Eiichi Katsumoto

Distribution of Fos- and Jun-related proteins and activator protein-1 composite factors in mouse brain induced by neuroleptics

The mechanisms by which the direct actions of neuroleptics are translated into therapeutic effects are unknown. We immunocytochemically investigated the expression of Fos- and Jun-related proteins and examined activator protein-1 DNA-binding activity in ddY mouse brain 120 min after the administration of haloperidol (1 mg/kg), (-)-sulpiride (20 mg/kg) and a selective dopamine D1 receptor antagonist, SCH23390 (1 mg/kg). The densities of Fos-, FosB-, Fra-1-, Jun- and JunD-immunoreactive nuclei induced by haloperidol and sulpiride in the hippocampus, piriform cortex and accumbens nucleus were higher than those in the control groups. The same regions showed higher densities of FosB-, Fra-1- and JunD-immunoreactive nuclei induced by SCH23390 compared with the control groups. We investigated further the activator protein-1 composite factors using super gel shift assays. These results suggested that induced Fos, FosB, Fra-1, Jun and JunD proteins constitute the activator protein-1 complex after the administration of haloperidol and sulpiride. In contrast, FosB, Fra-1 and JunD appear to constitute the activator protein-1 complex after the administration of SCH23390. Therefore, the diversity of activator protein-1 composite factors suggests that various kinds of gene are induced to act by some neuroleptics.

The comparative effects of haloperidol, (−)-sulpiride, and SCH23390 on c-fos and c-jun mRNA expressions, and AP-1 DNA binding activity

The c-fos and c-jun mRNA expressions were measured by the RNase protection assay method following intraperitoneal injection of haloperidol, (D1 and D2 receptor antagonists), (-)-sulpiride, (D2 receptor antagonist), and SCH23390, (D1 receptor antagonist). Haloperidol and (-)-sulpiride increased their mRNA expressions in a dose-dependent manner, peaking at 30 min after injection followed by a gradual decline. The SCH23390 did not induce expression of either c-fos or c-jun mRNA. A significant decrease of c-fos as well as c-jun mRNA expression was found due to pretreatment with SCH23390 (1 mg/kg i.p.) followed by injection of (-)-sulpiride (20 mg/kg i.p.). The results suggest that the expression of these mRNAs is closely related to the dopamine D2-like antagonism. Administration of haloperidol or (-)-sulpiride increased AP-1 DNA binding activity with similar manner of dose-dependence, whereas their activities were reduced by Fos and Jun antibodies, implying that AP-1 components, transcriptional factors, forming due to Fos and Jun were actually activated by either haloperidol or (-)-sulpiride.

Alterations of Glutamate Transporter Gene Expression in Epileptic EL Mice Brains

Intmduc%ion: Glutamate is considered to be an important excitatory amino acid and has been implicated in the pathophysiology of cpilcpsy. Glutamate transporters help to maintain the extracellular glutamate concentration and have been identified i n inousc brains, incl tiding GLT‐I and GLAST in astroglia and EAAC‐I in neurons. In an amygdala‐kindling study, alteration of glutamate transporter proteins showed that GLAST protein was down‐regulated in the piriform cortcx/amygclala rcgion of the amygdala‐kindled Iiits. In contrast, kindling induced an increase in EAAC‐ I levels in the pirilorrn cortex/amygdala and hippocampus. No changes i n GLT‐I wcrc obscrvcd in any region. The EL mouse, an inbred mutant strain, is considered to be a model for human complex partial seidures. This study was carried out to reveal the relationship bctwccn cpilcptogcncsis and glutamate transporters in EL mice.

Effects of a Pentylenetetrazol‐Induced Convulsion on Mouse Cortical Protein Kinase C and Ca2+/Calmodulin‐Dependenl Protein Kinase I1 Activity.

and CBZ, may possibly enhance the transformations of AEDs into epoxide, which may result in increased AED teratogenesis. On the other hand, the effects of AEDs on the detoxification or elimination process of epoxide have not yet been fully clarified. Our study clearly showed that therapeutic concentrations of VPA show inhibitory effects on the activities of EH and most of the GST isozymes, and that these effects are concentration dependent. Therapeutic concentrations of CBZ also inhibited the activity of GST 7-7, found primarly in the placenta. These results suggest that detoxification of the epoxide of AEDs could be inhibited under AED polytherapy by using VPA or CBZ or both. This may partly explain the increased incidence of malformations in offspring exposed to VPA-CBZ, administered with or without other AEDs. PHT, PB, and ZNS had no inhibitory effects on the activities of EH and GST. However, the possibility that PHT and PB have inducing effects on epoxide formation should be taken into account. In this regard, combinations of AEDs should be carefully selected in the treatment of women of childbearing age with epilepsy. (This work was supported by Grant-in-aid No. 05454309 for Scientific Research from the Ministry of Education and Culture and grant from the Hirosaki Research Institute for Neurosciences.)

Alteration of Fos-and Jun-Related Antigens After Seizures in Epileptic EL Mice.

analysis with anti-PKC-isozyme-specific antibodies was studied. The content of 80-K PKC /3 in the cytosolic fraction increased, whereas that in the membrane fractions decreased at 10 min after convulsion. The SO-K PKC /3 was detected in the cytosolic fraction during the tonic convulsion and 10 min after the convulsion phase. The findings in immunoblot analysis obtained by using anti-PKC a specific antibodies were similar to those obtained by using anti-PKC /3 specific antibodies, but those of PKC y with polyclonal proteins were not altered by the PTZ-induced convulsion. Our findings suggest that cytosolic PKC activity is increased by translocation of PKC a and p from the membrane to cytosol and release of 50-K PKC fragments. Although we could not demonstrate the mechanism of activation of membranous CaMKII by a PTZ convulsion, the activation of PKC in the cytosol and CaMKII activity in the membrane may be involved in the mechanism of the convulsion, such as termination of the convulsion or intracellular changes such as expression of immediate early genes.

The alteration of AP-1 DNA-binding activity in seizure-prone EL mouse brain.

The EL mouse being an inbred mutant strain is thought to be a model for human complex partial seizures.’V2 Our previous studies have shown that c

P-4-6 The effect of neuroleptics on the AP-1 DNA binding activity and the induction of fos and jun

os mRNA and Fos expressions in EL mouse brain by toss-up induced seizures prolonged more than those in EL and ddY mice by bemegride-induced ones.3 The results suggest that extended c

The difference among effects of haloperidol, (−)-sulpiride and SCH23390 on the expression of c-fos and c-jun mRNA inductions

os expression seems to be associated with seizuresusceptibility being reinforced by repeated seiz~res .~ Fos and Jun comprised homodimer or heterodimer, transcriptional factor of AP-1 which appeared to be AP-1 DNA elements concerning the promoter regions within target genes. This study was performed to survey AP-1 DNA-binding activities in EL and ddy mice brains after seizures and compare with those of interictal periods by gel retardation assay. Twenty-five week old mice and mice of various ages were used. Nuclear extracts were prepared from mice brain as described by Dignam et aL4 A 25 pg protein was subjected to gel retardation assay. The retardation assay was carried out using [32P]-dATP end-labeled 23-mer oligonucleotides (5’-TGATCCGGCTGACTCATCACTAG-3’) containing AP-1 consensus sequences. Gel was performed electrophoresis, then dried and autoradiographed. Figure 1 shows the difference of AP-1 DNA-binding activities, in particular the different seizures induced by toss-up and bemegride-induced injection. As seen in Fig. 1 a-d, the activities of toss-up induced seizures in EL[s] continued to be longer than those of bemegride induced seizures in two groups of EL and ddY mice. Figure 2 shows AP-1 DNA-binding activities during interictal periods of EL and ddY mice during the growth. The activities of third to fourth week after birth of EL mice were higher level compared with other age groups. The activities decreased with growth of ddY mice and attained to very low level on week 25. In contrast to this, EL held a higher expression than ddY throughout the age tested. Our present results are fairly consistent with those of Yoneda et al. who