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D-20443 is an experimental antiepileptic drug. Its mechanism of antiepileptic action is unknown. We evaluated the anticonvulsant effectiveness of D-20443 against sound-induced seizures in genetically epilepsy-prone rats (GEPRs). This compound produced anticonvulsant effects against sound-induced seizures in moderate seizure GEPRs (GEPR-3s) at significantly lower doses than in severe seizure GEPRs (GEPR-9s). Based on these data and on the responses of GEPRs to other antiepileptic drugs, we predict that D-20443 will be a broad spectrum antiepileptic agent in humans. That is, we predict that D-20443 will suppress both tonic/clonic and absence seizures in humans.

The X gene of human hepatitis B virus encodes the polypeptide HBx which transactivates viral and host genes through a variety of cis-acting enhancer elements present in RNA polymerases I, II and III promoters. To better understand the mechanism of X transactivation, we cloned cDNAs of proteins that bind HBx. Here we demonstrate that one of these cDNAs is a full-length cDNA of human RPB5, a subunit shared by RNA polymerases. The HBx transactivation domain and the central region of human RPB5 were necessary for the specific binding of the two proteins as shown by: (i) in vitro assays using deletion mutants of fusion proteins; (ii) in vivo assays which detect associated proteins by co-immunoprecipitation of the non-fused proteins from transfected HepG2 cells. Over-expressed HBx seemed to associate with assembled forms of endogenous human RPB5 in HBx-transfected cells, since the endogenous RPB5 co-immunoprecipitated with HBx. The HBx binding region of human RPB5 by itself stimulated chloramphenicol acetyltransferase activities from several different reporters having X-responsive element(s). Our results support the idea that the interaction of HBx and human RPB5 can facilitate HBx transactivation and that human RPB5 has a domain which can communicate with transcriptional regulators.

This study was designed to demonstrate a role of serotonin in the anticonvulsant effect of fluoxetine, a serotonin reuptake inhibitor, in genetically epilepsy-prone rats. When varied doses of 5-hydroxytryptophan (12.5, 25, 50 mg/kg) were administered i.p. along with a fixed dose of fluoxetine (15 mg/kg) to severe seizure genetically epilepsy-prone rats, the severity of audiogenic seizures was decreased dose-dependently, and the combination treatment also produced a marked potentiation of the anticonvulsant effect when compared with administration of either drug alone. Pretreatment of severe seizure genetically epilepsy-prone rats with p-chlorophenylalanine depleted brain serotonin and reduced the anticonvulsant effectiveness of fluoxetine. By using intracerebral microdialysis, the depletion of serotonin after p-chlorophenylalanine treatment was confirmed by measuring thalamic extracellular serotonin and 5-hydroxyindoleacetic acid concentrations during basal release and in response to a challenge dose of fluoxetine. We concluded that serotonergic transmission may be involved in the anticonvulsant effect of fluoxetine in severe seizure genetically epilepsy-prone rats.

The human hepatitis B virus (HBV) X gene seems to be essential for establishment of viral infection, and the X gene product, HBx, transactivates virus and host genes through a wide variety of cis-elements, whereas regulation of HBx has not been fully understood. We found that transactivation-negative HBx mutants truncated at the C-terminal portion specifically repressed the HBx transactivation in trans. The ability to trans-repress the HBx transactivation is confined to the N-terminal third of HBx. Transactivation-positive constructs of HBx were divided into two groups by their sensitivity to trans-repression due to the presence of the N-terminal third. Thus the regulatory domain, the N-terminal third, is separated from the transacting domain and responsible for the negative regulations, the trans-repression and sensitivity to X trans-repression. A possible direct association between the HBx regulatory domains was tested by far-Western blotting using purified fused forms of HBx proteins. The regulatory domain was found to associate preferentially with the full HBx or the regulatory domain, but not with the transacting domain. Taken together, it is possible that HBx has a self-regulatory mechanism that avoids excessive HBx transactivation and is important for regulation of X gene expression.

Selectivity of lead effect on dopamine beta-hydroxylase activity in regions of brai nfrom rats postnatally exposed to lead was tested. Three groups of animals were prepared; (1) Rats exposed to lead at a low dose (0.05% PbAcetate: PbAc); (2) Rats exposed to lead at a high dose (0.2% PbAc); (3) Age-matched normal control rats. At 2, 4, 6 and 8 weeks of age weight of whole brain and body in each group were measured. At the same ages activities of dopamine beta-hydroxylase and Na+K(+)-ATPase were measured in 5 brain regions of each animal. Exposure of rats to lead generally decreased Na+/K(+)-ATPase activity and showed alternative changes of dopamine beta-hydroxylase activity were detected without concomitant changes of Na+/K(+)-ATPase activity were telencephalon and pons/medulla at 2 weeks of age and telencephalon, diencephalon and pons/medulla at 4 weeks of age and midbrain and pons/medulla at 6 weeks of age and cerebellum at 8 weeks of age in rats exposed to lead at a low dose, and those in rats exposed to lead at a high dose were midbrain at 6 weeks of age and cerebellum at 8 weeks of age. These data imply that noradrenergic nervous system in the brain regions described above could selectively be affected by lead.

Alterations of tyrosine hydroxylase activity in various regions of brain from rats postnatally exposed to lead were tested. Three groups of animals were prepared; (1) Rats exposed to lead at a low dose (0.05% lead acetate, PbAc); (2) Rats exposed to lead at a high dose (0.2% PbAc); (3) Age-matched normal control rats. At 2, 4, 6, and 8 weeks of age, weight of brain and body, and concentrations of lead in whole brain of animals in each group were measured. Activities of tyrosine hydroxylase and Na(+)-K+ ATPase were also measured at the same ages in 4 brain regions of each animal. Body weight gain was decreased after 6 weeks of age in rats exposed to lead at a high dose. Concentrations of lead in whole brain were increased from 0.37 to 0.83 (ng/mg wet tissue) in these animals. Exposure of rats to lead generally increased tyrosine hydroxylase activity and decreased Na(+)-K+ ATPase activity. However, changes of tyrosine hydroxylase activity were detected without concomitant changes of Na(+)-K+ ATPase activity in pons-medulla at 2 weeks of age and telencephalon at 6 weeks of age in rats exposed to lead at a low dose, and in midbrain at 4 and 6 weeks of age in rats exposed to lead at a high dose. These data imply that catecholaminergic nervous system in the brain regions described above could be selectively affected by lead.