THE EFFECTS OF ENDOCRINE DISRUPTORS ON FEMALE GONADAL AXIS: AN UPDATE.

Endocrine disruptors (EDs) are considered to have an impact on the function of reproductive axis at different levels as well on reproductive organs in both sexes. Complexity of female reproductive system influenced with various stressors including EDs lead to morphological and functional alterations. This is resulting in modulation of neuroendocrine regulation with consequent developmental irregularities and derangements, causative infertility, endometriosis as well as premature ovarian insufficiency or polycystic ovary syndrome. A number of experimental clues was obtained on female animal models using various EDs such as synthetic estrogens and phytoestrogens, neurotransmitters, pesticides or various chemicals. These substances lead towards consequent derangement of the neuroendocrine control of reproduction from early phases of reproductive development towards different phases of adult reproductive period. This text will address some novel insights into the effects of EDs on neuroendocrine regulation of gonadal axis, effects on ovaries as well on endometrium during implantation period.

The effect of subchronic supplementation with folic acid and l-arginine on homocysteine-induced seizures.

The aim of the present study was to examine the effect of subchronic co-administration of folic acid (F) and l-arginine (A) on behavioural and electroencephalographic (EEG) characteristics of dl homocysteine thiolactone (H) induced seizures in adult rats. The activity of membrane ATPases in different brain regions were also investigated. Rats were treated with F, A, or vehicle for 15 days (regimen: F 5 mg/kg + A 500 mg/kg (F5A500); F 10 mg/kg + A 300 mg/kg (F10A300)). Seizures were elicited by convulsive dose of H (H, F5A500H, F10A300H) Subchronic supplementation with F and A did not affect seizure incidence, number of seizure episodes, and severity in F5A500H and F10A300H groups vs. H group. However, a tendency to increase latency and decrease the number of seizure episodes was noticed in the F10A300H group. EEG mean spectral power densities during ictal periods were significantly lower in F10A300H vs. H group. The activity of Na+/K+-ATPase and Mg2+-ATPase was significantly increased in almost all examined structures in rats treated with F and A. We can conclude that subchronic supplementation with folic acid and l-arginine has an antiepileptic effect in dl homocysteine thiolactone induced epilepsy.

The Influence of Finasteride on Mean and Relative Spectral Density of EEG Bands in Rat Model of Thioacetamide-Induced Hepatic Encephalopathy.

Liver failure is associated with a neuropsychiatric syndrome, known as hepatic encephalopathy (HE). Finasteride, inhibitor of neurosteroid synthesis, may improve the course of HE. The aim of our study was to investigate the influence of finasteride on mean and relative power density of EEG bands, determined by spectral analysis, in rat model of thioacetamide-induced HE. Male Wistar rats were divided into groups: (1) control; (2) thioacetamide-treated group, TAA (900 mg/kg); (3) finasteride-treated group, FIN (150 mg/kg); and (4) group treated with finasteride (150 mg/kg) and thioacetamide (900 mg/kg), FIN + TAA. Daily doses of FIN (50 mg/kg) and TAA (300 mg/kg) were administered during 3 subsequent days, and in FIN + TAA group FIN was administered 2 h before every dose of TAA. EEG was recorded 22-24 h after treatment and analyzed by fast Fourier transformation. While TAA did not induce significant changes in the beta band, mean and relative power in this band were significantly higher in FIN + TAA versus control group (p < 0.01). TAA caused a significant decline in mean power in alpha, theta, and delta band, and in FIN + TAA group the mean power in these bands was significantly higher compared with control. While in TAA group relative power was significantly decreased in theta (p < 0.01) and increased in delta band (p < 0.01) versus control, the opposite changes were found in FIN + TAA group: an increase in theta (p < 0.01) and a decrease in delta relative power (p < 0.01). In this study, finasteride pretreatment caused EEG changes that correspond to mild TAA-induced HE.

Homocysteine thiolactone-induced seizures in adult rats are aggravated by inhibition of inducible nitric oxide synthase.

Homocysteine and its metabolites (homocysteine thiolactone (HT)) induce seizures via different but still not well-known mechanisms. The role of nitric oxide (NO) in epileptogenesis is highly contradictory and depends on, among other factors, the source of NO production. The aim of the present study was to examine the effects of aminoguanidine, selective inhibitor of inducible NO synthase (iNOS), on HT-induced seizures. Aminoguanidine (50, 75, and 100 mg/kg, intraperitoneally (i.p.)) was injected to rats 30 min prior to inducing HT (5.5 mmol/kg, i.p.). Seizure behavior was assessed by seizure incidence, latency time to first seizure onset, number of seizure episodes, and their severity during observational period of 90 min. Number and duration of spike and wave discharges (SWDs) were determined in electroencephalogram (EEG). Seizure latency time was significantly shortened, while seizure incidence, number, and duration of HT-induced SWD in EEG significantly increased in rats receiving aminoguanidine 100 mg/kg before subconvulsive dose of HT. Aminoguanidine in a dose-dependent manner also significantly increased the number of seizure episodes induced by HT and their severity. It could be concluded that iNOS inhibitor (aminoguanidine) markedly aggravates behavioral and EEG manifestations of HT-induced seizures in rats, showing functional involvement of iNOS in homocysteine convulsive mechanisms.

Exercise decreases susceptibility to homocysteine seizures: the role of oxidative stress.

The aim of the study was to examine the effects of chronic exercise training on seizures induced by homocysteine thiolactone (HCT) in adult rats. Rats were assigned to: sedentary control; exercise control; sedentary+HCT; exercise+HCT group. Animals in the exercise groups ran 30 min daily on a treadmill for 30 consecutive days (belt speed 20 m/min), while sedentary rats spent the same time on the treadmill (speed 0 m/min). On the 31st day, the HCT groups received HCT (8.0 mmol/kg), while the control groups received vehicle. Afterwards, convulsive behavior and EEG activity were registered. Lipid peroxidation, superoxide dismutase (SOD) and catalase (CAT) activity were ascertained in the rat hippocampus. No signs of seizures were registered in sedentary and exercise control rats. Seizure latency was increased, while number of seizure episodes and spike-and-wave discharges (SWD) in EEG were decreased in the exercise+HCT compared to the sedentary+HCT group. Seizure incidence, the severity thereof and duration of SWDs were not significantly different between these groups. Exercise partly prevented increase of lipid peroxidation and decrease of the SOD and CAT activity after HCT administration. These results indicate beneficial effects of exercise in model of HCT-induced seizures in rats, what could be, at least in part, a consequence of improved antioxidant enzymes activity.

Spectral analysis of thioacetamide-induced electroencephalographic changes in rats.

Thioacetamide (TAA) is widely used as a model of hepatic encephalopathy (HE). The aim of our study was to investigate the effects of TAA on electroencephalographic (EEG) changes in rats and to compare them with human HE. Male Wistar rats were divided into groups: (1) saline-treated group and (2) TAA-treated groups: TAA(300) (300 mg/kg), TAA(600) (600 mg/kg), and TAA(900) (900 mg/kg). Daily dose of TAA (300 mg/kg) was administered intraperitoneally once (TAA(300)), twice (TAA(600)), or thrice (TAA(900)) in subsequent days. EEG changes were recorded about 24 h after the last dose of TAA. Absolute and relative power density in alpha bands were significantly higher in TAA(300) versus control group. In TAA(300), absolute beta power density was higher and relative beta power density was lower versus control group. Absolute alpha, theta, delta, and relative theta power were significantly lower, while relative power in delta band was significantly higher in TAA(900) versus control group (p < 0.01). In conclusion, decrease in EEG voltage with an increase in delta relative power, which correspond to the EEG manifestations of severe HE in humans, was observed in TAA(900) group. Electrical activity in TAA(300) group correlates with mild HE in humans.

The correlation between lipid peroxidation in different brain regions and the severity of lindane-induced seizures in rats.

The aim of this study was to investigate the dynamics of lipid peroxidation and the possible correlation between lipid peroxidation in different brain regions and behavioral manifestations in lindane-induced seizures in rats. Male Wistar rats were divided into the following groups: 1. control, saline-treated group; 2. dimethylsulfoxide (DMSO)-treated group; 3. lindane-treated group (8 mg/kg), intraperitoneally. Animals were sacrificed 0.5 or 4 h after treatment and the malondialdehyde level and superoxide dismutase (SOD) activity were determined in various brain regions spectrophotometrically. Behavioral changes were classified according to the descriptive scale (0--no response, 1--head nodding, lower jaw twitching; 2--myoclonic body jerks, bilateral forelimb clonus with full rearing; 3--progression to generalized clonic convulsions followed by tonic extension of fore- and hind limbs and tail; 4--status epilepticus). A significant rise in the malondialdehyde level was detected in the cerebral cortex, hippocampus, and thalamus of lindane-treated animals 0.5 and 4 h after administration (P < 0.05). SOD activity (total and mitochondrial) was significantly decreased in the hippocampus and the cortex of lindane-treated animals at both time points (P < 0.05). An initial fall in SOD activity was detected in the thalamus 4 h after lindane administration (P < 0.05). A positive correlation between seizure severity and the malondialdehyde level was found in the hippocampus at both time points (P < 0.01). These results suggest that lipid peroxidation may contribute to the neurotoxic effects of lindane in early acute lindane intoxication and that behavioral manifestations correlate with lipid peroxidation in the hippocampus of lindane-treated rats.

Oxidative stress in liver and red blood cells in acute lindane toxicity in rats.

The aim of our study was to determine the role and dynamics of oxidative and nitrosative stress, as well as superoxide dismutase (SOD) and catalase activity in the hepatocytes and erythrocytes in early phase of acute lindane intoxication. Male Wistar rats were divided into groups: control, dimethylsulfoxide and lindane-treated groups (L, 8 mg/kg, intraperitoneally). Animals were sacrificed 0.5 and 4 hours after treatment (L(0.5) and L(4) groups, respectively). Oxidative and nitrosative stress parameters and antioxidant enzymes were determined spectrophotometrically. Liver and plasma thiobarbituric acid reactive substances (TBARS) concentration were significantly increased 0.5 after lindane administration (p < .01), with subsequent additional rise within 4 hours (p < .01), while plasma nitrite + nitrate level was significantly higher only 4 hours after lindane treatment. Total liver SOD activity was significantly increased in L(4) group in comparison with control group (p < .01). In conclusion, oxidative and nitrosative stress play an important role in early phase of acute lindane hepatotoxicity. Antioxidant capacity of hepatocytes is partly increased, due to an adaptive increase in SOD activity.

Homocysteine, folic acid and coronary artery disease: possible impact on prognosis and therapy.

Within the past four decades, the efforts of investigators worldwide have established the amino acid homocysteine (Hcy) as an important factor in arteriosclerosis and ageing. The amino acid homocysteine is a unique candidate for the study of different age-related pathological conditions, namely vascular diseases, dementia disorders and late-life depression, due to its multiple roles in different pathways leading to atherosclerosis and neurotoxicity. Especially, the role of homocysteine in predicting risk for atherothrombotic vascular disease has been evaluated in several observational studies in a large number of patients. These studies show that the overall risk for vascular disease is small, with prospective, longitudinal studies reporting a weaker association between homocysteine and atherothrombotic vascular disease compared to retrospective case-control and cross-sectional studies. Furthermore, randomised controlled trials of homocysteine-lowering therapy have failed to prove a causal relationship. On the basis of these results, there is currently insufficient evidence to recommend routine screening and treatment of elevated homocysteine concentrations with folic acid and other vitamins to prevent atherothrombotic vascular disease.

Effect of acute lindane and alcohol intoxication on serum concentration of enzymes and fatty acids in rats.

This study examines possible synergistic effects of lindane and ethanol on inducing liver injury and serum fatty acid derangement in adult male Wistar rats. When administered together, ethanol and lindane-induced even more pronounced increase of alanine aminotransferase (165 +/- 10 U/L) and gamma-glutamyltranspeptidase activity (10.3 +/- 0.6 U/L) than after isolated administration of either substance. In addition, separate administration of lindane and ethanol was followed by a significant decrease of linoleic acid level in the serum (301 +/- 38 mg/L, 276 +/- 35 mg/L vs. 416 +/- 48 mg/L). However, when ethanol administration was followed by lindane injection, serum linoleic acid was at the similar level found in the control group (516 +/- 62 mg/L). Ethanol-treated rats that received lindane 30 min after ethanol administration have shown a marked increase of palmitic (421 +/- 27 mg/L) and linolic acid level (43 +/- 5 mg/L) in comparison with rats that have been treated only with ethanol (316+/-26 mg/L for palmitic and 32 +/- 2 mg/L for linolic acid) or lindane (295 +/- 26 mg/L for palmitic and 301 +/- 38 mg/L for linolic acid). Linolic acid level was significantly greater in comparison with control group (29 +/- 1 mg/L). In conclusion, this study found enough evidence to support the hypothesis that acute ethanol intoxication potentiates lindane-induced liver injury and enhances lipid derangement.

Ethanol suppressed seizures in lindane-treated rats. Electroencephalographic and behavioral studies.

This study examines the effects of ethanol on lindane-induced seizures in rats. The animals were divided into following groups: 1. saline, 2. DMSO (dimethylsulfoxide), 3. lindane dissolved in DMSO in the dose of 4, 6 or 8 mg/kg (L(4), L(6) and L(8) groups, respectively), 4. ethanol 2 g/kg administered 30 min prior to lindane (protected groups AL(4), AL(6) and AL(8)) and 5. ethanol alone (2 g/kg). In order to determine ethanol concentration in plasma, blood samples were collected by cardiac puncture 30 and 60 min after ethanol injection. For EEG and power spectra recordings, electrodes were implanted into the skull. The lindane treatment resulted in a dose-dependent increase of seizure incidence and severity. The rats displayed severe seizure patterns characterized by high voltage spike-wave complexes, poly-spikes and sleep-like patterns in EEG, while the power spectra were intensively elevated in comparison to the corresponding controls. Ethanol alone led to increased EEG power spectra, which became dominant in the range of 0-4 Hz. For evaluation of anticonvulsant ethanol action we compared latency to seizure, incidence and seizure severity (scale from 0 to 4) in the examined groups. Ethanol diminished seizure incidence in AL(4) and AL(6) groups, decreased intensity of convulsions, and prolonged duration of latency period in AL(8) group. We observed suppression of the EEG signs of lindane-provoked epileptiform activity in AL(4) and AL(6), but not in AL(8) group. These results suggest that ethanol acted protectively on lindane-induced seizures and suppressed behavioral and epileptic EEG spiking activity.

Valproate and delta-sleep peptide display high efficacy against metaphit-induced audiogenic seizure in rats.

The effects of valproate (VPA) and delta sleep-inducing peptide (DSIP) on metaphit-induced generalized, audiogenic seizure in adult rat males were compared. The animals were i.p. injected with: (1) Saline; (2) metaphit (mp, 10 mg kg(-1)); 3. metaphit (10 mg kg(-1)) and 8 h later with DSIP (0.1, 0.2, 0.4 or 1.0 mg kg(-1)), 4. metaphit (10 mg kg(-1)) and 8 h later with VPA (50, 75 or 100 mg kg(-1)); 5. DSIP alone (1.0 mg kg(-1)) and 6. VPA, alone (100 mg kg(-1)). The rats were exposed to sound stimulation at hourly intervals and the behavior and EEG were analyzed. The EEG signals in metaphit rats appeared as a sleep-like pattern and spike-wave complexes with increased power spectra. Valproate and DSIP reduced the incidence of seizure and prolonged duration of latency in a dose-dependent manner. ED50 of valproate in the 1st hour after administration was 63.19 mg kg(-1) and that of DSIP 3.19 mg kg(-1) four hours after injection. This suggests that VPA, reached a peak of action immediately after the application, while DSIP had a prolonged action, mildly reducing, but not abolishing metaphit seizure. None of the applied VPA and DSIP doses eliminated the metaphit-provoked EEG signs of epileptiform activity.

[The delta-sleep inducing peptide and its effect on the electroencephalogram and power spectrum density in rats with metaphit-induced epilepsy]. / Delta-peptid spavanja i njegov uticaj na elektroencefalogram i spektralnu gustinu snage kod pacova s metafitskom epilepsijom.

INTRODUCTION: Sleep has many common features with epilepsy (spontaneously, recurring event and EEG hypersynchrony including EEG potentials that look very similar to epileptiform sharp waves) [1]. Monnier et al. [4] reported the presence of a sleep-inducing factor inducing sleep with predominant EEG activity in the 8 band (1-4 Hz), and it was the reason for the term delta sleep-inducing peptide (DSIP). Metaphit was synthesized by Rafferty et al. (1985) [7] and was shown to increase general brain excitability and induce audiogenic seizures in small rodents. The effects of a natural somnogenic nonapeptide DSIP on metaphit-induced audiogenic epilepsy in rats were studied with the aim of shedding more light on answering the question whether DSIP could be included in the list of antiepileptic agents. MATERIALS AND METHODS: Adult, 2-month-old male Wistar rats (170-200 g) were used. None of the animals screened for audiogenic susceptibility showed seizure activity. Audiogenic stimulation was used for 60 s using an electric bell (100 +/- 3 dB 5-8 kHz). Rats were divided into four groups: 1. Control, saline-injected (n = 6); 2. metaphit administered (10 mg/kg; n = 12); 3. metaphit + DSIP (1 mg/kg), (n = 14) group, DSIP administered after 8th to investigate blocking effect on fully developed metaphit seizure. 4.DSIP alone (1 mg/kg, n = 6). RESULTS: In control saline-injected animals AGS provoked no convulsive response. Metaphit injection produced after 30 min initial EEG changes in the form of synchronized spikes and fast high-voltage activity that are typical seizure manifestations, power spectra increased and became more intense in the period of sound onset and seizure events. Our results demonstrate that DSIP acted increasing the EEG output in the 8 range and significantly elevated the mean power spectra in all checked experimental points. Besides, DSIP decreased the incidence and duration of convulsive component, as well as mean seizure grade in metaphit-induced seizures. DISCUSSION: Metaphit induces a generalized, reflex epilepsy thus providing an experimental model of choice for the studies of the mechanism of epilepsy development and blockade of NMDA/PCP receptors. In our previous studies a competitive NMDA antagonist CPP [9] and a noncompetitive antagonist MK-801 [8] were used. Non-competitive, selective NMDA antagonists MK-801, PCP and ketamin expressed a partial agonist motor action (myoclonic jerks, ataxia and tremor of the whole body) in audiogenic epilepsy prone mice. DSIP produced no harmful effects even when overdosed or any effect over "normality" [4, 5]. DSIP has a capacity of suppressing various forms of convulsive activity in different animal species. It was suggested that it exerts an anticonvulsant action by influencing neurotransmitter (dopaminergic, adrenergic, GABA-ergic) and neuromodulator (peptidergic) brain systems [12, 13]. CONCLUSION: Our results, together with the fact that DSIP penetrates through the blood brain barrier after systemic administration and that overdoses of this natural peptide produce no harmful effects, strongly suggest that it could be an important therapeutic agent for the treatment of sleep disturbances. Also, our data demonstrating reduction in incidence, severity and duration of seizure components, suggest that this agent might be a suitable candidate as an antiepileptic drug.

N-Methyl-D-aspartic acid- and metaphit-induced audiogenic seizures in rat model of seizures.

The effects of NMDA (N-methyl-D-aspartic acid) on metaphit (1-[1(3-isothiocyanatophenyl)-cyclohexyl]piperidine)-induced audiogenic seizures in adult male Wistar albino rats were studied with the aim of developing a suitable animal model of seizures. The animals were divided into four experimental groups: 1, saline control; 2, metaphit-injected; 3, metaphit + NMDA administered and 4, NMDA-treated. Upon the treatment, the rats were exposed to sound stimulation (100 +/- 3 dB, for 60 s) at hourly intervals and the incidence and severity (running, clonus and tonus) of seizures were analysed. In group 3, only the animals which did not exhibit any metaphit-induced audiogenic seizures over 8 h were given a subconvulsive NMDA dose after the eighth audiogenic testing. For EEG recordings, three gold-plated screws were implanted into the rat skull. In most animals, metaphit led to EEG abnormalities and elicited epileptiform activity recorded as spikes, polyspikes and spike-wave complexes. Maximum incidence and severity of metaphit-induced convulsions occurred 8 h after injection (incidence 9/12), abating gradually until disappearing 30 h later. NMDA alone provoked no seizure response but the initial signs characterized by isolated spike activity evolving into sporadic slow-wave complexes, thus representing a proconvulsive brain state, were observed. This compound led to stereotyped behaviour seen as asymmetric posture, loss of righting reflex and tonic hind limb extension lasting for 60-90 min. It also potentiated metaphit-induced audiogenic seizures. Potentiation of metaphit-related audiogenic seizures by NMDA was recorded in three out of 17 rats that had never displayed seizures in eight previous testings, with a maximum incidence of eight out of 17 animals, 13-14 h after metaphit administration and seizures lasted for 10 h.

The effect of delta sleep-inducing peptide on the EEG and power spectra in rat.

The effects of delta sleep-inducing peptide (DSIP) on the EEG and power spectra of adult male Wistar rats (b.w. 180-220 g) were studied by power spectra analyses of EEG wave forms recorded continuously for 12 h after DSIP administration. The animals were given DSIP i.p. (1 mg/kg). Saline-injected rats served as the corresponding control. Recorded bursts of high amplitude EEG in the 1-9 Hz range (delta and theta) were found to be more frequent in DSIP-treated animals, while power spectra and (delta) wave activity were enhanced in comparison with the control and a statistically significant increase was registered in all experimental points after DSIP (2 h P < 0.05; 4 h P < 0.05; 5 h P < 0.05; 6 h P < 0.05; 7 h P < 0.01; 11 h P < 0.05). In addition, DSIP significantly elevated both the EEG output in the (delta) range and sleep activity. These results suggest that DSIP should be considered as a potential agent for the treatment of sleep disturbances in human medicine.

[Potentiation of metaphit-induced audiogenic epilepsy with N-methyl-D-aspartate in rats]. / Potencijacija audiogene metafitske epilepsije en-metil-de-aspartatom kod pacova.

INTRODUCTION: Audiogenic seizures (AGS) are induced by high intensity sound stimulation in genetically susceptible rats or in animals subjected to chemical or electrical manipulation. Epileptic seizure may result from an impaired balance between excitation and inhibition in the CNS. The effect of NMDA (N-methyl-D-aspartic acid) on metaphit 1-(1(3-isothiocyanatophenyl-ciclohexyl)-piperidine) induced audiogenic seizures was evaluated in rats. METHODS: Male Wistar albino rats were divided into 4 groups: 1) saline; 2) metaphit (10 mg/kg); 3) metaphit + NMDA; 4) NMDA (70 mg/kg). Animals were injected with metaphit intraperitoneally (i.p.) and exposed to sound stimulation (100 +/- 3 dB, 60 s) at hourly intervals. The incidence and severity (running, clonus and tonus) of seizures were analyzed. NMDA alone was administered i.p. to 6 rats. In group metaphit + NMDA only animals which did not exhibit any seizure during 8 hours were injected with NMDA i.p. after the 8th audiogenic testing. For electroencephalograph (EEG) recordings three gold-plated screws were used. Convulsive behaviour was assessed by incidence of motor seizure and by seizure severity grade, determined by use of a descriptive rating scale with range of 0-3; 0-no response; 1-wild running only; 2-wild running followed by clonic seizures of all four limbs with body rollover; 3-wild running progressing to generalized clonic convulsions and then a tonic extension of the fore and hind limbs and tail. Sound onset, seizure events, and sound offset, along with the animals behaviour (convulsive or other) were recorded as the correlates to the respective EEG responses. RESULTS: In most animals the administration of metaphit (10 mg/kg) resulted in electrographic abnormalities, elicited epileptiform activity in the form of spikes, polyspikes and spikewave complexes (Fig. 1.). Maximum incidence and severity of metaphit convulsions occurred 8 h after the injection (9/12, 75%) (Fig. 2, 3.), then abated gradually and disappeared 30 h later. NMDA (70 mg/kg) alone induced no seizure response but isolated spiking activity, and sporadic slow-wave complexes were recorded (Fig. 4). NMDA induced stereotyped behaviour in the form of asymmetric posture, loss of righting reflex and tonic hindlimb extension, which lasted for 60-90 min. Subconvulsive dose of NMDA potentiated the metaphit-induced audiogenic seizures in rats. Two hours after NMDA administration 3 of 17 metaphit-treated rats convulsed, which in 8 previous testings never displayed seizures. Maximum incidence was 8 of 17 (53%), 5-6 h after NMDA administration and seizures lasted for 9 hours. DISCUSSION: Several authors reported that metaphit dose of 10 mg/kg accompanied by some REM sleep deprivation (REM-D) procedures [4], or subconvulsive doses of NMDA [25] provoked seizures of higher intensity and incidence. Metaphit treatment (10 mg/kg) followed 24 h later by NMDA dose of 50 mg/kg provoked no spontaneous convulsions, while metaphit in combination with a higher NMDA dose of 70 mg/kg resulted in spontaneous and AGS-induced seizures only in one time point [25]. It was found that the incidence and severity of convulsive responses were highest 8-12 h after metaphit injection (10 mg/kg) [23, 24]. Although about 8 h after metaphit administration the power spectra increased and were more intense in the period of sound onset and seizure events. CONCLUSION: The results of the present study strongly suggest that treatment of adult rats with the combination of metaphit and NMDA in the doses employed here followed by AGS provides a suitable animal model for examinations of epileptic seizures.

Inhibition of metaphit-induced audiogenic seizures by APV in rats.

The influence of APV ((+/-)-2-amino-5-phosphonovaleric acid) on EEG activity and behavior was studied on a model of epilepsy induced by intraperitoneal administration of metaphit (1-(1-(3-isothiocyanatophenyl)-cyclohexyl)-piperidine). Male Wistar rats received an injection of metaphit (10 mg/kg) and were subjected to intense audio stimulation (100+/-3 dB, 60 s) at hourly intervals during the experiment. The seizures were classified according to a four point scale ranging from 0 (no seizure) to 3 (tonic convulsions). In our report we studied the time course which revealed the maximum incidence and severity of seizures 7-12 h after the injection (10 out of 12 rats, with severity of 2.25+/-0.32). APV (0.05, 0.1, 0.2 and 0.3 micromol) was injected intracerebroventricularly at the time of fully developed convulsions. APV inhibited seizures in a dose-dependent manner. The minimum dose, which completely blocked seizures in all animals, was 0.3 micromol, while ED50 were 0.11, 0.10 and 0.07 micromol against running, clonus and tonus, respectively. In contrast to behavioral inhibition of convulsions, metaphit-provoked epileptiform activity was not abolished by APV, and represented a prerequisite for the reappearance of behavioral seizures. It is suggested that APV is rather an anticonvulsant than an antiepileptic agent in this model of epilepsy.

The anticonvulsant potency of (+/-)-2-Amino-7-phosphonoheptanoic acid and (+/-)-2-amino-5-phosphonovaleric acid against metaphit-induced audiogenic seizures in rats.

Male Wistar albino rats were subjected to sound stimulation (100+/-3 dB, 60 s) at hourly intervals after intraperitoneal injection of metaphit (10 mg kg-1). The incidence and severity of audiogenic convulsions increased with time, reached a peak 7-12 h after metaphit administration (ten out of 12 and 2.25+/-0.32), and then gradually decreased until 31 h post-injection when no animal displayed signs of seizure. In order to test anticonvulsant activity on fully developed seizures, antagonists were delivered intracerebroventricularly after the 8th testing. The doses of AP7 were 0.005, 0.01, 0.02, 0.03 and 0.05 micromol, and 0.05, 0.1, 0.2 and 0.3 micromol of AP5, all in 5 microliters of physiological saline. Antagonists inhibited metaphit-induced audiogenic seizures in a dose-related fashion. The minimum doses that completely abolished convulsions were 0.03 and 0.3 micromol for AP7 and AP5, respectively. For suppression of running, clonus and tonus AP7 was 16-18 times more potent than AP5. These results indicate that AP7 is substantially more potent than AP5 against all phases of metaphit-induced audiogenic seizures in rats. (c) 1998 The Italian Pharmacological Society.

APH, an N-methyl-D-aspartate receptor antagonist, blocks the metaphit-induced audiogenic seizures in rats.

The effect of the competitive antagonist of the N-methyl-D-aspartate (NMDA) receptor, (+/-)2-amino-7-phosphonoheptanoic acid (APH) on electrocorticographic (ECoG) activity and behavior was studied in the model of epilepsy induced by systemic application of metaphit (1-(1-(3-isothiocyanatophenyl)-cyclohexyl)-piperidine). Male Wistar rats were injected with metaphit intraperitoneally (10 mg/kg, i.p.), and exposed to intense audio stimulation (electric bell generating 100 +/- 3 dB at animal level for 60 s) 1 h after administration and at 1-h intervals thereafter. ECoG tracings showed appearance of paroxysmal activity in form of spikes, spike-wave complexes and ECoG seizures. Audiogenic seizures consisted of wild running followed by clonic and tonic convulsions. Each behavioral seizure response had a characteristic ECoG correlate. The incidence and severity of seizures increased with time, reaching a peak 8-12 h after metaphit administration, and then gradually decreased until 31 h, when no animal responded to sound stimulation. APH was injected intracerebroventricularly (0.005, 0.01, 0.02, 0.03 and 0.05 mumol icv in 5 microL of sterile saline) after the 8th hour of audiogenic testing (AGS). APH inhibited seizures in a dose-dependent manner. The minimum dose which blocked seizures in all animals was 0.03 mumol. However, ECoG signs of seizure susceptibility were not suppressed by APH. After varying periods of time, behavioral seizures reappeared. It seems that APH blocks epileptiform propagation, but has less influence on the epileptogenic activity caused by metaphit.