Elucidation of the relationship between pregnancy and epilepsy remains one of the actual problems in neurology [1] -[3] . The mechanisms responsible for hormonal modulation of seizure susceptibility, however, are still poorly understood as it has been shown that women can be either seizure-sensitive or seizure-nonsensitive to sexual steroids [4] [5] depending on the phase of menstrual or estrous cycle in epileptic subjects. Researchers paid relevant attention to the importance of steroidal hormones in epilepsy after Gowers W.R. [6] had described the intensification of seizure reactions in women during menstruation. These observations led to a convention that reduction of seizures in a midluteal phase should be associated secondarily with progesterone secretion, whereas seizures’ intensification during menstruation is due to a reduction of progesterone level. It has been also demonstrated that administration of estrogen appears to exacerbate epilepsy, whereas progesterone and some of its five reduced metabolites have potent inhibitory effects on epileptic seizures [7] -[9] . However some studies showed that progesterone had no effect on kindling in the adult animal [10] .

Numerous studies over the last few decades have identified sex- and cycle-dependent differences in seizure susceptibility in rodents. These studies used various methods to induce seizures, including kindling, electroshock, and chemoconvulsant drug administration [11] -[13] . However, these studies failed to provide a clear picture of the relationship between ovarian hormone levels and seizure susceptibility.

It is worth to mention that, when such clinical observations are made, one has to bear in mind epilepsy type, localization of epileptic locus, etc. Quite often such approach lacks due attention. Besides, the study of interdependence between epileptic fits and pregnancy in humans is complicated from the ethic point of view and also because of anti-epileptic therapy. Therefore, one of the ways to investigate this problem is using laboratory animal models, in which interactions between seizures and the gestation can be studied under more controlled conditions than it is possible in human population.

Kindling is the animal model for human temporal lobe epilepsy. During kindling initial subconvulsive stimulation of limbic structures causes short-lasting electroencephalographic (EEG) paroxysmal discharges, without any behavioral seizure manifestations. In response to such stimulation the augmentation of EEG seizure discharges and their propagation in different brain structures develop, followed by generalization of seizure activity and behavioral manifestation [14] .

Genetically epilepsy-prone rats are used as an animal model of epilepsy. In this context the animals, which develop clonic-tonic behavioral seizure reactions to a strong sound stimulus, deserve special attention. The neuronal substrate of audiogenic seizure activity is localized in the nuclei of inferior colliculi [15] [16] .

There were two main objectives of the study: first, the clarify the effect of rat’s gestation and postgestation periods on already formed epileptogenic focus (model of kindling); second, the effect of gestation and postgestation periods on the development and course of audiogenic seizure activity.

Rats with body mass of 200 - 250 g were housed under the standard laboratory conditions with a “12 h light - 12 h dark” cycle, constant temperature of 22˚C ± 2˚C, and water and food ad libitum. Animal care and handling throughout the experimental procedures were in accordance with the European Community Council Directive of 24 November 1986 (86/609/EEC). Use of the animals in the experiments, animal care, and post-experimental euthanasia were performed in a strict adherence to the officially adopted rules for animal use and care in biological laboratories [17] .

Subject to the specificity of the study, experiments were carried out on female rats kept in isolation from males. Wistar (n = 16) and genetically audiogenic seizure-determined rats (n = 16) of Krushinski-Molodkina (KM) line were used in the study [18] .

The course of seizure activity is known to be altered with the day/night circadian rhythm [19] . Taking this into account, the observation on the course of seizure reactions was performed at the same time of a day (at 11.00 - 12.00).

Model of kindling. In female rats forty-fold electric stimulation (50 Hz, duration 5 sec, current 400 - 450 µA, at 5 min intervals) was applied to the ventral hippocampus (−4.8 mm from bregma caudally; 5.2 mm from the midline laterally; 6.5 mm from dura mater ventrally) during one day. At kindling, initial subconvulsive stimulation of the hippocampus evoked short-lasting (3 - 5 sec) EEG paroxysmal discharges, with no behavioral manifestations of seizures. The enhancement of EEG seizure discharges, propagation in various brain structures followed by generalization of seizures and behavioral manifestation occurred in response to every consecutive subconvulsive stimulation. The kindling state suggests formation of epileptogenic locus in the animal’s brain. At 24 h, 2 and 4 weeks after the procedure was completed 5 test stimulation was applied to the same structure with 5 min intervals. In the process of kindling the EEG and behavioral parameters of seizures were assessed. For this reason the Racine [20] scale: 0—nomal behavior, arrest; 1—shaking movement, facial clonus; 2—nodding of the head, backward deviation; 3—forepaw clonus; 4—standing on hindpaws, forepaw clonus; 5—fall on a side or on the back.

Audiogenic seizures. Genetically seizure-determined KM rats were placed in an audiogenic stimulation chamber. The chamber represented 60 × 60 × 60 cm plexiglass box, in the upper part of which a standard wall bell was attached. The animal was in the chamber and an audiogenic stimulus was delivered to it. A high pitch sound stimulus was presented to rats (bell intensity 110 dB, time 60 sec), in response to which they developed seizure reactions. Motor components of seizure activity were estimated by a slightly modified Jobe [21] scale: 0—fear reaction; 1—facial muscle clonus; 2—head tremble, jaw myoclonus; 3—wild run, forepaw myoclonus; 4—myoclonus of fore- and hindpaws, fall on a side; 5—clonus of the fourpaws, skeletal muscle rigidity, ataxia, asphyxia. The mentioned KM line rats fall into two sublines: a) animals, which, in response to a high pitch sound stimulus, develop fear reaction and facial muscle clonus—conventionally referred to as the first subline and b) animals developing fear and wild run to sound stimulus followed by clonic-tonic behavioral seizures— conventionally refered to as the second subline.

Audiogenic kindling. To obtain audiogenic kindling we used the KM line first subline rats—the animals, which, in response to sound stimulus, developed only fear reaction and facial muscle myoclonus. It is remarkable that they developed seizure activity evidenced in behavioral clonic-tonic responses after reiterative sound stimulus. It was shown that triggering of various brain structures (limbic and diencephalic structures, neocortex) occurs in parallel with such multiple repetition of sound stimulus. Involvement of various brain structures in seizure activity in response to sound stimuli is called “audiogenic kindling”—similar to the phenomenon described by Goddard [14] on reiterative stimulation of the limbic structures. In our experiments, in order to obtain audiogenic kindling, sound stimulus was delivered twice in the morning (10.00) and in the afternoon (16.00).

Ovariectomy. Ovariectomy surgery was performed in ether-anesthetized rats by bilateral excision of ovaries. Experiments were conducted 12 - 14 days after ovariectomy.

Ascertaining of rat’s gestation. The animal’s gestation was established examining ovarial smear (after administration of saline solution) by means of microscopic observation of spermatozoids and later by palpation.

Pseudogestation model. To receiving pseudogestation state in ovariectomized rats, progesterone (60 mg/kg) was injected intraperitoneally. Later on, proceeding from the goal of experiment, “knock-out” of allopregnanolone was performed by the intraperitoneal injection of finasteride, a blocker of progesterone metabolism (60 - 70 mg/kg).

The results obtained were statistically processed by the methods of descriptive statistics and paired 2-tailed t-test (α < 0.05). Data were considered valid with p < 0.05.

The data obtained in this study showed that 1) there is a reduction of both EEG and behavioral seizures during gestation in the presence of hippocampal epileptogenic locus (kindling model); 2) suppression of audiogenic seizures and the development of audiogenic kindling occurs in gestation; 3) substantial augmentation of seizures follows bilateral ovariectomy; 4) suppression of seizures occurs in the state of pseudogestation; 5) it is assumed that, in the gestation period the intensification of the endogenous inhibitory mechanisms should follow the course of seizure activity.