsant, antinociceptive and anxiolytic properties. Although it has been previously shown that Gabapentin binds with high affinity to the α2δ subunit of voltage-operated Ca2+ channels (VOCC), little is known about the functional consequences of this interaction. Here, we investigated the effect of Gabapentin on VOCCs and synaptic transmission in rat hippocampus and neocortex using whole-cell patch clamp and confocal imaging techniques. Gabapentin (100–300 μM) did not affect the peak amplitude or voltage-dependency of VOCC currents recorded from either dissociated or in situ neocortical and hippocampal pyramidal cells. In contrast, Gabapentin inhibited K+-evoked increases in [Ca2+] in a subset of synaptosomes isolated from rat hippocampus and neocortex in a dose-dependent manner, with an apparent half-maximal inhibitory effect at ∼100 nM. In hippocampal slices, Gabapentin (300 μM) inhibited the amplitude of evoked excitatory- and inhibitory postsynaptic currents recorded from CA1 pyramidal cells by 30–40%. Taken together, the results suggest that Gabapentin selectively inhibits Ca2+ influx by inhibiting VOCCs in a subset of excitatory and inhibitory presynaptic terminals, thereby attenuating synaptic transmission.

Introduction

Gabapentin (1-(aminomethyl)cyclohexane acetic acid, Neurontin®) is an anticonvulsant used as add-on therapy for the treatment of epileptic seizures. In addition, Gabapentin exhibits antinociceptive and anxiolytic effects Macdonald and Kelly, 1993, Morris, 1999. Despite the plethora of literature on the therapeutic efficacy of Gabapentin, there is no consensus on its molecular mechanism of action (Taylor et al., 1998). A single high affinity (KD∼35 nM) binding site for [3H]Gabapentin has been described in the outer layers of the cortex, and the dendritic regions of the hippocampal CA1 area Suman-Chauhan et al., 1993, Hill et al., 1993, and it has been suggested that this binding site corresponds to the auxiliary α2δ subunit of voltage-operated Ca2+ channels (VOCCs; Gee et al., 1996). However, there are conflicting reports on the functional effects of Gabapentin on VOCCs.

It has been reported that Gabapentin inhibits high voltage-activated Ca2+ channels in dissociated rat cortical pyramidal cells, medium spiny striatal neurons and large globus pallidus cells Stefani et al., 1998, Stefani et al., 2001 and in isolated dorsal root ganglion neurones Alden and Garcia, 2001, Sutton et al., 2002. In contrast, Rock et al. (1993) reported no modulation of VOCCs by Gabapentin (up to 1 mM) in rat nodose and dorsal root ganglion neurons. Moreover, Gabapentin does not inhibit VOCCs in cardiac myocytes (Alden and Garcia, 2001) and in hippocampal granule cells from patients with temporal lobe epilepsy (Schumacher et al., 1998).

In contrast to the unequivocal effects of Gabapentin on postsynaptic VOCCs, there is now substantial evidence that Gabapentin inhibits presynaptic VOCCs and attenuates neurotransmitter release from central nerve terminals Dooley et al., 2000, Fink et al., 2000, Meder and Dooley, 2000, Fink et al., 2002. In addition, it has been reported that Gabapentin inhibits the frequency, but not the amplitude, of miniature excitatory postsynaptic currents (EPSCs) in the superficial lamina of the spinal cord dorsal horn (Shimoyama et al., 2000). These results suggest a presynaptic site of action for Gabapentin.

In this study, we compared the effect of Gabapentin on pre- and postsynaptic Ca2+ influx in rat hippocampus and neocortex. In addition, we investigated the effect of Gabapentin on synaptic transmission in hippocampus. Parts of this study have appeared as abstract (van Hooft et al., 2000).

Section snippets

Dissociated cells and slices

Dissociated cells and slices were prepared as described before (Vreugdenhil and Wadman, 1992). Transverse or saggital hippocampal and neocortical slices (250 μm) from young adult male Wistar rats were cut on either a tissue chopper or a vibroslicer. For the preparation of dissociated cells, subslices of areas CA1, CA3 and neocortex were incubated (1 h for CA1 and CA3, 30 min for neocortex) in dissociation solution containing (in mM): NaCl (120), KCl (5), CaCl2 (1), MgCl2 (1), NaHCO3 (10),

Gabapentin does not inhibit postsynaptic Ca2+ currents

Dissociated pyramidal neurons from hippocampal CA1 and CA3 areas and neocortex were whole-cell voltage clamped, and VOCCs were evoked by depolarization. Activation- and inactivation curves were similar to those described previously (Fig. 1A; Vreugdenhil and Wadman, 1992), and were not affected in the presence of 100 μM Gabapentin (Table 1). Fig. 1C shows that the application of 100 μM Gabapentin did not affect the peak amplitude of the VOCC recorded from dissociated hippocampal CA1 pyramidal

Discussion

Our results demonstrate that Gabapentin inhibits presynaptic Ca2+ influx, and that Gabapentin inhibits excitatory as well as inhibitory synaptic transmission in hippocampus and neocortex. We did not detect an inhibitory effect of Gabapentin on postsynaptic VOCCs recorded in either dissociated cells or slices from hippocampus and neocortex. However, VOCCs were rapidly and completely blocked by Cd2+ in these preparations (Fig. 1C), indicating that a putative blocking effect of Gabapentin could

Acknowledgements

We thank Parke-Davis Pharmaceuticals/Pfizer (Hoofddorp, The Netherlands) for the generous gift of Gabapentin and Fernando Lopes da Silva for his comments on the manuscript. This study was financially supported by a Netherlands Organization of Scientific Research (NWO) grant 903-42-065 to JAvH.

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