Effect of diphenylhydantion on cortical epileptogenic foci in the rat

In acute experiments we studied the effect of diphenylhydantoin (DPH, 60 mg/kg i.p.) on the activity of cortical penicillin foci in 81 male rats. DPH reduced the discharge frequencies (measured by means of histograms of the intervals between adjacent discharges) and limited the projection of the discharges into non-primary cortical areas. The results for the administration of DPH before formation and after stabilization of the focus were qualitatively the same. DPH statistically significantly slowed down synchronization of the activity of two symmetrical cortical foci, but did not affect the progress of the synchronization of two asymmetrical foci, which was also very slow in the controls. DPH did not inhibit the possibility of triggering focal discharges by peripheral nerve stimulation. A detailed evaluation of focal activity allows the anti-epileptic effect of DPH to be demonstrated even in a model as potent as the penicillin focus in animals without general anaesthesia.     

The role of the corpus callosum in the interhemispheric transmission of epileptic electrical activity.

In acute experiments on 49 curarized adult rats without general anaesthesia, we studied the transmission of discharges of cortical penicillin foci between the two hemispheres after transecting the corpus callosum. Projected discharges of the cortical penicillin focus appeared in the contralateral hemisphere later than in the controls and had a very different shape. The interhemispheric response of the experimental rats consisted of a small positive and a small negative deflection with long latent periods. Focal discharges could be triggered by electrical stimulat;on of the contralateral hemisphere only irregularly and for short periods of time. In rats with a transected corpus callosum, two symmetrical cortical foci at first behaved independently of each other; their synchronization then slowly improved, but never attained 100 per cent. The corpus callosum is the preferential pathway for interhemispheric transmission of focal activity. Transection of this pathway makes the transmission conditions much worse, but further connections, with a longer conduction time and lower efficacy, gradually come into action.     

Influence of clonazepam on cortical epileptogenic foci in the rat

The antiepileptic action of clonazepam was studied on epileptogenic foci induced by penicillin in sensorimotor cortex in acute experiments in rats. Clonazepam (1 mg/kg intraperitoneally) only moderately decreased the frequency of interictal discharges of single cortical focus and delayed the propagation of discharges into the ipsilateral occipital region. On the contrary, clonazepam failed to influence the callosal projection of interictal discharges in single unilateral as well as in two symmetrical foci. Spontaneous transition of interictal discharges into ictal phases regularly seen when two symmetrical foci were formed was only delayed but not blocked by clonazepam. It may be concluded that clonazepam exhibits only a weak anticonvulsant action against cortical foci and against secondary generalization of epileptic activity.     

Influence of dietary fat on the promotion of diethylnitrosamine-induced hepatocarcinogenesis in female rats

The effect of varying the amount and type of dietary fat on the promotion of gamma-glutamyltranspeptidase (GGT)-positive foci and hepatocarcinomas in female rats was studied. In the first study, two-thirds of the rats were first intubated with diethylnitrosamine (DEN, 10 mg/kg) 20 hr after partial hepatectomy; 1 week later, rats were fed one of three purified diets (a low-fat diet similar to the AIN-76 diet, a high saturated fat diet, or a high polyunsaturated fat diet) with or without 0.05% phenobarbital in the diet for 10 months. Increasing the fat level of the diet did not increase the number of GGT-positive foci arising spontaneously or induced by DEN alone. When phenobarbital was present in the diet, feeding the high polyunsaturated fat diet slightly increased the number of GGT-positive foci and the incidence of tumors. The low-fat diet, however, increased the incidence of fatty liver. We therefore reexamined the effect of diet on promotion by phenobarbital, using an additional low-fat diet with cornstarch rather than sucrose as the carbohydrate source. In this experiment, both high-fat diets slightly enhanced the induction of GGT-positive foci; the carbohydrate source had no effect. The incidence of tumors was not affected by diet in this experiment, but the incidence of fatty liver was again enhanced by feeding a diet high in sucrose. We conclude that increasing the fat level of the diet does not promote the development of DEN-initiated GGT-positive foci or carcinomas in female rats. Increasing the dietary fat level, however, may enhance promotion of liver foci by phenobarbital. Finally, increasing the sucrose content of the diet increases the incidence of fatty liver.     

Producing the cortical phenomenon of "kindling" during acute experiments on cats

Acute experiments on cats enabled spontaneous epileptogenic foci to be produced by means of electrical stimulation of the cerebral cortex. The central area of the suprasylvian gyrus was stimulated by 5-sec stimuli at 3-sec intervals. The strength of neuronal response gradually increased until spontaneous electrical activity set in as stimulation was set at a fixed intensity which had originally evoked local residual discharge at stimulation foci only, i.e., the phenomenon of "kindling" was observed. When "kindling" was produced in the cortical area under study, bursts of spindle activity were recorded on all ECoG with increasing frequency. Recordings of spindle activity then changed to a "spike-wave" pattern of activity. The results of these investigations, which were performed on an isolated strip of cortex, point to the involvement of subcortical structures in mediating "kindling" of the cortical focus.Institute of Clinical and Experimental Neurology, Ministry of Public Health of the Georgian SSR, Tbilisi. Translated from Neirofiziologiya, Vol. 17, No. 5, pp. 601–606, September–October, 1985.     

Effect of nicotinamide on epileptic activity in the cerebral cortex

The experiments on cats showed that intravenous administration of nicotinamide suppresses the epileptic activity in a solitary epileptic focus as well as in the complex of epileptic foci produced by strychnine application to various cortical zones under the influence of the most powerful focus that plays the role of a determinant. After the intravenous injection of nicotinamide (50-70 mg/kg) the complex was destabilized and broken down. The epileptic activity in the dependent foci of the complex disappeared first in the more remote from the determinant focus and then in the nearer one. The determinant focus was the last to disappear. The inhibitory effect of nicotinamide is associated with antiepileptic activity. Nicotinamide is suggested to be one of the endogenous drugs which may suppress brain hyperactivity and activate the antiepileptogenic system.     

Comparison of phenobarbital-and carcinogen-induced aldehyde dehydrogenases in the rat

There is a genetically determined variation in the inducibility of a high-K_m cytoplasmic aldehyde dehydrogenase activity in the rat liver by treatment with phenobarbital. In the present experiments this activity increased after phenobarbital administration in the phenobarbital-responsive rats also in the intestinal postmitochondrial supernatant fraction. Phenobarbital-nonresponsive rats did not exhibit such an increase after drug treatment. Intraperitoneal administration of 2,3,7,8,-tetrachlorodibenzo-pdioxin, strongly enchanced the cytoplasmic enzyme activity in the liver of both responsive and nonresponsive rats. This effect was also seen in the serum but not in the intestinal or hte kidney. Intragastric administration of 3-methylcholanthrene, 3,4,-benzpyrene or chrysene induced the activity in liver and intestine but not in serum or kidney. The activity in liver was also induced by long-term feeding with 2-acetamido-fluorene. The activities induced by tetrachlorodibenzodioxin or the carcinogens had similar behaviour in isoelectric focusing in gel slabs and in gel chromatography, suggesting a possible common identity of these induced enzymes. The activity induced by these agents could be clearly differentiated both from the activity induced by phenobarbital and from the normal cytoplasmic activities.     

Effects of steroid hormones and xenobiotics on the pubertal development of UDP-glucuronosyltransferase activities towards androsterone and 4-nitrophenol in Wistar rats.

Oestradiol benzoate, testosterone propionate, progesterone, corticosterone, 3-methylcholanthrene and phenobarbital were administered to Wistar rats at the pubertal period, and their effects on hepatic UDP-glucuronosyltransferase activities were determined. Pretreatment with oestradiol benzoate had a temporary suppressive effect on androsterone UDP-glucuronosyltransferase activity in rats with the high-activity phenotype of androsterone glucuronidation. The effect was marked in 40-day-old rats, but was not found in older rats. Androsterone UDP-glucuronosyltransferase activity was induced by phenobarbital in rats with the high-activity phenotype, but not in rats with the low-activity phenotype. Foster-feeding experiments showed that breast milk did not alter the genetically determined expression of androsterone UDP-glucuronosyltransferase activity in Wistar rats. In contrast, 4-nitrophenol UDP-glucuronosyltransferase activity was not affected by steroid hormones, but was highly induced by 3-methylcholanthrene.     

The participation of cortical brain areas in the processes of perceiving and reproducing human emotional states]

New method of mapping intracortical interactions was used to study the participation of cortical brain areas in the processes of perception and of mental reproduction of emotional states in humans. When an emotion was identified, the activity focus was observed in the left temporal cortex. If emotion was not identified, the temporal focus did not appear, but activity foci were seen in frontal regions of both hemispheres. When emotional states were mentally reproduced, activity foci were encountered mostly in the frontal cortical areas.     

Effects of phenobarbital and 3-methylcholanthrene on the hepatic cytochrome P-450 metabolism of various alkoxyresorufin ethers in the cotton rat (Sigmodon hispidus)

1. Microsomes isolated from phenobarbital and 3-methylcholanthrene induced cotton rats (Sigmodon hispidus) were tested for o-dealkylase activity with methoxy-, ethoxy-, pentoxy- and benzoxyresorufin ethers. 2. The activity of 3-methylcholanthrene induced microsomes was greater than controls. 3. Activity of phenobarbital induced microsomes was not different from controls. 4. There was a distinct difference between male and female animals. 5. The results obtained from cotton rats are markedly different from results obtained from Sprague-Dawley (S-D) rats.     

Stability of microsomal mono-oxygenase during incubations for the liver microsomal assay with S9 fractions of mouse liver under various inductions

Aminopyrine-N-demethylase and p-nitroanisole-O-demethylase activities were determined in incubation mixtures for the liver microsomal assay at time zero and after 1 h of incubation in the conditions for the mutagenic assay. The experiments were performed with the S9 liver fraction of mice in the basal state and induced with sodium phenobarbital, β-naphthoflavone or both. Lipid peroxidation was also determined.

The experiments were repeated with female mice and also in the presence of styrene, as an example of a xenobiotic substance. The activity of pNAD was much more stable than that of APD in all the conditions tested. The pattern of stability, however, was similar for the two activities: more stable than controls with S9 fractions from β-NF-induced mice, less stable than controls in PB-induced mice, intermediate between controls and PB-induced mice in those with combined induction by PB + βNF. Styrene 50 mM in the incubation mixtures led to a marked inactivation of enzymic activity, similar in all cases and reaching about 90% in 1 h. S9 fractions from female mice gave enzymes slightly more stable in almost all cases. Lipid peroxidation was appreciably more elevated in basal than in induced animals.

It was concluded that, for a mutagenesis test on an unknown xenobiotic, S9 fractions from mice following PB and β-NF induction are to be preferred from the point of view of activation.     

Formation of epileptic activity complexes under the effect of the determinant focus in the cortex isolé]

In experiments on cats foci of enhanced excitation working in independent regimes were created in the neuronal-isolated cortex using weak strychnine solutions. Creation of a hyperactive focus by means of application of strychnine concentrated solutions or crystals leads to an increase in the amplitude and discharge frequency in other foci, synchronism in discharges of these foci and their unification into a single functional complex, working in the hyperactive focus regime. The latter, in such a way, presented the determinant structure. Under the influence of the determinaant focus there was also a marked generalization of the convulsive activity manifested by an embrace with epileptical discharges of the cortex area intact from strychnine. The results obtained indicate that the established relations between the foci of hyperactivity in the cortx as well as the effects of the determinant structure may be realized by the cortex mechanisms proper.     

Modeling of determinant and dependent foci of epileptic activity in the rat cerebral cortex]

Foci of increased excitability were created in acute experiments on rats by means of weak strychnine solutions working at independent regimens. The hyperactive excitability focus induced by means of concentrated strychnine solutions played the role of a determinant structure. The importance of the latter is in the fact that it determines the activity character of other epileptogenic foci, enhances their convulsive activity, unites them into a single functional complex and determines the behaviour of the complex as a whole. This complex can be destroyed by depression of the determinant focus activity. Switching off any dependent foci included into this complex fails to destroy that latter. Results of the investigations confirmed on the new model the general concept of the role played by the determinant structures in the brain activity.     

Formation of epileptic activity complexes in the cerebral cortex as a result of a determinant focus induced by acetylcholine]

Foci of increased excitability were created by means of weak strychnine and penicillin dilutions in experiments on cats. These foci worked at individual regimens. Creation with acetylcholine and proserine of a hyperactive focus led to increase of the amplitude and frequency of convulsive discharges at first in the nearest activity foci, and then in the ones remote from the hyperactive focus. The next stage was attended by qualitative changes in the activity pattern of strychnine and penicillin foci (the appearance of acetylcholine activity in them) and by the formation of a single functional focal complex working in the regimen of acetylcholine focus. Thus, the latter played the role of a determinant structure. Suppression of the determinant focus activity led to disappearance of the acetylcholine activity in all the other foci, restoration of the initial (penicillin and strychnine) activity, and to the epileptic complex decay.     

Electroencephalographic study of the effect of neurotoxin DSP-4 in iron model of chronic focal epilepsy.

The effect of the noradrenergic neurotoxin N-(2-chloroethyl)-N-ethyl-2-bromobenzylamine (DSP-4) on electroencephalographic activity (EEG) was studied in the model of chronic focal epilepsy induced by intracortical injection of FeCl3 in the rat. EEG activity was recorded from the epileptogenic focus (ipsilateral and contralateral) in chronic experiments before and after DSP-4 treatment. In some experiments EEG activity was also simultaneously recorded from the cortical epileptogenic focus and locus coeruleus before and after DSP-4 treatment to study the effect of iron-induced seizure activity and of DSP-4 on the locus coeruleus electrical activity. The results showed that DSP-4 aggravated the iron-induced epileptiform activity as well as the locus-coeruleus electrical activity. The data also showed that, induction of epilepsy by FeCl3 is accompanied by enhancement of the locus coeruleus electrical activity. Our study demonstrates that DSP-4 intensifies and modifies the epileptic activity in the iron-induced chronic epilepsy model and that the effects of toxin persist for a longer duration.     

Expression of four phenobarbital-inducible cytochrome P-450s in liver, kidney, and lung of rats

Specific antibodies were prepared against cytochromes P450 PB-1, PB-2, PB-4, and PB-5 purified from hepatic microsomes of male rats treated with phenobarbital. With these antibodies, the levels of these four cytochrome P450s in hepatic, renal, and pulmonary microsomes of male rats that were untreated, treated with phenobarbital, or treated with 3-methylcholanthrene were examined. P450 PB-1 and PB-2 were present in moderate amounts in hepatic microsomes of untreated male rats and were induced 2- to 3-fold with phenobarbital. Also, the expression of these forms was suppressed by 3-methylcholanthrene. These forms were not detected in the renal or pulmonary microsomes of untreated rats or rats treated with phenobarbital or 3-methylcholanthrene. P450 PB-4 and PB-5 were found in the hepatic microsomes of untreated male rats at a low level but were induced with phenobarbital more than 50-fold. P450 PB-4 and PB-5 were not detected in renal microsomes; only P450 PB-4 or a closely related form was present in the pulmonary microsomes of untreated male rats, and its level was not changed by phenobarbital treatment. The constitutive presence of P450 PB-4 in pulmonary microsomes was confirmed by the investigation of testosterone metabolism. Purified P450 PB-4 had high testosterone 16 alpha- and 16 beta-hydroxylation activity in a reconstituted system. The testosterone 16 beta-hydroxylation activity of hepatic microsomes was induced with phenobarbital, and more than 90% of the testosterone 16 beta-hydroxylation activity of hepatic microsomes from rats treated with phenobarbital was inhibited by anti-P450 PB-4 antibody.(ABSTRACT TRUNCATED AT 250 WORDS)     

Potentiation of thioacetamide hepatotoxicity by phenobarbital pretreatment in rats. Inducibility of FAD monooxygenase system and age effect

The ability of phenobarbital to induce the expression and activity of microsomal drug monooxygenases in the liver presents one of the most important issues in the field of chemical interactions and in the toxicity of xenobiotics. The model of rat liver injury induced by a single dose of thioacetamide (500 mg/kg intraperitoneally) was used to study the effect of phenobarbital (80 mg/kg/day intraperitoneally) for 5 days prior to thioacetamide. Serum parameters of liver injury such as aspartate aminotransferase activity, gamma-glutamyl transferase activity and the total bilirubin levels, as well as the activities of hepatic FAD and cytochrome P450 microsomal monooxygenases, were assayed in 2- and 12-month-old rats. Samples of blood and liver were obtained from controls (injected at 0 h with 0.5 ml of 0.9% NaCl) and at 12, 24, 48, 72 and 96 h of thioacetamide intoxication either to non-treated or phenobarbital pretreated rats. Potentiation of thioacetamide hepatotoxicity by phenobarbital pretreatment was demonstrated at morphological level, and by significant increases in the activities of serum aspartate aminotransferase and gamma-glutamyl transferase, and in the levels of total bilirubin. The extent of potentiation of thioacetamide-induced liver injury by phenobarbital pretreatment was similar in both age groups. Microsomal FAD monooxygenase activity, the enzyme responsible for thioacetamide biotransformation, was significantly enhanced (twofold) by phenobarbital pretreatment, and also underwent a further increase following thioacetamide, preceding the peak of necrosis. Cytochrome P450 monooxygenases were induced by phenobarbital pretreatment more than sixfold, and sharply decreased when phenobarbital was withdrawn and thioacetamide administered, showing at 48 h intoxication values close to basal. Phenobarbital pretreatment potentiated thioacetamide necrogenicity, and this potentiation was parallel to the induction of the microsomal FAD monooxygenase system, both by phenobarbital and by thioacetamide itself. The extent of thioacetamide-induced liver injury was significantly higher in 12-month-old rats, but the effect of phenobarbital pretreatment was similar in both age groups.     

Electrocorticographic development of epileptogenic foci in the occipital region of the rat cerebral cortex

The development of cortical penicillin foci in the occipital region was studied in rats whose ages ranged from five days up to the adult age. The local application of penicillin induced the formation of an epileptogenic focus for the first time at the age of seven days. With advancing age, the amplitude of focal discharges increased, the duration of the individual components of the discharge shortened, its originally negative-positive configuration changed to a triphasic form and in the third week of life initial positivity, for a time, become the dominant component of the discharge. Projection of the discharges to the contralateral hemisphere was found to be inconstant in the second postnatal week, but appeared regularly from the age of 14 days. Synchronization of the discharges of two symmetrical foci was very poor in 7-day-old young, but improved noticeably by the 14th day; it was never complete, however, even in adulthood. The activity of symmetrical foci changed spontaneously to ECoG seizures, which were most common in 7-day-old young (in which ictal activity was usually not generalized, however) and were least frequent in 14-day-old animals. Focal discharges could not be reliably triggered by electrical stimulation of the contralateral cortex until the age of 18 days and later. The occipital part of the cortex develops somewhat later than the sensorimotor, frontal region, and during its development there also appeared phenomena which are not present in the frontal cortex.     

Long term phenobarbital administration does not promote the multiplication of hepatocytes replicating after single cyproterone acetate administration

Cyproterone acetate (CPA) is a synthetic antiandrogenic compound which is widely used in clinic but suspected to be hepatocarcinogenic. CPA is also mitogenic in rat liver. Using genetic labeling of dividing cells, we examined whether hepatocytes dividing in response to acute CPA administration could give rise to preneoplastic foci after administration of a tumor promoter: phenobarbital. CPA was administered orally to rats and dividing hepatocytes were genetically labeled using retroviral vectors carrying the beta-galactosidase gene. After labeling rats were given phenobarbital for 10 months and sacrificed. The presence of beta-galactosidase labeled hepatocytes as well as preneoplastic hepatocytes was assessed by immunohistochemistry. Genetic labeling of hepatocytes was obtained in all animals. At the end of phenobarbital administration, no hepatic tumors were observed. Preneoplastic foci were not increased in treated animals as compared to control rats. Moreover beta-galactosidase positive hepatocytes were never detected in preneoplastic foci. Finally, the size of the beta-galactosidase positive clusters was smaller in treated animals as compared to control rats. We conclude that acute CPA administration is not carcinogenic in rat liver and does not initiate preneoplastic hepatocytes capable to give rise to foci after phenobarbital promotion. Therefore the mitogenic property of CPA is distinct from its putative carcinogenic activity. Finally, analysis of the size of beta-galactosidase positive cells clusters demonstrate that phenobarbital does not induce hepatocyte proliferation in rats.     

Influence of the sensorimotor cortex on single neurons of the nucleus gracilis in the cat

The aim of this study was to investigate the functional role of the cortical projections to gracile nucleus. In unanesthetized cats single nuclear units projecting to the thalamus were tested for microstimulation of cortical foci (area 4) able to evoke single joint movements in contralateral hindlimb. A very significant percentage of gracile cells was influenced, very often in excitatory manner, if their receptive field was overlaying or very close to the joint controlled by a given cortical focus. Conversely, when the location of the receptive field was more distant, the percentage of responses and the incidence of excitatory effects decreased, inhibitions occurring more frequently. From a functional point of view, such an organization of the cortico-gracile control could be effective in modulating transmission of exteroceptive information from the region of the motor target (facilitation) as well as from adjacent ones (suppression). This arrangement could provide an higher resolution of afferent messages, in relation with the cortically induced movements.