Maria H. Millan

Induction of audiogenic seizure susceptibility by focal infusion of excitant amino acid or bicuculline into the inferior colliculus of normal rats.

N-Methyl-D-aspartate (NMDA) (10 to 20 nmol) or bicuculline (15 to 50 pmol) in 0.5 microliter was infused bilaterally into the inferior colliculus or the deep layers of superior colliculus (DLSC) in normal rats, and the response to high intensity acoustic stimulation was examined. Thirty-five percent of rats receiving NMDA infusions and 42% of animals receiving bicuculline infusions into the inferior colliculus exhibited sound-induced seizures exclusively that were behaviorally similar to audiogenic seizures displayed by genetically epilepsy-prone rats. Rats receiving microinjections into the DLSC did not display sound-specific seizures. A combined pattern of spontaneous and sound-induced seizures was seen in some rats with both drugs and loci of microinjection. These data and previous studies support a role for increased excitant amino acid action and decreased efficacy of GABA in the inferior colliculus as important mechanisms involved in genetic susceptibility to audiogenic seizures.

Excitant amino acids and audiogenic seizures in the genetically epilepsy-prone rat. II. Efferent seizure propagating pathway ☆

The afferent pathway involved in initiation of audiogenic seizures in the genetically epilepsy-prone rat was investigated by bilateral microinfusion of the excitant amino acid antagonist 2-amino-7-phosphonoheptanoate into the major brain stem and subcortical nuclei of the auditory system. This antagonist has been shown to possess anticonvulsant properties in other seizure models, and an excitant amino acid has been implicated as a putative neurotransmitter in several of these nuclei. Seizure severity was significantly reduced following infusion of this agent into the cochlear nucleus, superior olivary complex, inferior colliculus, and medial geniculate body. Many of these animals exhibited a complete blockade of seizures. The smallest effective dose in the cochlear nucleus and the medial geniculate body was 5 nmol per side. The smallest effective dose in the olive was 1 nmol, and in the inferior colliculus 0.1 nmol per side was protective. The onset of anticonvulsant effectiveness was earliest in the inferior colliculus. These findings showed that the inferior colliculus was the most sensitive auditory center to the anticonvulsant action of 2-amino-7-phosphonoheptanoate and that imbalance between inhibitory and excitatory transmission within this brain structure may be crucial in the initiation of audiogenic seizures in the genetically epilepsy-prone rat.

Extracellular amino acid levels in hippocampus during pilocarpine-induced seizures.

Extracellular levels of aspartate, glutamate and glutamine were monitored by microdialysis in the dorsal hippocampus of freely moving rats following the administration of a convulsant dose of pilocarpine (400 mg/kg, i.p.). Rats were either pretreated with the glutamate uptake inhibitor, 1-trans-pyrrolidine-2,4-dicarboxylic acid (PDC, 1 mM in the perfusion medium, -25 min), or received pilocarpine directly. All rats injected with pilocarpine (with or without PDC pretreatment) developed limbic seizures (latency 15.4 +/- 2.4 min). Without PDC pretreatment there were no significant changes in extracellular levels of aspartate, glutamate and glutamine following pilocarpine administration until the onset of limbic seizures when glutamine levels fell by 35%. Following PDC pretreatment there were large and sustained increases in extracellular hippocampal aspartate (250%) and glutamate (55%) levels, but no significant change in the glutamine level. When pilocarpine was administered to this group of rats, there were further selective, significant, transient increases in the extracellular levels of aspartate (31%) and glutamate (18%) which preceded the onset of seizures. Aspartate and glutamate levels were not significantly increased (relative to PDC controls) during seizures. The conditions for pilocarpine-induced increases in aspartate and glutamate release were established in parallel groups of anaesthetised rats where pilocarpine was administered via a microdialysis probe in the dorsal hippocampus. Following the infusion of 10 mM pilocarpine there were large and rapid increases in the levels of aspartate (143%) and glutamate (179%), which were completely abolished by the absence of calcium in the perfusion medium, or by the presence of atropine (20 mM) or tetrodotoxin (1 microM).

Focal injection of 2-amino-7-phosphonoheptanoic acid into prepiriform cortex protects against pilocarpine-induced limbic seizures in rats

Intraperitoneal injection of pilocarpine (380 mg/kg) produces motor limbic seizures in rats. Focal injection into the prepiriform cortex (PC) of an N-methyl-D-aspartate receptor antagonist, 2-amino-7-phosphonoheptanoic acid (APH), 1-10 pmol, potently protects against these seizures and their pathological consequences. Sites similarly sensitive to the protective action of APH are found along a substantial part of the anterior-posterior extension of the piriform cortex. More caudal injection sites, located at the level of lateral septum are less sensitive. The anticonvulsant action of APH along the extent of the PC is localised in the vicinity of the injection site, as shown by autoradiography following focal injection of tritiated APH.

Decrease in excitatory transmission within the lateral habenula and the mediodorsal thalamus protects against limbic seizures in rats

We have used limbic convulsions induced by systemic pilocarpine in rats combined with focal intracerebral injections concurrently to study the initiation and spread of seizure activity. Protection against pilocarpine-seizure development by antagonism of excitatory or facilitation of inhibitory neurotransmission at focal sites establishes the anatomical circuits involved in the propagation of seizures. The excitatory amino acid antagonist 2-amino-7-phosphonoheptanoate (APH, selective for the NMDA preferring glutamate receptor subtype) is potently anticonvulsant after bilateral focal injections into the habenula or mediodorsal thalamus. The dose of APH required to give sustained protection against pilocarpine-induced convulsions is 10 pmol for lateral habenula and 50 pmol for mediodorsal thalamus. The GABA agonist muscimol produces a similar sustained protection following focal injections (100 pmol/side) into either the lateral habenula or the mediodorsal thalamus. An overall decrease in the efferent neurotransmission of these two brain regions results in a strong anticonvulsant effect indicating their importance in modulating limbic seizure activity.

Induction of audiogenic seizures in normal and genetically epilepsy-prone rats following focal microinjection of an excitant amino acid into reticular formation and auditory nuclei

An excitant amino acid (EAA), N-methyl-D-aspartate (NMDA), induces susceptibility to seizures when bilaterally microinjected into subcortical auditory nuclei of normal rats. Thirty-five percent of animals exhibit only audiogenic seizures (AGS) after infusions of NMDA into inferior colliculus (IC). Infusions into cochlear nucleus and medial geniculate body never produce susceptibility to AGS without non-audiogenic seizures (N-AGS). The overall seizure incidence (AGS and N-AGS) with IC infusions is 100%, but the incidence is less than 50% with infusions into cochlear nucleus or medial geniculate body. Although AGS susceptibility is induced by NMDA infusions in normal animals, the seizures are submaximal in severity and lack tonic components. Bilateral infusions of NMDA into IC or reticular formation of the substrain of genetically epilepsy-prone rats (GEPRs) that exhibits submaximal AGS (GEPR-3s) do not increase seizure severity. These data along with studies showing increased EAA levels and excitotoxic-like damage in the IC of the GEPR and blockade of AGS with an EAA receptor antagonist or synthesis inhibitor suggest that an EAA in the IC is involved in initiation of AGS in the GEPR. However, EAA action in the GEPR IC is not sufficient to induce the complete spectrum of seizure behaviors, and additional mechanisms may be required for induction of maximal severity audiogenic seizures.

2-Amino-7-phosphonoheptanoic acid (2-APH) infusion into entopeduncular nucleus protects against limbic seizures in rats

Motor limbic seizures occur following a systemic injection of pilocarpine (380 mg/kg) in rats. Focal injection of the selective N-methyl-D-aspartate receptor antagonist, (+/-)-2-amino-7-phosphonoheptanoic acid (2-APH, 5-20 pmol), bilaterally into the entopeduncular nucleus (EP) prior to pilocarine blocks these seizures. Muscimol (50 pmol), a potent gamma-aminobutyric acid receptor agonist, injected bilaterally into EP produces a similar protection against pilocarpine-induced seizures. Thus by blocking excitatory neurotransmission or facilitating inhibition within the EP, the severity of limbic seizures can be reduced.

Neurotransmission in the pedunculopontine nucleus and pilocarpine-induced motor limbic seizures in rats

Systemic injection of the cholinergic agonist, pilocarpine (380 mg/kg, i.p.) initiates a sequence of events leading to motor limbic seizure activity. Focal injection of the excitatory amino acid antagonist, 2-amino-7-phosphonoheptanoic acid (50 pmol-1 nmol) into the pedunculopontine nucleus (PPN), prior to pilocarpine injection, results in a powerful anticonvulsant action. The GABA agonist, muscimol (25-50 pmol) also afforded protection against pilocarpine-evoked convulsions when injected focally into the PPN. The results suggest that an overall inhibition of PPN output neurons is required for anticonvulsant action.

The high pressure neurological syndrome in genetically epilepsy prone rats : protective effect of 2-amino-7-phosphono heptanoate

Genetically epilepsy prone rats (GEPR) are hypersensitive to various epileptogenic treatments and undergo characteristic generalized seizures when exposed to potent acoustic stimulation. We have studied the sensitivity of GEPR to high atmospheric pressure. Threshold pressures for behavioral symptoms of the high pressure neurological syndrome (HPNS) were recorded in normal Sprague-Dawley (SD) and GEPR (which originate from the SD strain) of both sexes. The threshold pressure (TP) for tremor and for convulsion was significantly lower in GEPR than in SD rats. The protective action of the NMDA receptor antagonist D-2-amino-7-phosphono-heptanoate (D-APH) was tested on both strains of rats. D-APH, 90 mg/kg ip was more protective against tremor in SD than in GEPR. Female GEPR were not protected against tremor. Protection against clonic seizures was similar in both sexes of GEPR and female SD rats while SD males were not significantly protected. None of the animals treated with D-APH developed the tonic phase of seizures. Blockade of the NMDA receptor with D-APH brought the threshold for convulsions in GEPR to a similar pressure to that obtained in SD vehicle-injected controls. This findings suggests the involvement of the excitatory amino acid system in the hypersensitivity of GEPR to high atmospheric pressure.

Dual inhibitory action of enadoline (CI977) on release of amino acids in the rat hippocampus

The effect of the kappa-opioid receptor agonist enadoline (CI977, (5R)-(5 alpha,7 alpha,8 beta)-N-methyl-N-[7-(1-pyrrilidinyl)-1-oxaspiro [4,5]dec-8-yl-4-benzofuranacetamide monohydrochloride), on the release of amino acids was studied in the hippocampus of freely moving rats. K+, 100 mM, or veratrine, 100 microM, were applied for 10 min via the dialysis probe, either alone (control groups) or together with CI977 (after a 10 min pretreatment with CI977 in the perfusion medium). To test the specificity of the response to CI977, nor-binaltorphimine, a selective kappa-opioid receptor antagonist, was delivered together with CI977 in two groups of animals. To test the effect of systemic injection, CI977 was given subcutaneously 30 min prior to either stimulus. K(+)-induced release of glutamate and aspartate was significantly reduced by CI977, 2.5 mM; release of gamma-aminobutyric acid (GABA) was reduced by 250 microM CI977 in the probe. The effect of CI977 on release of glutamate and aspartate, but not of GABA, was reversed by nor-binaltorphimine (45 microM). Systemic treatment with CI977, 1 or 10 mg/kg, did not reduce K(+)-induced release of glutamate. Veratrine-induced release of aspartate and glutamate was significantly inhibited by 25 microM and release of GABA by 250 microM CI977 in the probe, and this effect was not modified by nor-binaltorphimine (58 microM). Systemic injection of CI977 1 mg/kg significantly reduced veratrine-induced release of glutamate. These results indicate that CI977 regulates release of amino acids by two independent mechanisms.(ABSTRACT TRUNCATED AT 250 WORDS)