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- Title
Loss of GABAergic Interneurons in Seizure-induced Epileptogenesis.
- Authors
Kobayashi, M.; Buckmaster, P.S.; Sayin, U.; Osting, S.; Hagen, J.; Rutecki, P.; Sutula, T.; Dinocourt, C.; Petanjek, Z.; Freund, T.F.; Ben-Ari, Y.; Esclapez, M.; Dudek, F. Edward
- Abstract
Reduced Inhibition of Dentate Granule Cells in a Model of Temporal Lobe Epilepsy Patients and models of temporal lobe epilepsy have fewer inhibitory interneurons in the dentate gyrus than do controls, but it is unclear whether granule cell inhibition is reduced. We report the loss of γ-aminobutyric acid (GABA)ergic inhibition of granule cells in the temporal dentate gyrus of pilocarpine-induced epileptic rats. In situ hybridization for GAD65 messenger RNA (mRNA) and immunocytochemistry for parvalbumin and somatostatin confirmed the loss of inhibitory interneurons. In epileptic rats, granule cells had prolonged excitatory postsynaptic potentials (EPSPs), and they discharged more action potentials than did controls. Although the conductances of evoked inhibitory postsynaptic potentials (IPSPs) recorded in normal artificial cerebrospinal fluid (aCSF) were not significantly reduced, and paired-pulse responses showed enhanced inhibition of granule cells from epileptic rats, more direct measures of granule cell inhibition revealed significant deficiencies. In granule cells from epileptic rats, evoked monosynaptic IPSP conductances were less than 40% those of controls, and the frequency of GABA[sub A] receptor–mediated spontaneous and miniature IPSCs (mIPSCs) was less than 50% that of controls. Within 3 to 7 days after pilocarpine-induced status epilepticus, miniature IPSC frequency had decreased, and it remained low, without functional evidence of compensatory synaptogenesis by GABAergic axons in chronically epileptic rats. Both parvalbumin- and somatostatin-immunoreactive interneuron numbers and the frequency of both fast- and slow-rising GABA[sub A] receptor–mediated mIPSCs were reduced, suggesting that loss of inhibitory synaptic input to granule cells occurred at both proximal/somatic and distal/dendritic sites. Reduced granule cell inhibition in the temporal dentate gyrus preceded the onset of spontaneous recurrent seizures by days to weeks, so it may contribute, but is insufficient, to cause epilepsy. Spontaneous Seizures and Loss of Axo-axonic and Axo-somatic Inhibition Induced by Repeated Brief Seizures in Kindled Rats Repeated brief seizures evoked by kindling progressively increase seizure susceptibility and eventually induce spontaneous seizures. Previous studies demonstrated that the initial seizures evoked by kindling increase paired-pulse inhibition at 15- to 25-millisecond interpulse intervals in the dentate gyrus and also induce apoptosis, progressive neuronal loss, mossy fiber sprouting, and neurogenesis, which could potentially alter the balance of excitation and/or inhibition and modify functional properties of hippocampal circuits. In these experiments, paired-pulse inhibition in the dentate gyrus was reduced or lost after approximately 90 to 100 evoked seizures in association with emergence of spontaneous seizures. Evoked inhibitory postsynaptic currents (IPSCs) examined by single-electrode voltage-clamp methods in granule cells from kindled rats experiencing spontaneous seizures demonstrated altered kinetics (reductions of ∼48% in 10% to 90% decay time, ∼40% in τ, and ∼65% in charge transfer) and confirmed that decreased inhibition contributed to the reduced paired-pulse inhibition. The loss of inhibition was accompanied by loss of subclasses of inhibitory interneurons labeled by cholecystokinin and the neuronal γ-aminobutyric acid (GABA) transporter GAT-1, which project axo-somatic and axo-axonic GABAergic inhibitory terminals to granule cells and axon initial segments. Seizure-induced loss of interneurons providing axo-somatic and axo-axonic inhibition may regulate spike output to pyramidal neurons in CA3 and could play an important role in generation of spontaneous seizures. The sequence of progressive cellular alterations induced by repeated seizures, particularly loss of GABAergic interneurons providing axo-somatic and axo-axonic inhibition, may be important in the development of intractable epilepsy. Loss of Interneurons Innervating Pyramidal Cell Dendrites and Axon Initial Segments in the CA1 Region of the Hippocampus Following Pilocarpine-induced Seizures In the pilocarpine model of chronic limbic seizures, vulnerability of γ-aminobutyric acid (GABA)ergic interneurons to excitotoxic damage has been reported in the hippocampal CA1 region. However, little is known about the specific types of interneurons that degenerate in this region. To characterize these interneurons, we performed quantitative analyses of the different populations of GABAergic neurons labeled for their peptide or calcium-binding protein content. Our data demonstrate that the decrease in the number of glutamic acid decarboxylase (GAD) messenger RNA (mRNA)-containing neurons in the stratum oriens of CA1 in pilocarpine-treated rats involved two subpopulations of GABAergic interneurons: interneurons labeled for somatostatin only (O-LM and bistratified cells) and interneurons labeled for parvalbumin only (basket and axo-axonic cells). Stratum oriens interneurons labeled for somatostatin/calbindin or somatostatin/parvalbumin were preserved. The decrease in number of somatostatin- and parvalbumin-containing neurons was observed as early as 72 hours after the sustained seizures induced by pilocarpine injection. Many degenerating cell bodies in the stratum oriens and degenerating axon terminals in the stratum lacunosum-moleculare were observed at 1 and 2 weeks after injection. In addition, the synaptic coverage of the axon initial segment of CA1 pyramidal cells was significantly decreased in pilocarpine-treated animals. These results indicate that the loss of somatostatin-containing neurons corresponds preferentially to the degeneration of interneurons with an axon projecting to stratum lacunosum-moleculare (O-LM cells) and suggest that the death of these neurons is mainly responsible for the deficit of dendritic inhibition reported in this region. We demonstrate that the loss of parvalbumin-containing neurons corresponds to the death of axo-axonic cells, suggesting that perisomatic inhibition and mechanisms controlling action-potential generation also are impaired in this model.
- Subjects
TEMPORAL lobe epilepsy; SPASMS; EPILEPSY; BRAIN diseases; INTERNEURONS
- Publication
Epilepsy Currents, 2003, Vol 3, Issue 5, p159
- ISSN
1535-7597
- Publication type
Article
- DOI
10.1046/j.1535-7597.2003.03503.x