An electrographic analysis of the synchronous discharge patterns of GEPR-9s generalized seizures.

Previous results from our Laboratory have shown a synchronous discharge pattern (less than 1 ms apart) in monopolar recordings from electrodes placed in the cortex, inferior colliculus, and medulla of seizing GEPR-9s. However, the wave morphology of the ictal EEG is quite different for electrodes placed in different anatomical structures. These results lead us to hypothesize that wave morphology was indicative of neural circuitry involved in the GEPR9 seizure and that volume conduction was accounting for synchronous epileptiform EEG pattern. We decided to approach the problem by using a set of two experiments. Experiment 1: Perform a complete precollicular transection in GEPR-9s before inducing seizure in order to observe changes in EEG morphology after forebrain circuitry removal. Experiment 2: A novel methodological approach using a three-dimensional bipolar array enabled the reconstruction of a vector indicative of to which direction is voltage increasing. Such time-varying vector is indicative of the source direction of the high-amplitude epileptiform EEG signal. By placing such an array of electrodes, used to record the 3 bipolar EEGs, in the forebrain, midbrain, and hindbrain, we were able to use a simple intersection method to infer source localization. Our results suggest that the slow wave component of the GEPR9 epileptiform ictal EEG pattern is associated with a midbrain-forebrain circuit while the spike component is associated with a midbrain-hindbrain substrate. These results are supported by experiment 1 in which only the spike component of EEG remained after the precollicular transection.

A comprehensive electrographic and behavioral analysis of generalized tonic-clonic seizures of GEPR-9s.

This study records noise-free intracerebral EEG of the genetically epilepsy prone rat (GEPR-9), along with behavioral correlates, during a seizure on unanesthetized freely behaving unrestrained animals. The GEPR-9 exhibits acoustically triggered generalized tonic-clonic seizures, and often times the EEG, recorded with conventional techniques, has resulted in data with imbedded movement artifact. For noise-free video-EEG recordings, we used a previously developed system that consists of a head connector with a FET preamplifier and battery, signal conditioning device (5000x gain, 1 Hz-100 Hz filters), A/D converter and video/PC-PC/video computer boards for recording image data. Each animal was implanted with three monopolar/referential electrodes chosen among the following areas: cortex, inferior colliculus, reticular formation and caudal medulla. The video-EEG data were quite similar for all recorded animals: (1) basal desynchronized EEG before sound stimulus; (2) increase in EEG frequency after stimulus and before seizure onset; (3) high-amplitude polyspikes during massive myoclonic thrusts with or without a very fast running episode; (4) an electrodecremental response during tonic extension; (5) wave and spike complex during forelimb and hindlimb tonic rigidity and posttonic clonus; (6) low-amplitude EEG during postictal depression. Time sequenced spectral analysis also highlights the epileptiform EEG pattern during seizure with high reproducibility between animals. While testing seizure naive GEPR-9s, there was a clear evolution from modest epileptiform EEG activity on the first acoustic stimulation to progressively higher amplitude, duration and frequency epileptiform EEG activity throughout seizure repetition.

Comparative fos immunoreactivity in the brain after forebrain, brainstem, or combined seizures induced by electroshock, pentylenetetrazol, focally induced and audiogenic seizures in rats.

To help discern sites of focal activation during seizures of different phenotype, the numbers of Fos immunoreactive (FI) neurons in specific brain regions were analyzed following "brainstem-evoked," "forebrain-evoked" and forebrain/brainstem combination seizures induced by a variety of methods. First, pentylenetetrazol (PTZ, 50 mg/kg) induced forebrain-type seizures in some rats, or forebrain seizures that progressed to tonic/clonic brainstem-type seizures in other rats. Second, minimal electroshock induced forebrain seizures whereas maximal electroshock (MES) induced tonic brainstem-type seizures in rats. Third, forebrain seizures were induced in genetically epilepsy-prone rats (GEPRs) by microinfusion of bicuculline into the area tempestas (AT), while brainstem seizures in GEPRs were induced by audiogenic stimulation. A final set was included in which AT bicuculline-induced forebrain seizures in GEPRs were transiently interrupted by audiogenic seizures (AGS) in the same animals. These animals exhibited a sequence combination of forebrain clonic seizure, brainstem tonic seizure and back to forebrain clonic seizures. Irrespective of the methods of induction, clonic forebrain- and tonic/clonic brainstem-type seizures were associated with considerable Fos immunoreactivity in several forebrain structures. Tonic/clonic brainstem seizures, irrespective of the methods of induction, were also associated with FI in consistent brainstem regions. Thus, based on Fos numerical densities (FND, numbers of Fos-stained profiles), forebrain structures appear to be highly activated during both forebrain and brainstem seizures; however, facial and forelimb clonus characteristic of forebrain seizures are not observable during a brainstem seizure. This observation suggests that forebrain-seizure behaviors may be behaviorally masked during the more severe tonic brainstem seizures induced either by MES, PTZ or AGS in GEPRs. This suggestion was corroborated using the sequential seizure paradigm. Similar to findings using MES and PTZ, forebrain regions activated by AT bicuculline were similar to those activated by AGS in the GEPR. However, in the combination seizure group, those areas that showed increased FND in the forebrain showed even greater FND in the combination trial. Likewise, those areas of the brainstem showing FI in the AGS model, showed an even greater effect in the combination paradigm. Finally, the medial amygdala, ventral hypothalamus and cortices of the inferior colliculi showed markedly increased FND that appeared dependent upon activation of both forebrain and brainstem seizure activity in the same animal. These findings suggest these latter areas may be transitional areas between forebrain and brainstem seizure interactions. Collectively, these data illustrate a generally consistent pattern of forebrain Fos staining associated with forebrain-type seizures and a consistent pattern of brainstem Fos staining associated with brainstem-type seizures. Additionally, these data are consistent with a notion that separate seizure circuitries in the forebrain and brainstem mutually interact to facilitate one another, possibly through involvement of specific "transition mediating" nuclei.

Penetrance and expressivity of genes involved in the development of epilepsy in the genetically epilepsy-prone rat (GEPR).

In order to understand the level of complexity of the epileptic phenotype in the two strains of Genetically Epilepsy-Prone Rats (GEPRs), we determined two important measures of genetic complexity, penetrance and expressivity. Penetrance is the percentage of animals of a specific genotype who express the phenotype associated with that underlying genotype. Expressivity refers to the degree that a particular genotype is expressed as a phenotype within an individual. Incomplete penetrance and variable expressivity are caused by genetic and environmental variation. In this paper we have studied the epileptic phenotype for 20,373 rats. Animals were tested on three occasions for audiogenic seizure and given an audiogenic response score (on a scale of 0-9, 0 being no seizure and 9 being the most severe). The GEPR-3 and GEPR-9 animals both show incomplete penetrance and variable expressivity of the underlying genetic predisposition. The GEPR-9 strain has more animals that have variable levels of seizure predisposition (as measured by a scoring system that denotes the severity of generalized tonic/clonic seizures) and a greater percentage of animals that exhibit no susceptibility to such seizures induced by sound. Both strains have a number of animals that are not susceptible to sound-induced GTCSs and that exhibit some variability in seizure severity. The GEPR-9 males show greater differences in expressivity and penetrance compared to GEPR-9 females. The GEPR-3 animals also show sex-associated variable penetrance and expressivity of the epileptic phenotype, although the differences are much smaller. These findings are the first step toward the mapping of the underlying quantitative trait loci (QTL) for seizure in these animals.

Fos expression and 2-deoxyglucose uptake following seizures in developing genetically epilepsy-prone rats.

Juvenile genetically epilepsy-prone rats (GEPR)-3s display one of three types of seizures in response to sound: a typical class 3 seizure consisting of an explosive running/bouncing episode followed by a clonic seizure (audiogenic response score, ARS-3); an ARS-3 seizure followed by a forebrain seizure that includes facial and forelimb (F&F) clonus with rearing (ARS-3f); or, a running/bouncing episode followed by a severe tonic seizure with complete hindlimb extension (ARS-9) not accompanied with subsequent F&F clonus. The adult seizure phenotype, manifest in all GEPR-3s by age 45 days of age, consists of an ARS-3 not followed by F&F clonus or tonic extension. The present studies sought to determine the neuronal networks activated during these various developmental convulsive patterns by examining anatomical patterns of [(14)C]2-deoxyglucose (2-DG) uptake or immediate-early-gene (Fos) expression subsequent to seizures. Many, but not all, brain areas of control rats showed age-related increases in Fos expression in response to the acoustic stimulation. An age effect was not observed in 2-DG uptake. In GEPRs, the profiles of Fos expression and 2-DG uptake following seizures were often parallel; however, there were notable exceptions. For example, increased 2-DG uptake in the cochlear nuclei, central region of the inferior colliculi, and the substantia nigra were not accompanied by increased Fos expression in these areas regardless of the seizure phenotypes. Reciprocally, other regions, particularly in the amygdala, ventromedial hypothalamus and parabrachial areas, displayed intense seizure related Fos labeling without detectable increases in 2-DG uptake. Fos and 2-DG uptake patterns in response to acoustic stimulation varied according to brain region, seizure phenotype and severity. In general, the degree of 2-DG uptake correlated with seizure severity. For example, the ARS-9 seizures, being the most intense, resulted in significant increases in 2-DG uptake in almost all brain regions examined. 2-DG uptake following the ARS-3f and ARS-3 seizures, although increased, did not reach statistical significance in most brain areas. In contrast to the 2-DG findings, a seizure-severity dependent effect was not seen with Fos. Rather, the induction of Fos associated with acoustic stimulation and seizure was more associated with age and seizure-phenotype. Thus, the developmental profiles of Fos expression and 2-DG uptake in response to seizures are distinctly different and concurrent examination of both markers is useful in the identification of brain circuitry involved in seizure development.

Effect of norepinephrine release on adrenoceptors in severe seizure genetically epilepsy-prone rats.

The genetically epilepsy-prone rat (GEPR) seizure model is characterized by extensive abnormalities in brain noradrenergic function. Earlier studies had suggested that GEPRs might not regulate adrenoceptors in a normal fashion. The purpose of the present study was to determine if GEPR-9s are capable of up and down regulation of alpha(1)- and beta-adrenoceptors in response to increments or decrements in extracellular norepinephrine. Seizure induction has been shown to increase extracellular norepinephrine. Chronic sound or electroshock-induced seizures caused down regulation of beta-adrenoceptors in frontal cortex and in hippocampus from GEPR-9s. Similarly, chronic daily treatment with the norepinephrine reuptake inhibitor desmethylimipramine produced down regulation of beta-adrenoceptors in frontal cortex and in hippocampus from GEPR-9s. As is the case in neurologically normal animals, chronic electroshock-induced seizure did not cause down regulation of beta-adrenoceptors in 6-hydroxydopamine pretreated GEPR-9s. Chronic electroshock treatment also caused up-regulation of alpha(1)-adrenoceptors in frontal cortex but not in hippocampus. In 6-hydroxydopamine pretreated GEPR-9s, chronic electroshock treatment caused a further up-regulation of alpha(1)-adrenoceptors in frontal cortex but not in hippocampus. Taken together, these results indicate that GEPR-9s are capable of up and down regulation of alpha(1)- and beta-adrenoceptors in a manner that is qualitatively similar to the regulation of these receptors in normal animals. Whether the regulation of brain adrenoceptors is quantitatively different in GEPRs from normal animals remains to be established.

Subcutaneous microdialysis in rats correlates with carbamazepine concentrations in plasma and brain.

Microdialysis simplifies the measurement of pharmacokinetics in pharmacodynamically relevant tissues. Microdialysis permits serial measurements of unbound drug concentration. The objective of the present work was to study rats to correlate plasma carbamazepine pharmacokinetics with subcutaneous and brain tissues. Microdialysis probes were inserted into the jugular vein, the brain, and subcutaneous tissue in Sprague-Dawley rats. After receiving single doses of carbamazepine, 12 mg/kg i.p., pharmacokinetic sampling occurred simultaneously from three microdialysis sites. Microdialysis sampled unbound carbamazepine and carbamazepine-10, 11-epoxide concentrations. Concentrations measured in brain, subcutaneous, and plasma correlated with each other. Except where differences were anticipated, pharmacokinetic parameters, including half-life and time to maximum concentration, were the same regardless of measurement site. The present study suggests microdialysis may allow pharmacokinetic measurements in peripheral physiological spaces that are surrogates for the pharmacologically relevant tissue.

Effect of repeated seizure experiences on tyrosine hydroxylase immunoreactivities in the brain of genetically epilepsy-prone rats.

The genetically epilepsy-prone rat (GEPR) is a model of generalized tonic/clonic epilepsy, and has functional noradrenergic deficiencies that act as partial determinants for the seizure predisposition and expression. The present study investigated the effect of repeated seizure experiences by acoustic stimulation (110 dB, 10 times) on the immunoreactivities of tyrosine hydroxylase (TH), a rate-determining enzyme in the synthesis of norepinephrine, in brain regions of GEPRs. TH immunoreactivity in locus coeruleus, the major noradrenergic nucleus in brain, was lower in GEPRs than control Sprague-Dawley rats. It was also decreased in several regions including inferior colliculus of GEPRs. Repeated experiences of audiogenic seizures further decreased TH immunoreactivities in locus coeruleus and inferior colliculus of GEPRs. The results from the present study suggest that the lower immunoreactivities of TH in locus coeruleus and inferior colliculus contribute, at least in part, to the noradrenergic deficits in GEPRs, and repeated seizure experiences further intensified these noradrenergic deficits, which may be related to the altered seizure expression by repetitive audiogenic seizure in GEPRs.

Carbamazepine pharmacokinetics-pharmacodynamics in genetically epilepsy-prone rats.

Carbamazepine produces dose-related anticonvulsant effects in epilepsy models including the genetically epilepsy-prone rat (GEPR) model and the rat maximal electroshock model. Dose-response relationships are quantitatively different among the models. Against electroshock seizures in Sprague-Dawley rats the ED50 dose is 7.5 mg/kg whereas the ED50 against audiogenic seizures in severe seizure GEPRs (GEPR-9s) is 3 mg/kg. In contrast, the ED50 in moderate seizure GEPRs (GEPR-3s) is 25 mg/kg. The present study was designed to ascribe dose-response differences among the three rat strains to pharmacokinetic or pharmacodynamic factors. After systemic carbamazepine, pharmacokinetic studies revealed differences in area under the concentration-vs.-time curve. In other experiments, carbamazepine-induced serotonin release from hippocampus was used as a pharmacodynamic marker. In a concentration-controlled design using intracerebral microdialysis, hippocampal carbamazepine infusions produced similar concentration-response relations for the three rat strains. These data support the hypothesis that dose-response differences among the three rat strains are primarily pharmacokinetic in nature.

Morphological deficits in noradrenergic neurons in GEPR-9s stem from abnormalities in both the locus coeruleus and its target tissues.

The epileptic condition of the genetically epilepsy-prone rat (GEPR) appears to be caused partially by deficiencies in the locus coeruleus (LC) innervation of the superior colliculus (SC). Previous studies provide quantitative documentation of noradrenergic morphological deficits in the moderately epileptic GEPR-3. The present findings extend these studies by applying cell culture methodology to assessments of the severely epileptic GEPR-9. Our data show that total neurite length, the number of neurite branch points per cell, the cross-sectional area of cell bodies, and the cell perimeter are deficient in noradrenergic neurons in LC + SC cocultures derived exclusively from GEPR-9s compared to analogous cocultures obtained solely from nonepileptic control rats. Partial restoration of LC neuron morphology toward normal occurs when the GEPR-9 SC component of the coculture is replaced with nonepileptic control SC. Finally, when the GEPR-9 SC is cocultured with the control LC, a partial morphological deficit occurs in the otherwise normal noradrenergic neurons. However, the magnitude of this deficit is less than that observed in noradrenergic neurons of the GEPR-9 LC cocultured with the control SC. These data support the hypothesis that the developmental deficiencies of noradrenergic neurons of the GEPR-9 are derived from two sources, the LC and its target tissue, in this case, the SC. Also, intrinsic abnormalities of the LC appear to make a more pronounced contribution to the noradrenergic deficits than do those which reside in the SC.

Effect of precollicular transection on audiogenic seizures in genetically epilepsy-prone rats.

Previous studies have demonstrated that generalized tonic-clonic seizures (GTCS) consisting of running/bouncing clonic and tonic extension can still be elicited in rats after brain transections which separate forebrain from brain stem, showing that forebrain circuitry is not required for GTCS. Inasmuch as sound-induced generalized tonic-clonic seizures in rodents are characterized by running-bouncing clonic and tonic convulsions, we have hypothesized that these are brain stem seizures that can occur independently of the forebrain. To test this hypothesis, we examined the response of two strains of genetically epilepsy-prone rats (GEPR-3s and GEPR-9s) to seizure-evoking auditory stimuli 3 h after a precollicular transection or sham surgery performed under ether anesthesia. In addition, the effect of a precollicular transection on audiogenic seizures was evaluated in normal rats made susceptible to such seizures by infusing NMDA into the inferior colliculus. Following the transection 58% of GEPR-9s displayed a sound-induced tonic-clonic convulsion and the remaining 42% exhibited a sound-induced seizure when subjected to stimulation 5 min after a subconvulsant dose of pentylenetetrazol (PTZ). While sham surgery and the precollicular transection both reduced sound-induced seizure severity in GEPR-3s, the full seizure response could be elicited by sound stimulation following a subconvulsant dose of PTZ. Moreover, the audiogenic seizures in normal rats rendered susceptible by NMDA were unaltered by the precollicular transection. These findings show that the anatomical circuitry required for generalized tonic-clonic seizures evoked by sound stimulation in rodents resides within the brain stem.

Effect of alterations in extracellular norepinephrine on adrenoceptors: a microdialysis study in freely moving rats.

Chronic electroshock treatment (once daily for 12 days) increases extracellular norepinephrine in the frontal cortex and hippocampus as measured by microdialysis. This chronic treatment produced an elevation of basal norepinephrine overflow into extracellular space while both the first and the twelfth treatments produced a transient increase in norepinephrine overflow of about 40 min. Acutely, desmethylimipramine (10 mg/kg) treatment significantly increased extracellular norepinephrine. While chronic desmethylimipramine (once daily for 10 days) increased basal overflow of norepinephrine in the frontal cortex and hippocampus, the tenth daily administration of desmethylimipramine did not produce a statistically significant increase in extracellular norepinephrine. Both daily electroshock and daily desmethylimipramine produced down regulation of beta-adrenoceptors in the hippocampus and the frontal cortex. Chronic electroshock caused up regulation of alpha-adrenoceptors in the frontal cortex but not in the hippocampus while chronic desmethylimipramine administration did not alter alpha-adrenoceptors in either structure. Depletion of norepinephrine with reserpine or with 6-hydroxydopamine prevented the down regulation of beta-adrenoceptors while depletion of this neurotransmitter did not prevent the electroshock-induced up regulation of alpha-adrenoceptors in the frontal cortex. These data suggest that down regulation of beta-adrenoceptors is mediated through increases in extracellular norepinephrine. In contrast, up regulation of alpha-adrenoceptors appears to be independent of norepinephrine release and does not require the presence of noradrenergic neurons in order to be induced by electroshock.

A noradrenergic and serotonergic hypothesis of the linkage between epilepsy and affective disorders.

Noradrenergic and/or serotonergic deficits, as well as other abnormalities, may contribute to predisposition to some epilepsies and depressions. Evidence for this hypothesis stems from several sources. Epidemiological investigations are intriguing but incomplete. Pharmacological studies show that noradrenergic and/or serotonergic transmission are both anticonvulsant and antidepressant. Therapeutically pertinent investigations show that antidepressant drugs have anticonvulsant properties, whereas antiepileptic drugs are effective in the management of affective disorders. Additional investigations demonstrate that seizures, whether spontaneously occurring or therapeutically induced, protect against depression. Through studies of innate pathophysiology, noradrenergic and serotonergic deficits have been identified in individuals with depression and in animal models of epilepsy, as well as in some humans with epilepsy. Vagal nerve stimulation, a treatment already known to be effective in the epilepsies, is presently under investigation for effectiveness in affective disorder. New evidence suggests that vagal nerve stimulation exerts at least some of its therapeutic effects through its capacity to increase noradrenergic and serotonergic transmission. Finally, emerging evidence supports the concept that some genetic mammalian models of the human epilepsies exhibit analogous manifestations of depression.

Carbamazepine-induced release of serotonin from rat hippocampus in vitro.

PURPOSE: Carbamazepine is one of several antiepileptic drugs (AEDs) that release the inhibitory neurotransmitter serotonin as part of their pharmacodynamic action on brain neurons. We undertook this study to investigate the cellular processes by which carbamazepine (CBZ) releases serotonin from brain tissue. METHODS: Tissue slices were prepared from hippocampi of Sprague-Dawley rats. These hippocampal slices were preincubated in vitro in a buffer so that neurons within the slice would take up tritium-labeled serotonin. Subsequently the slices were superfused with buffer containing CBZ or other chemicals (or both) that increase the overflow of serotonin radioactivity. RESULTS: Carbamazepine produced a concentration-dependent (50, 125, 250, or 500 microM) increase in basal overflow of serotonin radioactivity from superfused rat hippocampal slices in vitro. In contrast, these concentrations did not alter potassium-stimulated release, suggesting that the CBZ-induced release does not depend on depolarization or exocytosis. Blockade of the neuronal membrane serotonin transporter by fluoxetine (1 microM) or citalopram (2 microM) did not alter overflow of serotonin radioactivity produced by 250 microM CBZ. p-chloramphetamine (10 microM) produced a substantial increase in overflow of serotonin radioactivity, and this effect appears to be antagonized by 250 microM CBZ. Uptake of [3H]-labeled serotonin into hippocampal synaptosomes was inhibited by CBZ with a median inhibitory concentration (IC50) of 511+/-33 microM and a Hill coefficient of 0.87+/-0.11, suggesting competitive inhibition of uptake by CBZ. CONCLUSIONS: We conclude that CBZ (a) releases serotonin from hippocampal slices independent of exocytosis and by a mechanism not involving the neuronal membrane serotonin transporter, and (b) at high enough concentration, blocks the neuronal serotonin transporter.

Anticonvulsant effect of enhancement of noradrenergic transmission in the superior colliculus in genetically epilepsy-prone rats (GEPRs): a microinjection study.

An expanding body of data has indicated that the seizure prone state in genetically epilepsy-prone rats (GEPRs) is partially caused by deficits in central nervous system noradrenergic transmission. Several lines of evidence suggest that the noradrenergic terminals in the superior colliculus (SC) may act as determinants of seizure predisposition in the GEPR. In order to assess the role of the noradrenergic transmission in the SC in the regulation of seizure severity, several drugs with different mechanisms of enhancing noradrenergic transmission were bilaterally microinfused into the SC of GEPR-9s (severe seizure GEPRs). The rats were tested for audiogenic seizure intensity at 0.25, 1, 2, 3, and 4 h after treatments. Bilateral infusion of vehicle produced no reduction in the severity of the audiogenic seizure. Desipramine (2, 4, 8 micrograms/side), nisoxetine (2, 4, 8 micrograms/side), and idazoxan (0.25, 1, 4 micrograms/side) all decreased the seizure severity in a dose-dependent fashion. Significant decreases in the seizure severity were also observed after administration of methoxamine (0.15 microgram/side) or phenylephrine (0.15 microgram/side). Pretreatment with prazosin (1 microgram/side) significantly diminished the anticonvulsant effectiveness of methoxamine and nisoxetine while prazosin, by itself, had no effects on the seizure intensity. These results suggest that noradrenergic transmission in the SC may be involved in the seizure regulation in GEPR-9s, and that this regulation may be mediated, at least in part, through alpha 1 receptors.

Neurite extension of developing noradrenergic neurons is impaired in genetically epilepsy-prone rats (GEPR-3s): an in vitro study on the locus coeruleus.

A primary determinant of seizure susceptibility and severity in genetically epilepsy-prone rats (GEPRs), is a generalized deficiency in the central noradrenergic system of these animals. In particular, this deficiency includes reduced numbers of norepinephrine (NE) synaptic terminals in several brain areas and distinctly fewer NE axons within the auditory tectum. Two strains of GEPRs have been developed: GEPR-3s that have moderately severe clonic seizures and GEPR-9s that have severe tonic seizures culminating in complete hindlimb extension. Seizures in animals of each substrain are preceded by a brief episode of wild running. The developmental profile of NE axonal growth in GEPRs compared to control rats is not known, but may be causally related to NE deficiencies in this seizure model. The present study compared developmental neurite extension of fetal NE neurons in vitro between GEPR-3s and Sprague-Dawley control rats, the strain from which GEPR-3s were originally derived. Neurite arborization of individual NE neurons was assessed by quantitative morphometry following immunocytochemical identification of tyrosine hydroxylase (TH). Preliminary studies using explant and dispersed-cell cultures of control-rat tissues showed that optimal culture parameters to support neuritogenesis of LC neurons included the use of dispersed-cell cultures, Pronectin-F substrate, day-14 gestation donor-tissue, no use of cytosine-arabinofuranoside (ARA-c, a glial mitotic inhibitor) and the presence of co-cultured tectal tissue. Compared to fetal control-rat NE neurons co-cultured with fetal control-rat tectum, NE neurons derived from fetal GEPR-3 LC in co-culture with GEPR-3 tectum exhibited only 30% of the neurite extension of control-rat LC neurons and GEPR-3 LC neurons had a similarly deficient amount of branching. This study suggests, but does not prove, that deficiency in tectal NE in GEPR-3s involves a developmental deficiency in neurite extension from GEPR-3 LC neurons. Hypothetically, this deficiency may also contribute to the well described NE deficiency in other regions of the adult GEPR brain.

Effect of systemic ethanol on basal and stimulated glutamate releases in the nucleus accumbens of freely moving Sprague-Dawley rats: a microdialysis study.

This study was conducted to assess the impact of systemic ethanol (EOH) on the glutamatergic transmission in the nucleus accumbens (NACC). Extracellular concentrations of glutamate (GLU) in the NACC of freely moving Sprague-Dawley rats were monitored by intracerebral microdialysis. Intraperitoneal injection of EOH at a dose of 2 g/kg significantly decreased basal extracellular GLU by 21%. In addition, administration of the same dose of EOH significantly attenuated 150 mM K+-stimulated GLU release from the NACC by more than 50%. Since K+-stimulated GLU release has been demonstrated to derive largely from nerve terminal depolarization, reductions of K+-evoked GLU release may reflect in part the effect of EOH on the neurotransmitter pool. The present results suggest that EOH may suppress glutamatergic transmission in the NACC by lowering presynaptic GLU release.

Fos in locus coeruleus neurons following audiogenic seizure in the genetically epilepsy-prone rat: comparison to electroshock and pentylenetetrazol seizure models.

Seizures in genetically epilepsy-prone rats (GEPRs) may result from hypoactivity of locus coeruleus (LC) neurons during seizures. This study examined Fos-like-immunoreactivity (FLI) in the LC following audiogenic seizures in two strains of GEPRs (GEPR-9s and -3s), and following pentylenetetrazol (PTZ) or maximal electroshock seizures (MES) in normal rats. After tonic seizure, GEPR-9s showed an identical LC-FLI response to that of normal rats following tonic seizures induced by either PTZ or MES. GEPR-3s, having clonic seizures, had less FLI in the LC. Therefore, stimulus-transcription coupling in the GEPR LC is apparently normo-typic in its FLI response to seizure and thus is not likely the root cause of NE abnormalities in this seizure model.

Seizures and proto-oncogene expression of fos in the brain of adult genetically epilepsy-prone rats.

The mechanisms and brain circuitry that render genetically epilepsy-prone rats (GEPRs) susceptible to acoustically induced seizures are not completely known. The present study explores the neuroanatomy of acoustically induced seizures by immunohistochemical analysis of the proto-oncoprotein fos after intense acoustic stimulation (AS) with and without seizures. Acoustic stimulation induced tonic convulsions in GEPR-9s, but not in control rats. Locations of brain nuclei showing fos-like immunoreactive (FLI) neurons following AS with and without seizures were mapped. Semiquantitative methods were used to compare FLI neuron numerical densities in AS control rats and GEPRs. Many brain areas exhibited profound FLI in AS control rats and GEPRs. Unexpectedly, the cochlear nuclei and the central nucleus of the inferior colliculi (ICc), both of which are requisite for AGS initiation, exhibited a diminished fos expression in animals having seizures compared to AS controls. In contrast, GEPRs displayed a significant increase in FLI neurons within the dorsal cortex of the IC (ICd) compared to AS controls. This finding may suggest a seizure-related amplification of the auditory signal between the ICc and the ICd. Other nuclei, known to be involved in auditory transmission (i.e., superior olivary complex; trapezoid nucleus; dorsal nucleus of the lateral lemniscus, DNLL), did not show differential FLI densities between seizure and AS control animals. In contrast, seizure-induced FLI was observed in many nonauditory brain nuclei. Of particular interest was the identification of an intensely labeled nucleus in the GEPR. This nucleus resides in the most posterior and dorsal-lateral part of the pedunculopontine tegmental nucleus-pars compacta (PPTn-pc) immediately adjacent to the DNLL and extends posteriorly into the superior lateral subnucleus of the lateral parabrachial area (SLPBn). Therefore, we have tentatively termed this nucleus the PPSLPBn. The PPSLPBn lies in a region previously described as a mesencephalic locomotor region and a suspected functional involvement of this nucleus in display of seizure activity is under investigation. Other brain stem nuclei showing differential fos expression between GEPRs and AS control rats are also described.

Dizocilpine (MK-801) increases not only dopamine but also serotonin and norepinephrine transmissions in the nucleus accumbens as measured by microdialysis in freely moving rats.

The extracellular concentrations of dopamine (DA), norepinephrine (NE), and serotonin (5-HT) in the nucleus accumbens (NACC) of freely moving rats were monitored simultaneously via intracerebral microdialysis. Local infusion of the non-competitive N-methyl-D-aspartate (NMDA) receptor antagonist MK-801 (dizocilpine) (5-250 microM) produced significant increases in extracellular levels of DA, NE and 5-HT in a concentration-dependent fashion. Perfusion with tetrodotoxin (TTX, 1 microM) blocked the ability of focal MK-801 (50 microM) to increase DA, NE and 5-HT in the dialysate. Systemic administration of MK-801 (0.3 mg/kg, i.p.) also produced small, but statistically significant, increases in extracellular concentrations of DA, NE and 5-HT in the NACC. Our microdialysis results are consistent with the hypothesis that, in addition to dopaminergic, serotonergic and noradrenergic neurotransmissions in the NACC are involved in the mechanism by which MK-801 alters behavior in rats. Also, the present study gives further support to the concept that NMDA receptors within the NACC do not regulate DA release through direct excitatory control.