Abstract: The atypical antidepressant, mirtazapine enhances noradrenergic transmission, but its effects on serotonergic transmission remain to be clarified. The present study determined the effects of acute and chronic administration of mirtazapine on serotonergic transmissions in raphe nuclei and their innervation regions, frontal and entorhinal cortex, using multiple-probes microdialysis with real-time PCR and western blotting. Acute administration of mirtazapine did not affect extracellular serotonin level in raphe nuclei or cortex; however, chronic administration increased extracellular serotonin level in raphe nuclei without affecting that in cortex. Blockade of 5-HT1A receptor, but not that of the 5-HT2A/2C receptor, enhanced the effects of acute administration of mirtazapine on extracellular serotonin level in raphe nuclei. Chronic mirtazapine administration reduced the inhibitory function associated with somatodendritic 5-HT1A receptor in raphe nuclei, but enhanced postsynaptic 5-HT1A receptor in serotonergic innervated cortical regions. Chronic administration reduced the expression of mRNA and protein of serotonin transporter and 5-HT1A receptor in raphe nuclei, but not in the cortices. These results suggested that acute administration of mirtazapine probably activated serotonergic transmission, but its stimulatory action was abolished by activated inhibitory 5-HT1A receptor. Chronic administration of mirtazapine resulted in increased extracellular serotonin level via reduction of serotonin transporter with reduction of somatodendritic 5-HT1A autoreceptor function in raphe nuclei. These pharmacological actions of mirtazapine include its serotonergic profiles as noradrenergic and specific serotonergic antidepressant (NaSSA).
Abstract: Nocturnal frontal lobe epilepsy is seen exclusively during sleep and is characterized by three distinct seizure phenotypes: paroxysmal arousals, paroxysmal dystonia, and episodic wandering. Mutations of CHRNA4, CHRNB2, or CHRNA2 genes encoding alpha4, beta2 or alpha2 subunits of neuronal nicotinic ACh receptor (nAChR) have been identified in the individuals with sporadic type NFLE and pedigrees with autosomal dominant type of NFLE (ADNFLE). In the past decade, various electrophysiological studies have analyzed the functional abnormalities of ADNFLE/NFLE mutant nAChR; however, the detailed pathogenesis of ADNFLE/NFLE has remained to be clarified. Therefore, to explore the pathogenesis of ADNFLE/NFLE, genetic animal models harboring ADNFLE mutant Chrna4 genes have recently been established. The face, construct and predictive validities have been demonstrated in a transgenic rat strain bearing the S284L mutant Chrna4 gene. The in vivo analyses of the functional abnormalities using genetic ADNFLE/NFLE animal models suggest the putative mechanisms of the ADNFLE/NFLE seizure onset during slow wave sleep.
Abstract: A few mutations in the gene encoding the gamma 2 subunit of the gamma-aminobutyric acid receptor type A (GABRG2) have been reported in various types of epilepsy. The aim of this study is to investigate the role of GABRG2 in the pathogenesis of childhood epilepsy in a large Japanese cohort. Genetic analysis of GABRG2 was performed on 140 Japanese patients with various childhood epilepsies largely including Dravet syndrome and genetic epilepsy with febrile seizures plus. The mutational analysis identified one novel missense mutation of GABRG2 (c.236A>G: p.N40S) in a patient with generalized tonic-clonic seizures (GTCS). The mutation was heterozygous and replacing a highly conserved Asn residue with a Ser. The affected amino acid was located at residue 40 of the mature GABRG2 protein, which was near the first one of two high-affinity benzodiazepine-binding domains of the gamma2 subunit (Lys-41-Trp-82). This mutation in such an important position may hamper the function of the channel and contribute to the case's pathogenesis of GTCS.
Abstract: To clarify the antiepileptic mechanisms of valproate (VPA), we determined the effects of acute and sub-acute administrations of VPA on ryanodine receptor (RyR)-associated hippocampal releases of GABA and glutamate using microdialysis, as well expression of mRNA and protein of RyR subtypes in the rat hippocampus. Acute administration of therapeutic-relevant VPA did not affect the hippocampal extracellular levels of GABA or glutamate, whereas sub-acute administration increased GABA level without affecting that of glutamate. Perfusion with ryanodine increased the hippocampal extracellular level of glutamate (ryanodine concentration range: 1-1000μM) concentration-dependently; however, that of GABA was increased by 1-100μM ryanodine concentration-dependently but the stimulatory effects of 1000μM ryanodine on GABA release was not observed. Both acute and sub-acute administrations of therapeutic-relevant VPA inhibited ryanodine-induced responses of hippocampal extracellular glutamate level without affecting that of GABA. Especially, both acute and sub-acute administrations of VPA prevented the breakdown of GABA release induced by 1000μM ryanodine. Sub-acute administration of therapeutically-relevant dose VPA weakly increased RyR mRNA expression but we could not detect the changes of RyR protein expression in rat hippocampus. These results suggest that VPA inhibited the neurotransmitter release associated with RyR without affecting the expression of RyR protein. Therefore, the antiepileptic action of VPA seems to be mediated, at least in part, by an increase in basal GABA release and inhibition of RyR-associated glutamate release.
Abstract: Japanese is a free word-order language, and allows both subject-object-verb (SOV) and object-subject-verb (OSV) orders. Our previous study using near-infrared spectroscopy (NIRS) imaging revealed that OSV sentences induce more activation in the left frontal lobe than SOV sentences. The present study develops our previous experiment: (1) by adopting an event-related design, and (2) by using sentences involving the adverb naze 'why', which plays a prominent role in recent linguistic studies. The results of our new experiment indicated that the cerebral activation in O why SV sentences was significantly larger than that in S why OV sentences, in the right anterior prefrontal region, which is consistent with the assumption that O why SV order is derived from S why OV order. We speculate that the activation observed in the anterior prefrontal cortex during the processing of the sentences involving 'why' might be due to the processing of higher-order function in the cerebral cortex.
Abstract: To clarify the mechanisms of action of blonanserin, an atypical antipsychotic drug, we studied the effects of systemic administration of blonanserin and risperidone on extracellular levels of norepinephrine, dopamine, serotonin, GABA and glutamate in the medial prefrontal cortex using microdialysis, and neuronal firing in the ventral tegmental area, locus coeruleus, dorsal raphe nucleus and mediodorsal thalamic nucleus using radiotelemetry. The binding affinities of blonanserin to D(2) and 5-HT(2A) receptors in the rat brain were confirmed and found to be similar. Blonanserin transiently increased neuronal firing in locus coeruleus and ventral tegmental area but not in dorsal raphe nucleus or mediodorsal thalamic nucleus, whereas risperidone increased the firing in locus coeruleus, ventral tegmental area and dorsal raphe nucleus but not in mediodorsal thalamic nucleus. Blonanserin persistently increased frontal extracellular levels of norepinephrine and dopamine but not serotonin, GABA or glutamate, whereas risperidone persistently increased those of norepinephrine, dopamine and serotonin but not GABA or glutamate. These results suggest a pharmacological correlation between the stimulatory effects of these antipsychotics on frontal monoamine release and neuronal activity in monoaminergic nuclei. Inhibition of the α(2) adrenoceptor increased extracellular monoamine levels and enhanced blonanserin-induced increase in extracellular serotonin level. These results indicated that the combination of antagonism of D(2) and 5-HT(2A) receptors contribute to the rise in extracellular levels of norepinephrine and dopamine, and that α(2) adrenoceptors play important roles in frontal serotonin release. They also suggest that blonanserin-induced activation of monoaminergic transmission could be, at least partially, involved in atypical antipsychotic properties of blonanserin.
Abstract: The CACNA1A gene encodes the poreforming, voltage-sensitive subunit of the voltage-dependent Ca(v)2.1 calcium channel. Mutations in this gene have been linked to several human disorders, including familial hemiplegic migraine type 1, episodic ataxia type 2, and spinocerebellar ataxia type 6. In mice, mutations of the homolog Cacna1a cause recessively inherited phenotypes in tottering, rolling Nagoya, rocker, and leaner mice. Here we describe two knockdown mice with 28.4+/-3.4% and 13.8+/-3.3% of the wild-type Ca(v)2.1 quantity. 28.4+/-3.4% level mutants displayed ataxia, absence-like seizures and progressive cerebellar atrophy, although they had a normal life span. Mutants with 13.8+/-3.3% level exhibited ataxia severer than the 28.4+/-3.4% level mutants, absence-like seizures and additionally paroxysmal dyskinesia, and died premature around 3 weeks of age. These results indicate that knock down of Ca(v)2.1 quantity to 13.8+/-3.3% of the wild-type level are sufficient to induce the all neurological disorders observed in natural occurring Cacna1a mutants. These knockdown animals with Ca(v)2.1 calcium channels intact can contribute to functional studies of the molecule in the disease.
Abstract: To clarify the antiepileptic mechanisms of zonisamide (ZNS), we determined the interaction between ZNS and inositol-1,4,5-triphosphate receptor (IP3R) on exocytosis of GABA and glutamate in rat frontal cortex using microdialysis. ZNS increased basal GABA release, but not glutamate, concentration-dependently, and reduced concentration-dependently K(+)-evoked GABA and glutamate releases. Inhibition and activation of IP3R reduced and enhanced basal and K(+)-evoked GABA releases, respectively. The K(+)-evoked glutamate release was reduced and enhanced by IP3R antagonist and agonist, respectively, whereas basal glutamate release was increased by IP3R agonist but not affected by IP3R antagonist. Under extracellular Ca(2+) depletion, IP3R agonist increased basal GABA and glutamate releases. The latter effects of IP3R agonist were weakly enhanced by ZNS, but such stimulatory action of ZNS was abolished by extracellular Ca(2+) depletion. In contrast, ZNS inhibited the stimulatory effect of IP3R agonist on K(+)-evoked release. The stimulatory effect of IP3R agonist on basal release was regulated by N-type voltage-sensitive Ca(2+) channel (VSCC) rather than P- and L-type VSCCs, whereas the stimulatory effect of IP3R agonist on K(+)-evoked release was regulated by P- and L-type VSCCs rather than N-type VSCC. These results suggest that ZNS-activated N-type VSCC enhances IP3R-associated neurotransmitter release during resting stage, whereas ZNS-induced suppression of P- and L-type VSCCs possibly attenuates IP3R-associated neurotransmitter release during neuronal hyperexcitability. Therefore, the combination of both of these two actions of ZNS on IP3R-associated neurotransmitter release mechanism seems to be involved, at least in part, in the mechanisms of antiepileptic and neuroprotective actions of ZNS.
Abstract: We examined the relationship between the catechol-O-methyltransferase (COMT) Val158Met genotype and frontal lobe function by using multi-channel near-infrared spectroscopy (NIRS). The present study investigated oxygenated ([oxy-Hb]) and deoxygenated ([deoxy-Hb]) hemoglobin concentration changes during the performance of a verbal fluency task in the frontal region of 71 patients with panic disorder (PD). The activation of [oxy-Hb] on the right lateral prefrontal cortex was observed in the Met/Met genotype of the COMT gene polymorphism of PD patient groups in the analysis of NIRS, which seems to be related to the autonomic dysfunction in the pathogenesis of PD.
Abstract: A recent randomized control study demonstrated that zonisamide (ZNS), an antiepileptic drug, is effective in Parkinson's disease at the lower than the therapeutic doses against epilepsy (25-50 mg/day); however, the detailed mechanism of antiparkinsonian effects of ZNS remains to be clarified. To determine the mechanism of antiparkinsonian effect of ZNS, we investigated the effects of ZNS on extracellular levels of dopamine in the striatum (STR), glutamate in substantia nigra pars reticulata (SNr), GABA in globus pallidus (GP), subthalamic nucleus (STN) and SNr, using multiple microdialysis probes. Striatal perfusion of 1000 microM ZNS (within therapeutic-relevant concentration against epilepsy) increased extracellular levels of dopamine in STR, whereas 100 microM ZNS (lower than the therapeutic-relevant concentration against epilepsy but within the therapeutic rage against Parkinson's disease) did not affect it. Striatal perfusion of ZNS (100 and 1000 microM) decreased the extracellular levels of GABA in STN and glutamate in SNr, but decreased extracellular GABA level in GP without affecting GABA level in SNr. These concentration-dependent effects of ZNS on extracellular neurotransmitter levels were independent of dopamine and delta(2) receptors; however, blockade of delta(1) receptor inhibited the effects of ZNS. Furthermore, activation of delta(1) receptor enhanced the effects of ZNS on neurotransmitter level. These results suggest that ZNS does not affect the direct pathway but inhibits the indirect pathway, which is mediated by delta(1) receptor. Therefore, the antiparkinsonian effects of ZNS seem to be mediated through the interaction between lower than therapeutically-relevant concentration against epilepsy of ZNS (100 microM) and delta(1) receptor.
Abstract: To determine whether intrasubject reproducibility could be observed in the frontal cortex and to assess the mental-health status of subjects in each session.
Abstract: The atypical antipsychotic drug, quetiapine (QTP), is effective in schizophrenia and mood disorders, but induces seizures compared to typical antipsychotics.
Abstract: The mechanisms of paradoxical aggravation of epileptic seizures induced by selected antiepileptic drugs (AEDs) remain unclear. The present study addressed this issue by determining the seizure-threshold doses of carbamazepine (CBZ) and phenytoin (PHT), as well the dose-dependent effects of CBZ, PHT, and carbonic anhydrase-inhibiting AEDs, acetazolamide (AZM), topiramate (TPM), and zonisamide (ZNS), on neurotransmitter release in rat hippocampus. The dose-dependent effects of AEDs on hippocampal extracellular levels of glutamate (Glu), GABA, norepinephrine (NE), dopamine (DA), and serotonin (5-HT) were determined by microdialysis with high-speed and high-sensitive extreme liquid chromatography. Proconvulsive effects of AEDs were determined by telemetric-electrocorticography. Therapeutically relevant doses of AZM, CBZ, TPM, and ZNS increased hippocampal extracellular levels of GABA, NE, DA, and 5-HT, while PHT had no effect. Supratherapeutic doses of AZM, CBZ, PHT, TPM, and ZNS decreased extracellular levels of GABA, NE, DA, and 5-HT, without affecting Glu levels. Toxic doses of CBZ and PHT produced seizures (paradoxical intoxication), markedly increasing all transmitter levels, but TPM and ZNS even at toxic doses did not produce seizure. Co-administration experiments showed that therapeutically relevant doses of CBZ or PHT reduced the seizure-threshold doses of PHT or CBZ, respectively. In contrast, therapeutically relevant doses of AZM, TPM, and ZNS elevated the seizure-threshold doses of CBZ and PHT. These results suggested that blockade of high percentage of the population of voltage-dependent sodium channels by CBZ and PHT might be important in inducing paradoxical intoxication/reaction, and that inhibition of carbonic anhydrase inhibits this effect. TPM and ZNS are candidate first-choice agents in treatment of epilepsy when first-line AEDs are ineffective.
Abstract: BFNC is an autosomal dominant epileptic disorder caused by mutations of KCNQ2 or KCNQ3 potassium channel gene. W309R missense mutation in KCNQ3 gene was previously reported in a family with BFNC. In this study, potassium currents were recorded from HEK293 cells expressing both W309R mutant KCNQ3 and wild type KCNQ2 channels. We found a lack of potassium current in W309R mutant KCNQ3 and KCNQ2 channels, which can explain the hyper-excitability of CNS in patients with BFNC.
Abstract: Nilotinib is a second-generation BCR-ABL kinase inhibitor with improved potency and selectivity compared to imatinib. A Phase I/II dose-escalation study was designed to evaluate the efficacy, safety, and pharmacokinetics of nilotinib in Japanese patients with imatinib-resistant or -intolerant Philadelphia chromosome-positive (Ph+) chronic myelogenous leukemia (CML) or relapsed/refractory Ph+ acute lymphoblastic leukemia (ALL). A total of 34 patients were evaluated in this analysis and had a median duration of drug exposure of 293 (range 13-615) days. All 6 CML-CP patients without complete hematologic response (CHR) at baseline rapidly achieved CHR. A major cytogenetic response was achieved in 94% of patients with CML-CP, including a complete cytogenetic response in 69%. A major molecular response was achieved by 56%. These responses were also observed in patients with CML in advanced stages and Ph+ ALL. Non-hematologic adverse events were mostly mild to moderate. Grade 3 or 4 neutropenia and thrombocytopenia occurred in 50 and 28% of patients, respectively. Overall, the results of this study suggest that nilotinib induced significant responses in imatinib-resistant or -intolerant patients with CML-CP and CML in advanced stages and Ph+ ALL. The results of this study confirmed the efficacy and safety of nilotinib in Japanese patients.
Abstract: The underlying genetic abnormalities of rare familial idiopathic epilepsy have been identified, such as mutation in KCNQ2, a K(+) channel gene. Yet, few genetic abnormalities have been reported for commoner epilepsy, i.e., sporadic idiopathic epilepsy, which share a phenotype similar to those of familial epilepsy.
Abstract: The atypical antipsychotic drug, zotepine, is effective in treatment of schizophrenia and acute mania, but the incidence of seizures during treatment is higher than with other antipsychotics. In addition, the mechanisms underlying the clinical actions of zotepine remain uncharacterized.
Abstract: To investigate whether prefrontal function during a cognitive task reflects the severity of panic disorder, the prefrontal function during a word fluency task in 109 panic disorder patients with or without agoraphobia was measured by multi-channel near-infrared spectroscopy (NIRS). [Oxy-Hb] changes in the left inferior prefrontal cortex were significantly associated with the frequency of panic attacks, and, in addition, [deoxy-Hb] changes in the anterior area of the right prefrontal cortex were significantly associated with the severity of agoraphobia. These results suggest that the prefrontal function in patients with panic disorder is associated with the disease state of disease in patients with panic disorder.
Abstract: To determine the basic word order in Japanese, oxyhemoglobin concentration changes in the frontal region of 32 healthy men were measured by near-infrared spectroscopy. Our experiment used four types of sentences: (i) subject-object-verb (SOV), (ii) object-subject-verb (OSV), (iii) subject-naze (why)-object-verb (SwhyOV), and (iv) naze (why)-subject-object-verb (whySOV). The results have shown that although oxyhemoglobin changes in the OSV sentences were significantly larger than those in the SOV sentences in the left frontal lobe, such effects were not observed between SwhyOV sentences and whySOV sentences. These results are consistent with the view that both SwhyOV and whySOV are the basic, which suggests that neurolinguistic evidence has the potential to provide an important basis for determining the basic word order in a free word-order language.
Abstract: Psychotic-like experiences (PLEs) are considered predictive of mental health problems later in life. However, little has been known about the mental health status and psychopathological distress in adolescents with PLEs in the general population. To investigate the associations between PLEs and mental health status or psychopathologies in a community sample of adolescents in a school-based cross-sectional fashion, PLEs were studied using a self-rating questionnaire in 5073 Japanese junior-high school students aged 12-15 years. Mental health status was evaluated using the 12-item General Health Questionnaire (GHQ-12). Psychopathologies, lifestyle, victimization, and interpersonal and help-seeking attitudes were also studied using a self-rating questionnaire. Fifteen percent of the students reported definitely having experienced at least one PLE. A dose-response relationship between the severity of PLEs and the prevalence of poor mental health status was observed. PLEs were also significantly associated with psychopathologies (strong anxiety in the classroom: OR = 1.4, 95% CI 1.2-1.6; suicidal ideation: OR = 2.1, 95% CI 1.8-2.4; self-harm behaviors: OR = 1.4, 95% CI 1.0-1.9; difficulty falling asleep due to hypersensitivity to environmental noise: OR = 1.7, 95% CI 1.4-2.0; difficulty concentrating due to hypersensitivity to environmental noise: OR = 1.5, 95% CI 1.3-1.8; physically assaulting others: OR = 1.3, 95% CI 1.0-1.5; bullying others, OR = 1.3, 95% CI 1.1-1.5; irritability when exchanging e-mails: OR = 1.3, 95% CI 1.0-1.6). Adolescents with PLEs in the community suffer from a wide range of psychopathological problems during crucial developmental periods [corrected]
Abstract: Temporal low-voltage irregular delta-waves (TLID) are often found in elderly subjects. The physiological significance of TLID has not been clarified; however, our previous studies suggest that TLID are associated with mild cerebrovascular dysfunction.
Abstract: Mutations of genes encoding alpha4, beta2, or alpha2 subunits (CHRNA4, CHRNB2, or CHRNA2, respectively) of nAChR [neuronal nicotinic ACh (acetylcholine) receptor] cause nocturnal frontal lobe epilepsy (NFLE) in human. NFLE-related seizures are seen exclusively during sleep and are characterized by three distinct seizure phenotypes: "paroxysmal arousals," "paroxysmal dystonia," and "episodic wandering." We generated transgenic rat strains that harbor a missense mutation S284L, which had been identified in CHRNA4 in NFLE. The transgenic rats were free of biological abnormalities, such as dysmorphology in the CNS, and behavioral abnormalities. The mRNA level of the transgene (mutant Chrna4) was similar to the wild type, and no distorted expression was detected in the brain. However, the transgenic rats showed epileptic seizure phenotypes during slow-wave sleep (SWS) similar to those in NFLE exhibiting three characteristic seizure phenotypes and thus fulfilled the diagnostic criteria of human NFLE. The therapeutic response of these rats to conventional antiepileptic drugs also resembled that of NFLE patients with the S284L mutation. The rats exhibited two major abnormalities in neurotransmission: (1) attenuation of synaptic and extrasynaptic GABAergic transmission and (2) abnormal glutamate release during SWS. The currently available genetically engineered animal models of epilepsy are limited to mice; thus, our transgenic rats offer another dimension to the epilepsy research field.
Abstract: The present study investigated oxygenated ([oxy-Hb]) and deoxygenated ([deoxy-Hb]) hemoglobin concentration changes during the performance of a word fluency task in the frontal region of five drug-naive patients with panic disorder with or without agoraphobia and in 33 age-, sex-, and task performance-matched healthy volunteers by using multi-channel near-infrared spectroscopy (NIRS). The left inferior frontal [oxy-Hb] changes during performance of the task in patients with panic disorder were significantly smaller than those of healthy controls. This pilot study suggests the possibility that the left frontal lobe, required for cognitive function, is impaired in patients with panic disorder.
Abstract: Staphylococcal enterotoxins (SEs) produced by Staphylococcus aureus are the most recognizable bacterial superantigenic toxins causing food poisoning in humans throughout the world. However, it remains unclear how SEs induce emesis and its emetic signal pathway. We investigated a mechanism of SEA-induced emesis using a small emetic animal model, house musk shrew. SEA-induced emesis in the animals was inhibited by a 5-hydroxytryptamine (5-HT) synthesis inhibitor and a 5-HT(3) receptor antagonist. SEA could increase 5-HT release in the small intestine. Pre-treatment with 5,7-dihydroxytryptamine (5,7-DHT) markedly inhibited SEA-induced emesis. SEA-induced emesis was also abolished by surgical vagotomy. Furthermore, cannabinoid (CB) receptor agonists inhibited SEA-induced emesis, and the action was reversed by a CB1 antagonist. Both 5-HT release and CB1 receptor expression were found in the mucosal and myenteric plexus of the intestine. Moreover, a CB1 receptor agonist significantly decreased the 5-HT release in the intestine. These results demonstrate that SEA induces 5-HT release in intestine, rather than in brain, and that the 5-HT(3) receptors on vagal afferent neurons are essential for SEA-stimulated emesis. In addition, SEA-induced emesis is downregulated by the CB system through decreasing 5-HT release in intestine.
Abstract: To clarify the mechanisms of the pharmacological action of carbamazepine (CBZ), we determined the effect of CBZ on GABA and glutamate release associated with the ryanodine receptor (Ryr)-sensitive Ca(2+)-induced Ca(2+)-releasing system (CICR) in the rat hippocampus using microdialysis. The therapeutically relevant concentration of CBZ increased basal GABA release without affecting basal glutamate release; however, K(+)-evoked releases were concentration-dependently reduced by CBZ. Lower-concentration ryanodine increased basal and K(+)-evoked releases of GABA and glutamate in a concentration dependent manner, whereas higher-concentration ryanodine reduced them. These inflection points in the concentration-response curves of ryanodine for neurotransmitter release (critical concentrations) were shifted to the left by K(+)-evoked stimulation. The critical concentration of ryanodine in GABA release was lower than that in glutamate release. During the resting stage, the critical concentrations of ryanodine were unaffected by inhibition of L-type, N-type and P-type voltage-sensitive Ca(2+) channels (VSCCs) but were prevented by CBZ; however, during the neuronal hyperexcitable stage, the critical concentration was increased by CBZ, L-type and P-type VSCC inhibitors but not the N-type VSCC inhibitor. Therefore, a therapeutically relevant concentration of CBZ protects against the breakdown of the neurotransmitter release mechanism induced by hyperactivation of Ryr via inhibition of L-type and P-type VSCCs as well as inhibition of Ryr-sensitive CICR. These actions of CBZ appear to be involved, at least partially, in its anti-seizure mechanisms.
Abstract: Evidence that some types of epilepsies show strong genetic predisposition has been well documented. AP3M2 is considered to be an epileptogenic gene because AP3M2 knockout mice exhibit symptoms of spontaneous epileptic seizures. In order to investigate whether the AP3M2 gene causes susceptibility to epilepsy, we performed mutation screening of the genomic DNA of 190 patients with six epilepsy types; this screening involved all the 9 exons and the relevant exon-intron boundaries of AP3M2. Although neither missense nor nonsense mutations were detected, we identified 21 sequence variations, of which 16 variations were novel. Of the 21 variations, 11 were detected in 5' and 3' UTRs, while the remaining variations were detected in introns. Although the present study failed to identify the possible AP3M2 mutations that may cause epilepsy, our results suggest that some AP3M2 mutations still remain candidates for unmapped disorders including epilepsy, febrile seizure, and other neuronal developmental disorders associated with functional abnormalities of GABAergic transmission.
Abstract: Interleukin-1beta (IL-1beta) plays an important role in neuroprotective and neurodegenerative events in the central nervous system. To clarify the mechanism of controversial actions of IL-1beta, we determined the effect of IL-1beta, as well as the interaction between IL-1beta and Ca(2+)-induced Ca2+ releasing system (CICR), on adenosine releases in mice hippocampus using mini-slices method. Basal and K(+)-stimulated adenosine releases were regulated by two types of CICRs, including inositol-1,4,5-trisphosphate (IP3) receptor and ryanodine receptor. Lower concentration of IL-1beta increased both adenosine releases, whereas higher concentration did not affect their releases. The stimulatory effect of IL-1beta on basal adenosine release was reduced by removal of extracellular Ca2+ and IP3 receptor inhibitor, while the stimulatory effect of IL-1beta on K(+)-stimulated adenosine release was reduced by ryanodine receptor inhibitor. These results suggest that the potent effect of IL-1beta upon adenosine release might contribute to the neuroprotective action of IL-1beta, whereas IL-1beta-induced neurodegeneration might be due to the overload response of Ca2+ mobilization and the inactivation of adenosine exocytosis.
Abstract: Recent advances in DNA microarray technology have enabled the identification of small alterations throughout the genome. We used standard karyotype analysis, followed by DNA microarray analysis and PCR to precisely map the chromosomal 4p deletion and determine the deletion breakpoints in the genome of an epileptic patient. The karyotype of the patient was 46,XY,del(4)(p15.2p15.3) as determined by G-banding analysis. We used a high-density oligonucleotide genotyping array to estimate the size of the deletion (4.5 Mb) and to locate the breakpoints within a 9-kb region on one side of the deletion and a 100-kb region on the other side. We amplified by PCR and sequenced the genomic region encompassing the breakpoints, and mapped the deletion to regions extending from 21648457 to 26164287 and from 26164505 to 26167493, respectively (chromosome 4 of NCBI Homo sapiens Genome Build 35.1). The deletion involves 18 genes, one of which (CCKAR) is partially deleted.
Abstract: Recent clinical and basic studies have demonstrated that hyperactivation of interleukin-1beta (IL-1beta) plays important roles in generation of febrile and epileptic seizures. To clarify this mechanism, the present study determined the effects of IL-1beta on Ca2+-associated releases of glutamate and GABA in mouse hippocampus. Both basal and K+-evoked GABA releases were regulated by Ca2+ influx and Ca2+-induced Ca2+ releasing system (CICR). The K+-evoked glutamate release was also regulated by Ca2+ influx and CICR, whereas basal glutamate release was not affected by them. IL-1beta increased basal releases of glutamate and GABA depending on the activation of Ca2+ influx and ryanodine receptor (RyR)-sensitive CICR, but reduced K+-evoked releases depending on Ca2+ influx, RyR-sensitive and inositol 1,4,5-trisphosphate receptor (IP3R)-sensitive CICRs. During neuronal hyperexcitability, the effect of IL-1beta on GABA release was more predominantly modulated by Ca2+ influx and RyR-sensitive CICR than that on glutamate. These results indicate that hyperactivation of IL-1beta leads to imbalance between glutamatergic and GABAergic transmission via toxic overload response of Ca2+ influx and CICR.
Abstract: We investigated the effect of Zonisamide (ZNS), a newer anti-epileptic drug, on field potentials and neuropropagation in rat frontal cortex, with the aid of the 64-channel multi-electrode dish (MED64) system. The amplitude and propagation of field potentials were expressed dimensionally in the MED64 system. ZNS (3-100 microM) inhibited the amplitude and propagation of field excitatory postsynaptic potentials (fEPSP) in a concentration dependent manner. In contrast, ZNS could not suppress the amplitude and propagation of the presynaptic fiber volley (PrV) at clinically relevant concentrations (10-30 microM). Stimulating dependency with reduction fEPSP was seen in the presence of ZNS at clinically relevant concentrations, but not with PrV. The reduction of fEPSP amplitude was not accompanied by a change in paired-pulse facilitation. These data suggest that at clinically relevant concentrations of ZNS, the suppression of neuronal propagation is at least partially due to the postsynaptic mechanism, probably through alpha-amino-3-hydroxy-5-methyl-4-isoxazolepropionic acid (AMPA) receptors.
Abstract: This study describes comprehensive polling of transcription start and termination sites and analysis of previously unidentified full-length complementary DNAs derived from the mouse genome. We identify the 5' and 3' boundaries of 181,047 transcripts with extensive variation in transcripts arising from alternative promoter usage, splicing, and polyadenylation. There are 16,247 new mouse protein-coding transcripts, including 5154 encoding previously unidentified proteins. Genomic mapping of the transcriptome reveals transcriptional forests, with overlapping transcription on both strands, separated by deserts in which few transcripts are observed. The data provide a comprehensive platform for the comparative analysis of mammalian transcriptional regulation in differentiation and development.
Abstract: To clarify the antiepileptic and neuroprotective actions of zonisamide (ZNS), we determined acute effects of ZNS on exocytosis of GABA and glutamate associated with ryanodine-receptor (Ryr) in rat hippocampus using microdialysis. ZNS increased basal GABA release concentration-dependently without affecting basal glutamate release; however, K(+)-evoked glutamate and GABA releases were reduced by ZNS concentration-dependently. Inhibition of Ryr reduced K(+)-evoked GABA and glutamate releases without affecting their basal releases. Ryanodine affected GABA and glutamate releases biphasic concentration-dependently: lower concentration of ryanodine increased both basal and K(+)-evoked releases of GABA and glutamate, whereas higher concentration reduced them. The therapeutically relevant concentration of ZNS inhibited ryanodine-induced GABA and glutamate releases, and abolished the inflection point in concentration-response curve for ryanodine on neurotransmitter exocytosis. These data suggest that ZNS elevates seizure threshold via enhancement of GABAergic transmission during resting stage. ZNS inhibits propagation of epileptic hyperexcitability and Ryr-associated neuronal damage during neuronal hyperexcitable stage. These demonstrations indicate that the indirect inhibition of Ryr activities by ZNS during neuronal hyperexcitability appear to be involved in the mechanisms of action of antiepileptic and neuroprotective actions of ZNS.
Abstract: Dynamic changes in intracellular free Ca(2+) concentration play a crucial role in various neural functions. The inositol 1,4,5-trisphosphate (IP3) receptor (IP3R) and the ryanodine (Ry) receptor (RyR) are involved in Ca(2+)-induced Ca(2+)-release (CICR). Recent studies have shown that type 3 IP3R is highly expressed in rat hippocampal neurons after kainic acid (KA)-induced seizures and that dantrolene, a RyR antagonist, reduces KA-induced neuronal cell death. We investigated the RyR-associated effects of CICR agents on basal and K(+)-evoked releases of GABA and glutamate in rat hippocampus and the changes in expression of mRNA for RyRs in mouse brain after KA-induced seizures. The stimulatory effect of Ry on releases of GABA and glutamate was concentration-dependent in a biphasic manner. The inflection point in concentration-response curves for Ry on GABA release was lower than that for glutamate in both basal and K(+)-evoked conditions, suggesting that hyperactivation of RyR-associated CICR produces the imbalance between GABAergic and glutamatergic transmission. Following KA-induced seizures, transient up-regulation of brain-type RyR mRNA was observed in the hippocampal CA3 region and striatum, and signals for c-Fos mRNA increased transiently in the hippocampus, dentate gyrus and deeper layers of the neocortex. Thereafter, some dead neurons with single-stranded DNA (ssDNA) immunoreactive fragmented nuclei appeared in these areas. These findings suggest that intracellular Ca(2+) release via the RyR might be one of the mechanisms involved in KA-induced neuronal cell death.
Abstract: To search for clues to molecular genetics of common idiopathic epilepsy syndromes. Genetic defects have been identified recently in certain inherited epilepsy syndromes in which the phenotypes are similar to those of common idiopathic epilepsies.
Abstract: To explore the pharmacological mechanisms of topiramate (TPM), we determined the effects of TPM on monoamine (dopamine and serotonin) exocytosis associated with N-ethylmaleimide-sensitive factor attachment protein receptors and Ca(2+)-induced Ca(2+)-releasing systems, including inositol-triphosphate receptor and ryanodine receptor in freely moving rat pre-frontal cortex using in vivo microdialysis. During resting stage, Ca(2+) output from endoplasmic reticulum Ca(2+) store via inositol-triphosphate receptor regulates syntaxin-associated monoamine exocytosis mechanism, whereas during neuronal hyperexcitable stage, Ca(2+) output via ryanodine receptor regulates synaptobrevin-associated monoamine exocytosis mechanism. Basal monoamine releases were increased and decreased by therapeutically relevant and supratherapeutic concentration of TPM, respectively. The therapeutic-relevant concentration of TPM increased Ca(2+)-evoked release concentration-dependently; however, its stimulatory effect was attenuated in the supratherapeutic range. The K(+)-evoked releases were reduced by TPM concentration-dependently (from therapeutic to supratherapeutic ranges). The therapeutic-relevant concentration of TPM-induced elevation of basal release was reduced by cleavage with syntaxin and inhibition of inositol-triphosphate receptor predominantly, by cleavage with SNAP-25 and synaptobrevin weakly, but not by ryanodine receptor inhibitor. The therapeutic-relevant concentration of TPM-induced elevation of Ca(2+)-evoked release was reduced by cleavage with syntaxin and inositol-triphosphate receptor inhibitor selectively. The therapeutic-relevant concentration of TPM-induced reduction of K(+)-evoked monoamine release was abolished by cleavage with synaptobrevin, but was not affected by cleavage with SNAP-25 or synaptobrevin. The stimulatory effect of ryanodine receptor agonist on K(+)-evoked monoamine release was reduced by TPM, whereas that of inositol-triphosphate receptor agonist was not affected by TPM. Therefore, these results indicate that the combination of the effects of TPM on exocytosis mechanisms associated with SNARE and Ca(2+)-induced Ca(2+)-releasing systems, enhancement of inositol-triphosphate receptor/syntaxin and inhibition of ryanodine receptor/synaptobrevin in pre-frontal cortex, may be involved in clinical actions of TPM.
Abstract: Quinupristin-dalfopristin (Q-D) is a mixture of quinupristin and dalfopristin, which are semisynthetic antibiotics of streptogramin groups B and A, respectively.
Abstract: Psychomotor development of 71 children born to mothers with epilepsy was prospectively studied and compared to those of 99 controls matched for age, maternal educational level and age, and socioeconomic status. Intrauterine growth retardation disappeared before age 3 years. Assessment at age 1.5 years revealed that exposure to seizures, high dose of antiepileptic drugs (AEDs) in utero, and small head circumference at birth affected development quotient (DQ) scores of motor or linguistic abilities or both. DQ scores of motor ability of children of mothers with complex partial seizures were lower than those with simple partial seizures when assessed at age 3 years. Assessments at age 1.5 years revealed that the total daily dose of AEDs correlated negatively with DQ scores of motor ability, and at age 3 years, maternal educational level affected DQ scores of some fields, including linguistic ability. The effects of AED exposure in utero and the occurrence of maternal seizures on the development of offspring were found to matter more at the younger age, but later on, the child care environment and, in particular, maternal ability of child-rearing, became more important. Our findings indicate that careful and regular follow-ups are needed to monitor the developmental stages of children of mothers with epilepsy, and the introduction of a day nursery should be suggested if necessary.
Abstract: We investigated the marital status of the patients with epilepsy to clarify the clinical factors impeding improvement of the quality of life in adults with epilepsy.
Abstract: Phencyclidine (PCP) produces schizophrenia-like psychosis and acute PCP-intoxications; however, whether glutamate/NMDA-receptor blockade by PCP modulates or not these mechanisms has remained to be clarified. To clarify this mechanism, we determined interaction among voltage-gated Na(+)-channel inhibitor, tetrodotoxin (TTX), Golgi-disturbing-agent, brefeldin-A (BFA), and PCP on releases of glutamate, GABA, and monoamine in prefrontal-cortex (pFC), using microdialysis. PCP increased basal monoamine release, whereas it decreased basal GABA release, without affecting glutamate release. PCP increased K(+)-evoked monoamine release, whereas it decreased K(+)-evoked glutamate and GABA releases. TTX reduced basal monoamine and GABA releases without affecting glutamate release, whereas BFA did not affect them. Interestingly, BFA and TTX inhibited PCP-associated basal monoamine release and abolished PCP-induced reduction of basal GABA release without affecting glutamate release. BFA and TTX reduced K(+)-evoked releases of all neurotransmitters. BFA inhibited PCP-associated K(+)-evoked monoamine release, but TTX did not affect them. PCP-induced reduction of K(+)-evoked GABA and glutamate releases was abolished by TTX and BFA. These results indicate that PCP reduces GABAergic transmission via NMDA-receptor blockade and activates intracellular endoplasmic-reticulum-associated signal-transduction, resulting in enhancement of monoaminergic transmission in pFC. Thus, these PCP properties support the hypothesis that mechanisms of the neurological symptoms of acute PCP-intoxication, convulsion, and rhabdomyolysis may be involved in both reduction of GABAergic-transmission and activation of endoplasmic-reticulum-associated signal-transduction induced by PCP.
Abstract: To explore the exocytosis mechanism of dopamine and its precursor, 3,4-dihydroxyphenylalanine (DOPA), we determined the effects of protein-kinase, cyclic-AMP-dependent protein-kinase (PKA), Ca(2+)-phospholipid-dependent protein-kinase (PKC) and Ca(2+)-calmodulin-dependent protein-kinase II (CaMK-II) on dopamine and DOPA releases in rat striatum using microdialysis. Basal DOPA and dopamine releases were reduced by PKC and CaMK-II inhibitors predominantly, and PKA inhibitor weakly. Ca(2+)-evoked releases were reduced by PKC and CaMK-II inhibitors, but not by PKA inhibitor. K(+)-evoked (20 min) releases were reduced by PKA and CaMK-II inhibitors predominantly, and PKC inhibitor weakly. Sustained K(+)-evoked (120 min) releases of DOPA and dopamine were reduced by CaMK-II inhibitor, but not by PKC or PKA. DOPA accumulation was reduced by PKA and CaMK-II inhibitors strongly, and PKC inhibitor weakly. Therefore, the present study demonstrates that striatal DOPA exocytosis is regulated by a similar protein kinase-associated exocytosis mechanism as that of dopamine.
Abstract: AP-3 is a member of the adaptor protein (AP) complex family that regulates the vesicular transport of cargo proteins in the secretory and endocytic pathways. There are two isoforms of AP-3: the ubiquitously expressed AP-3A and the neuron-specific AP-3B. Although the physiological role of AP-3A has recently been elucidated, that of AP-3B remains unsolved. To address this question, we generated mice lacking mu3B, a subunit of AP-3B. mu3B-/- mice suffered from spontaneous epileptic seizures. Morphological abnormalities were observed at synapses in these mice. Biochemical studies demonstrated the impairment of gamma-aminobutyric acid (GABA) release because of, at least in part, the reduction of vesicular GABA transporter in mu3B-/- mice. This facilitated the induction of long-term potentiation in the hippocampus and the abnormal propagation of neuronal excitability via the temporoammonic pathway. Thus, AP-3B plays a critical role in the normal formation and function of a subset of synaptic vesicles. This work adds a new aspect to the pathogenesis of epilepsy.
Abstract: To explore the exocytosis mechanism of striatal 3,4-dihydroxyphenylalanine (DOPA), this study determined the interaction between voltage-sensitive Ca2+-channel (VSCC) and SNARE on releases of DOPA and glutamate in rat striatum using microdialysis. Inhibitors of VSCCs and SNAREs did not affect basal glutamate release but decreased basal DOPA release, however, blocking effects of P-type-VSCC and synaptobrevin inhibitors were weaker than those of N-type-VSCC and syntaxin. The K+-evoked releases of DOPA and glutamate were reduced by inhibitors of P-type-VSCC and synaptobrevin predominantly and by inhibitors of N-type-VSCC and syntaxin weakly. However, interaction study between VSCC and SNARE on K+-evoked DOPA release indicates that DOPA release is regulated by different exocytosis mechanism from glutamate and monoamine during the depolarization stage (N-type-VSCC/P-type-VSCC/synaptobrevin and/or combination with N-type-VSCC/synaptobrevin and P-type-VSCC/synaptobrevin). Therefore we conclude that striatal DOPA release might be regulated by its specific exocytosis mechanism via different from dopaminergic presynaptic vesicle.
Abstract: The entorhinal cortex (EC) is a major source of afferent input to the hippocampus via the perforant and temporoammonic pathways; however, the detailed transmission mechanism in the temporoammonic pathway remains to be clarified. Thus, we determined interaction among GABA(A), AMPA/glutamate receptors and protein kinases (PKA and PKC) in the exocytosis of GABA and glutamate using multiprobe microdialysis, as well as propagation of neuronal excitability using optical recording in the EC-Hippocampal formation. Multiprobe microdialysis demonstrated that EC-evoked GABA release in ventral CA1 was predominantly regulated by the PKC-related rather than PKA-related exocytosis mechanism and was augmented by the activation of glutamatergic transmission. Contrary to GABA release, EC-evoked glutamate release was predominantly regulated by PKA-related rather than PKC-related mechanisms and was suppressed by activation of GABAergic transmission. Optical recording demonstrated that there are two sub-pathways in the temporoammonic pathway; direct projects from EC layers (II-IV) to dendrites on pyramidal cells and GABAergic interneurons in ventral hippocampal CA1. PKC activation enhanced trisynaptic transmission, whether the GABA(A) receptor was functional or blocked, whereas PKC activation enhanced and inhibited temporoammonic transmission when the GABA(A) receptor was functional and blocked, respectively. Thus, GABAergic inhibition, which is regulated by PKC activity, in the temporoammonic pathway is more significant than that in the trisynaptic pathway.
Abstract: To clarify the striatal Ca2+-dependent monoaminergic exocytosis mechanisms, this study determined the effects of the Ca2+-induced Ca2+ releasing system (CICR), containing inositol-trisphosphate-receptor (IP3R) and ryanodine-receptor (RyR), on striatal releases of dopamine and its precursor, 3,4-dihydroxyphenylalanine (DOPA), using microdialysis. The basal dopamine release is regulated by IP3R but not by RyR, whereas basal DOPA release does not require CICR. The K+-evoked releases of DOPA and dopamine were enhanced by IP3R agonist, whereas RyR agonist reduced it. Additionally, inhibition of dopamine release induced by RyR hyperactivation was prevented by inhibition of L-type voltage-sensitive Ca2+-channel activity. These present results suggest that CICR-associated regulation of striatal releases of DOPA and dopamine is restrictive during the resting stage, whereas CICRs play an important role as a reserve mechanism of exocytosis of striatal DOPA and dopamine during the hyperexcitable stage.
Abstract: Microdialysis has become an effective and frequently used technique to study the extracellular levels of monoamine, i.e. dopamine, serotonin and norepinephrine in the central nervous system. However, the detailed exocytosis mechanisms of monoamine using microdialysis has remained to be clarified. The present report introduces methods for administration of voltage-sensitive calcium channel (VSCC) inhibitors and soluble N-ethylmaleimide-sensitive factor attachment protein receptor (SNARE) inhibitors to clarify the mechanisms of monoamine exocytosis using in vivo microdialysis. The N-type and P-type VSCCs are inhibited by perfusion with omega-conotoxin GVIA and omega-agatoxin IVA, respectively; however, their diffusion rate from internal to external spaces of the microdialysis probe is lower than 1%. Unlike VSCC inhibitors, SNAP-25, synaptobrevin and syntaxin can be cleavaed with botulinum toxin type A, B and C, respectively. These toxins (with molecular weights over 500,000) bind specifically to the presynaptic membrane via the heavy chain, while the light chain enters the cytosol, where it displays zinc-endopeptidase activity directed to proteins of the neuroexocytosis apparatus. Therefore, to prevent SNARE activity, botulinum toxins are microinjected. These two methods, perfusion with VSCC inhibitors and microinjection with botulinum toxins, can contribute to the clarification of the mechanisms of monoaminergic exocytosis.
Abstract: Chromatin structure plays a critical role in the regulation of transcription. Drosophila GAGA factor directs chromatin remodeling to its binding sites. We show here that Drosophila FACT (facilitates chromatin transcription), a heterodimer of dSPT16 and dSSRP1, is associated with GAGA factor through its dSSRP1 subunit, binds to a nucleosome, and facilitates GAGA factor-directed chromatin remodeling. Moreover, genetic interactions between Trithorax-like encoding GAGA factor and spt16 implicate the GAGA factor-FACT complex in expression of Hox genes Ultrabithorax, Sex combs reduced, and Abdominal-B. Chromatin immunoprecipitation experiments indicated the presence of the GAGA factor-FACT complex in the regulatory regions of Ultrabithorax and Abdominal-B. These data illustrate a crucial role of FACT in the modulation of chromatin structure for the regulation of gene expression.
Abstract: Little is known concerning promoters or gene therapy specific for ovarian cancer. To explore the potential use of IAI.3B isolated from ovarian cancer cells in gene therapy for ovarian cancer, we identified the promoter region of the IAI.3B gene and created a replication-selective adenovirus, AdE3-IAI.3B, driven by the promoter. Transient transfection experiments showed that the DNA segment located between -1816 and -1 bp resulted in preferential expression in ovarian cancer cells with negligible expression in squamous cell carcinoma and normal cells. The promoter activity of IAI.3B was almost the same as that of cytomegalovirus and an order of magnitude higher than those of midkine and cyclooxygenase-2 in ovarian cancer cells. AdE3-IAI.3B replicated as efficiently as the wild-type adenovirus and caused extensive cell killing in a panel of ovarian cancer cells in vitro. In contrast, squamous cell carcinoma and normal cells were not able to support AdE3-IAI.3B replication. In animal studies, AdE3-IAI.3B administered to flank and i.p. xenografts of ovarian cancer cells led to a significant therapeutic effect. These results demonstrate the usefulness of the IAI.3B promoter for generation of ovarian cancer-specific adenoviral vectors and provide a potential for the development of ovarian cancer-specific oncolytic viral therapies.
Abstract: Febrile seizures (FS) may represent the most common seizure disorder in childhood and are known to be associated with putative genetic predispositions. Nevertheless, molecular genetic approaches toward understanding FS have been just initiated this decade. Recently, several genetic loci for FS have been mapped thereby assuring the genetic heterogeneity of FS. However, the exact molecular mechanisms of FS are yet to be elucidated. Genetic defects have been recently identified in autosomal dominant epilepsy with FS plus or generalized epilepsy with FS plus. The underlying mutations were found in genes encoding several Na+ channel subunits and the gamma2 subunit of gamma amino-butyric acid (GABA)A receptors in the brain. Furthermore, both channels are also associated with severe myoclonic epilepsy in infancy, where the seizure attacks often begin with prolonged FS and are precipitated by fever even afterwards. Na+ channels are associated with other temperature-sensitive disorders, and GABA(A) receptors are known to play an important role in the pathogenesis of FS. These lines of evidence suggest the involvement of various Na+ channels, GABA(A) receptors and additional auxiliary proteins in the pathogenesis of frequent FS and even in simple FS. This hypothesis may facilitate our understanding of the genetic background of FS.
Abstract: Recently, mutations of KCNQ2 or KCNQ3, members of the KCNQ-related K(+)-channel (KCNQ-channel) family, were identified as cause of benign familial neonatal convulsions (BFNC). However, the exact pathogenic mechanisms of age-dependent development and spontaneous remission of BFNC remain to be elucidated. To clarify the age-dependent etiology of BFNC, we determined age-dependent functional switching of KCNQ-channels, GABAergic- and glutamatergic-transmission in rat hippocampus. The effects of inhibitors of KCNQ-channel, GABA- and glutamate-receptors on propagation of neuronal-excitability and neurotransmitter release were determined by 64-channel multielectrode-dish (MED64), whole-cell recording, in vitro release technique and in vivo microdialysis biosensor, using rat hippocampus from day of birth (P0) to postnatal-day 56 (P56). Inhibition of KCNQ-channels enhanced depolarization-induced glutamate and GABA releases during P0-P7, but not during P14-P28. Inhibition of KCNQ-channels magnified neuronal-excitability propagation from P0 to P14: maximal at P3, but this effect disappeared by P28. GABA(A)-receptor inhibition surprisingly reduced neuronal-excitability propagation during P0-P3, but not at P7. AMPA/glutamate-receptors inhibition reduced propagation of neuronal-excitability throughout the study period. KCNQ-channels inhibition shortened spike-frequency adaptation, but this stimulation was more predominant during P<7 than P>14. During the first week of life, KCNQ-channels performed as a predominant inhibitory system, whereas after this period GABAergic-transmission switched from excitatory to inhibitory function. Contrary, glutamatergic-transmission has acquired as excitatory function from P0. These findings suggest that the pathogenic mechanisms of age-dependent development and spontaneous remission of BFNC are, at least partially, associated with the interaction between age-dependent reduction of inhibitory KCNQ-channel activity and age-dependent functional switching of the GABAergic-system from excitatory to inhibitory action in neonatal CNS.
Abstract: Genetic defects have been recently identified in certain inherited epilepsy syndromes in which the phenotypes are similar to common idiopathic epilepsies. Mutations in the neuronal nicotinic acetylcholine receptor 4 and 2 subunit genes have been detected in families with autosomal dominant nocturnal frontal lobe epilepsy. Both receptors are components of neuronal acetylcholine receptor, a ligand-gated ion channel in the brain. Furthermore, mutations of two K+-channel genes were also identified as the underlying genetic abnormalities of benign familial neonatal convulsions. Mutations in the voltage-gated Na+-channel 1, 2 and 1 and the gamma aminobutyric acid (GABAA) receptor 2 subunit genes were found as a cause of generalized epilepsy with febrile seizures plus, a clinical subset of febrile convulsions. Na+-channels, GABAA receptor and their auxiliaries may be involved in the pathogenesis of this subtype and even in simple febrile convulsions. Mutation of a voltage-gated K+-channel gene can cause partial seizures associated with periodic ataxia type 1 and some forms of juvenile myoclonic epilepsy and idiopathic generalized epilepsy can result from mutations of a Ca2+-channel. This line of evidence suggests the involvement of channels expressed in the brain in the pathogenesis of certain types of epilepsy. Our working hypothesis is to view certain idiopathic epilepsies as disorders of ion channels, i.e. 'channelopathies'. Such hypothesis should provide a new insight to our understanding of the genetic background of epilepsy.
Abstract: To clarify the mechanisms of action of antiepileptic drugs (AEDs), carbamazepine (CBZ) and zonisamide (ZNS), on exocytosis mechanisms, the present study determined the concentration-dependent action of CBZ and ZNS, as well as the interaction between these AEDs and voltage-sensitive Ca(2+) channel (VSCC) activity on basal, Ca(2+)- and K(+)-evoked acetylcholine (ACh) release in frontal cortex of freely moving rat using in vivo microdialysis. Perfusion with therapeutic-relevant concentrations of CBZ and ZNS increased basal ACh release, which was regulated by N-type VSCC predominantly and P-type VSCC weakly, whereas supratherapeutic-relevant concentrations of these AEDs reduced this release. The 3.4 mM Ca(2+)-evoked release, which was regulated by N-type VSCC selectively, but not by P-type VSCC, was increased by therapeutic-relevant concentrations of CBZ and ZNS, whereas this release was reduced by supratherapeutic-relevant concentrations of them. The 50 mM K(+)-evoked release, which was regulated by P-type VSCC predominantly and N-type VSCC weakly, was decreased by CBZ and ZNS, in a concentration-dependent manner. These findings indicate that the interplay between enhancement of basal ACh release and reduction of depolarization-related ACh release in the frontal cortex are at least partially involved in a common mechanism of antiepileptic action between CBZ and ZNS.
Abstract: Benign familial neonatal convulsions (BFNC), a hereditary epilepsy, occurs specifically in newborns and remits spontaneously after this period. Several mutations of either KCNQ2 or KCNQ3, members of the KCNQ-related K+-channel (KCNQ-channel) family, were identified as a cause of BFNC. Such mutations impair KCNQ-related M- current, an element of the inhibitory system in the central nervous system (CNS), and therefore are thought to result in neuronal hyperexcitability.
Abstract: Carbamazepine (CBZ) and zonisamide (ZNS) are effective antiepileptic drugs (AEDs) for the treatment of epilepsy and mood disorder. One of the mechanisms of action of CBZ and ZNS is inactivation of voltage-gated Na+ channel (VGSC). However, the major mechanism(s) of action of these AEDs is not clear yet. We have been exploring novel targeting mechanisms for the antiepileptic actions of CBZ and ZNS during the past ten years. In this report, we describe our hypothesis regarding the new targeting mechanisms for the antiepileptic action of AEDs. We determined an interaction between these AEDs and inhibitors of both voltage-sensitive Ca2+ channels (VSCCs) and soluble N-ethylmaleimide-sensitive factor attachment protein receptor (SNARE) on neurotransmitter exocytosis using microdialysis. Perfusion with therapeutic concentrations of CBZ and ZNS increased basal neurotransmitter release. This stimulatory action was predominantly inhibited by inhibitors of N-type VSCC and syntaxin. CBZ and ZNS increased Ca2+-evoked release, an action selectively inhibited by inhibitors of N-type VSCC and syntaxin. CBZ and ZNS reduced K+-evoked release, an action predominantly inhibited by inhibitors of P-type VSCCs and synaptobrevin. These actions of CBZ and ZNS on neurotransmitter exocytosis could be observed under the condition of inhibition of VGSC using perfusion with tetrodotoxin. Our findings enhance our understanding of the mechanisms of action of CBZ and ZNS as AEDs, which possibly reduce P-type VSCCs/synaptobrevin-related exocytosis mechanisms during the depolarization stage, and simultaneously enhance N-type VSCCs/syntaxin-related exocytosis mechanisms at the resting stage.
Abstract: Genetic defects have been recently identified in certain inherited epilepsy syndromes in which the phenotypes are similar to those of common idiopathic epilepsies. Mutations in the neuronal nicotinic acetylcholine receptor alpha4 and beta2 subunit genes have been detected in families with autosomal dominant nocturnal frontal lobe epilepsy. Both receptors are components of neuronal acetylcholine receptor, a ligand-gated ion channel in the brain. Furthermore, mutations of two K+ channel genes also were identified as the underlying genetic abnormalities of benign familial neonatal convulsions. Mutations in the voltage-gated Na+-channel alpha1 and beta1 subunit genes were found as the cause of generalized epilepsy with febrile seizures plus, a clinical subset of febrile convulsions. Mutation of a voltage-gated K+-channel gene can cause partial seizures associated with periodic ataxia type 1 and some forms of juvenile myoclonic epilepsy can result from mutations of a Ca2+ channel. This line of evidence suggests the involvement of channels expressed in the brain in the pathogenesis of certain types of epilepsy. Our working hypothesis is to view certain idiopathic epilepsies as disorders of ion channels (i.e., "channelopathies"). Such a hypothesis should provide a new insight into our understanding of the genetic background of epilepsy.
Abstract: Epilepsy affects >0.5% of the world's population and has a large genetic component. The most common human genetic epilepsies display a complex pattern of inheritance, and the identity of the susceptibility genes is largely unknown despite recent advances in molecular biology. However, genetic identifiers of certain types of epilepsy with neurodegenerative characteristics and of a small number of familial idiopathic epilepsies have been uncovered to date. This article reviews recent progress made in molecular genetics of epilepsy, focusing mostly on idiopathic epilepsy together with our own discovery of novel mutations in the genes of autosomal dominant nocturnal frontal lobe epilepsy and benign familial neonatal convulsions (BFNCs), and the genetic locus of benign adult familial myoclonic epilepsy. Pathogenesis of epilepsy as a channelopathy and of BFNC also is discussed.
Abstract: Dipole source localization corresponding to interictal spikes were estimated using EEG dipole tracing with a realistic three-shell head model in three patients with cryptogenic gelastic epilepsy. The dipole sources in two patients, whose gelastic seizures were accompanied by a subjective feeling of mirth, were estimated in the right or left medio-basal temporal regions. In the other patient, with gelastic seizures without a sense of mirth, the dipole sources were localized in the right frontal region corresponding to the anterior cingulate. The results suggest that the neural activities in hippocampal regions are involved with the generation of gelastic seizures with a sense of mirth and those in the cingulate might be associated with the motor act of laughter.
Abstract: Epilepsy affects more than 0.5% of the world's population and has a large genetic component. The most common human genetic epilepsies display a complex pattern of inheritance and the susceptibility genes are largely unknown. However, major advances have recently been made in our understanding of the genetic basis of monogenic inherited epilepsies. Progress has been particularly evident in familial idiopathic epilepsies and in many inherited symptomatic epilepsies, with the discovery that mutations in ion channel subunits are implicated, and direct molecular diagnosis of some phenotypes of epilepsy is now possible. This article reviews recent progress made in molecular genetics of epilepsy, focusing mostly on idiopathic epilepsy, and some types of myoclonus epilepsies. Mutations in the neuronal nicotinic acetylcholine receptor alpha4 and beta2 subunit genes have been detected in families with autosomal dominant nocturnal frontal lobe epilepsy, and those of two K(+) channel genes were identified to be responsible for underlying genetic abnormalities of benign familial neonatal convulsions. The voltage-gated Na(+) -channel (alpha1,2 and beta1 subunit), and GABA receptor (gamma2 subunit) may be involved in the pathogenesis of generalized epilepsy with febrile seizure plus and severe myoclonic epilepsy in infancy. Mutations of Ca(2+)-channel can cause some forms of juvenile myoclonic epilepsy and idiopathic generalized epilepsy. Based upon these findings, pathogenesis of epilepsy as a channelopathy and perspectives of molecular study of epilepsy are discussed.
Abstract: To clarify the mechanisms of interaction between adenosine receptor subtypes (A1R and A2R) on 5-HT release, the present study determined the effects of adenosine receptor subtypes on voltage-sensitive Ca(2+)-channels (VSCCs), protein-kinases (PKs) and synaptic-proteins (SNAREs) related 5-HT release using microdialysis in freely moving rat. A1R-antagonists increased basal 5-HT release, which was reduced by inhibitors of N-VSCC, PKC and syntaxin predominantly, and by inhibitors of PKA and synaptobrevin weakly, but was not affected by P-VSCC inhibitor. In the presence of A1R-antagonist, A2R-agonists increased basal 5-HT release, whose action was inhibited by P-VSCC, PKA and synaptobrevin inhibitors predominantly and reduced by N-VSCC, PKC and syntaxin inhibitors weakly. Under the condition of adenylate-cyclase activation in the absence of A1R-antagonists, A2R-agonists increased basal 5-HT release. K(+)-evoked 5-HT release was enhanced by A1R-antagonist and A2R-agonist, whose actions were inhibited by P-VSCC, PKA and synaptobrevin inhibitors predominantly. These results suggest that an activation of A1R suppresses 5-HT release via an inhibition of N-VSCC/PKC/syntaxin and P-VSCC/PKA/synaptobrevin, and an activation of A2-R stimulates 5-HT release via an enhancement of P-VSCC/PKA/synaptobrevin. Therefore PKA activity plays an important role in the interaction between A1R and A2R on hippocampal 5-HT release.
Abstract: To clarify the mechanisms of interaction between adenosine A(1) receptor (A1-R) and adenosine A(2) receptor (A2-R) on neurotransmitter release, this study determined the functional interactions among adenosine receptors (AD-Rs), voltage-sensitive Ca(2+) channels (VSCCs), protein kinases (PKs), and synaptic proteins [N-ethylmaleimide-sensitive factor (NSF) attachment protein (SNAP) receptors] on hippocampal serotonin release using in vivo microdialysis in freely moving rat. Basal serotonin release was regulated by two functional complexes: N-type VSCC (N-VSCC)/calcium-phospholipid-dependent protein kinase (PKC)/syntaxin (major pathway) and P-type VSCC (P-VSCC)/cyclic AMP-dependent protein kinase (PKA)/synaptobrevin (minor pathway). However, K(+)-evoked serotonin release was regulated by N-VSCC/PKC/syntaxin (minor pathway) and P-VSCC/PKA/synaptobrevin (major pathway). A1-R antagonists increased basal serotonin release, which was reduced by inhibitors of N-VSCC, PKC, and syntaxin predominantly and by inhibitors of PKA and synaptobrevin weakly, but was not affected by P-VSCC inhibitor. In the presence of A1-R antagonist, A2-R agonists increased basal serotonin release, which was inhibited by inhibitors of P-VSCC, PKA, and synaptobrevin predominantly and reduced by inhibitors of N-VSCC, PKC, and syntaxin weakly. Under the condition of activation of adenylate cyclase in the absence of A1-R antagonists, A2-R agonists increased basal serotonin release. A1-R antagonist and A2-R agonist enhanced K(+)-evoked serotonin release, which was inhibited by inhibitors of P-VSCC, PKA, and synaptobrevin predominantly. These results suggest that an activation of A1-R suppresses serotonin release via inhibition of both N-VSCC/PKC/syntaxin and P-VSCC/PKA/synaptobrevin pathways, and an activation of A2-R stimulates serotonin release via enhancement of the P-VSCC/PKA/synaptobrevin pathway. Therefore, PKA activity plays an important role in the interaction between A1-R and A2-R on hippocampal serotonin release.
Abstract: To elucidate mechanisms of hippocampal serotonin release and possible mechanisms of clinical action of carbamazepine (CBZ), we determined interaction between antagonists of N-type (omega-conotoxin GVIA:GVIA), P-type (omega-agatoxin IVA:IVA) Ca(2+) channels, Na(+) channel (tetrodotoxin: TTX) and CBZ on hippocampal basal, Ca(2+)- and K(+)-evoked serotonin releases, using microdialysis in freely moving rats. Basal release was reduced by TTX, GVIA and IVA (GVIA>IVA). Ca(2+)-evoked release was reduced by GVIA but unaffected by TTX and IVA. K(+)-evoked release was reduced by TTX, GVIA and IVA (GVIA<IVA). TTX inhibited actions of IVA and GVIA on respective basal and K(+)-evoked releases, without affecting Ca(2+)-evoked release. Perfusion with 100 microM CBZ (estimated-concentration in hippocampal tissue: 19+/-2 microM) enhanced basal and Ca(2+)-evoked releases, but reduced K(+)-evoked release, whereas 1000 microM CBZ (estimated-concentration in hippocampal tissue: 188+/-16 microM) reduced three types of releases. Under condition of pretreatment with 100 and 1000 microM CBZ, TTX unaffected basal and K(+)-evoked releases. Under condition of pretreatment with 100 microM CBZ, IVA and GVIA unaffected basal and K(+)-evoked releases, respectively, but GVIA reduced basal, Ca(2+)-evoked releases and IVA also reduced K(+)-evoked release. Under condition of pretreatment with 1000 microM CBZ, GVIA unaffected three types of releases, and IVA unaffected basal release but reduced K(+)-evoked release. These findings contribute towards the possible mechanisms of concentration-dependent antiepileptic action of CBZ, which possibly inhibits Na(+) channel related neurotransmitter release mechanisms during K(+)-evoked stage, and simultaneously enhances N-type Ca(2+) channel related basal serotonin release at the resting stage.
Abstract: 1. To elucidate possible mechanisms underlying the effects of carbamazepine (CBZ), valproate (VPA) and zonisamide (ZNS) on neurotransmitter exocytosis, the interaction between these three antiepileptic drugs (AEDs) and botulinum toxins (BoNTs) on basal, Ca(2+)- and K(+)-evoked release of dopamine (DA) and serotonin (5-HT) were determined by microdialysis in the hippocampus of freely moving rats. 2. Basal release of monoamine was decreased by pre-microinjection of the syntaxin inhibitor, BoNT/C, but only weakly affected by the synaptobrevin inhibitor, BoNT/B. Ca(2+)-evoked release was inhibited by BoNT/C selectively. K(+)-evoked release was reduced by BoNT/B predominantly and BoNT/C weakly. 3. Perfusion with low and high concentrations of CBZ and ZNS increased and decreased basal monoamine release, respectively. Perfusion with VPA increased basal 5-HT release concentration-dependently, whereas basal DA release was affected by VPA biphasic concentration-dependently, similar to CBZ and ZNS. This stimulatory action of AEDs on basal release was inhibited by BoNT/C predominantly. 4. Ca(2+)-evoked monoamine release was increased by low concentrations of CBZ, ZNS and VPA, but decreased by high concentrations. These effects of the AEDs on Ca(2+)-evoked release were inhibited by BoNT/C, but not by BoNT/B. 5. K(+)-evoked monoamine release was reduced by AEDs concentration-dependently. The inhibitory effect of these three AEDs on K(+)-evoked release was inhibited by BoNT/B, but not by BoNT/C. 6. These findings suggest that the therapeutic-relevant concentration of CBZ, VPA and ZNS affects exocytosis of DA and 5-HT, the enhancement of syntaxin-mediated monoamine release during resting stage, and the inhibition of synaptobrevin-mediated release during depolarizing stage.
Abstract: We examined the employment and marital status of adult patients with epilepsy who did not have mental retardation and who had been treated at Hirosaki University Hospital, Hirosaki, Japan, for more than 5 years. The present study included 278 patients (142 males and 136 females) ranging from 20 to 60 years of age. We investigated the occupational status of the subjects and found that 168 had permanent jobs, but 41 patients were unemployed at the time of this survey. The proportion of the patients whose seizures were controlled at the time of this survey was 68% (114/168) in the group having permanent jobs, and 22% (9/41) in the unemployed group. Forty cases answered that they had resigned from their jobs due to occurrence of epileptic seizures. Of these patients, 13 were dismissed and 27 resigned voluntarily due to the potential for seizures. As to relationship between jobs and neuropsychiatric complications, the incidence of a past history of psychotic states in the unemployed group was significantly higher than that in the employed group. As to marital status, 13 males and 16 females (n = 29) had experienced divorce. Seven cases (two males and five females) had answered that epilepsy had been the reason for their divorce. We conclude that epilepsy or epileptic seizures have various negative effects on the patient's social life.
Abstract: To explore the pathogenesis of benign familial neonatal convulsions (BFNC), we determined effects of KCNQ-related M-channels (KCNQ-channels) on hippocampal glutamate (Glu) and gamma-aminobutyric acid (GABA) releases using microdialysis, and propagation of evoked field-potentials (FP) using multielectrode (64-ch)-dish system as two-dimensional monitoring. KCNQ-channel inhibitor, Dup996, enhanced hippocampal K(+)-evoked Glu and GABA releases without affecting basal releases of them. Dup996 unaffected FP-amplitude, but enhanced FP-propagation. The GABA(A)-receptor antagonist, bicuculline, enhanced the stimulatory effects of Dup996 on FP-propagation, however, this stimulatory effects of Dup996 were abolished by the alpha-amino-3-hydroxy-5-methylisoxazole-4-propionic acid (AMPA)/glutamate-receptor antagonist, DNQX. These results suggest that the occurrence of BFNC cannot be produced by KCNQ-channel dysfunction alone, but by reciprocal action between impaired KCNQ-channel and other unknown elements (possibly dysfunction of inhibitory neurotransmission system).
Abstract: To clarify the mechanisms of action of carbamazepine (CBZ), we investigated the effects of CBZ on acetylcholine (ACh) release and metabolism in rat striatum and hippocampus. Acute administration of effective dose of CBZ (25 mg/kg) increased both striatal and hippocampal extracellular levels of ACh, whereas a supraeffective dose of CBZ (50 mg/kg) did not affect the levels and a toxic dose of CBZ (100 mg/kg) decreased the extracellular ACh levels in both brain regions. Both acute and chronic administrations of CBZ (25 and 50 mg/kg, mg/kg per day) increased intracellular ACh levels in striatum and hippocampus. The striatal intracellular ACh levels were decreased by both acute and chronic administrations of CBZ (100 mg/kg, mg/kg per day), whereas the hippocampal intracellular ACh levels were not affected. The effective CBZ concentration did not affect cholinesterase activity, whereas supraeffective CBZ concentration reduced it weakly. Effective dose of CBZ enhanced ACh release and synthesis; however, supraeffective doses of CBZ reduced ACh release and synthesis without enhancement of ACh degradation, indicating that CBZ has biphasic effects on ACh release and synthesis. Thus, the present findings, the slight stimulation of ACh function by effective dose of CBZ, are involved, at least partially, in the antiepileptic and mood stabilizing mechanisms of action of CBZ.
Abstract: Effects of aromatherapy (odorless condition, lavender, and hiba oil) on mood and anxiety were investigated in 14 female patients who were being treated with chronic hemodialysis. A control period consisting of natural hospital smells was established before each test session, and then aromatic test conditions were systematically evaluated for odorless conditions as well as aromatic conditions containing lavender and hiba oil aromas. The effects of aromatherapy were measured using the Hamilton rating scale for depression (HAMD) and the Hamilton rating scale for anxiety (HAMA). Hiba oil aroma significantly decreased the mean scores of HAMD and HAMA, and lavender aroma significantly decreased the mean scores of HAMA. The mean scores of HAMD and HAMA in an odorless condition were not significantly different from those of the control conditions. These results indicate that in chronic hemodialysis patients hiba oil is an effective, non-invasive means for the treatment of depression and anxiety, and that lavender alleviates anxiety.
Abstract: Epilepsy is a common neurological disease and encompasses a variety of disorders with paroxysms. Although there is a genetic component in the pathogenesis of epilepsy, the molecular mechanisms of this syndrome remains poorly understood. Linkage analysis and positional cloning have not been sufficient tools for determining the pathogenic mechanisms of common idiopathic epilepsies, and hence, novel approaches, based on the etiology of epilepsy, are necessary. Recently, many paroxysmal disorders, including, epilepsy, have been considered to be due to ion channel abnormalities or channelopathies. Results of recent studies employing gene analysis in animal models of epilepsy and human familial epilepsies support the hypothesis that at least some of the so called idiopathic epilepsies, i.e. epilepsies currently, classified as idiopathic could be considered as a channelopathy. This hypothesis is consistent with the putative prerequisites for genes responsible for the majority of idiopathic epilepsies that can adequately explain the following characteristics of epilepsy. Neuronal hyperexcitability, dominant inheritance with various penetrance, pharmacological role of some conventional antiepileptic drugs, age dependency in the onset of epilepsy, and the involvement of genetic factors in the pathogenesis of post-traumatic epilepsy. Search for mutations in ion channels expressed in the central nervous system may help in finding defects underlying some of idiopathic epilepsies, thereby enhancing, our understanding of the molecular pathogenesis of epilepsy. A working hypothesis to view certain idiopathic epilepsies as disorders of ion channels should provide a new insight to our understanding of epilepsy and allow the design of novel therapies.
Abstract: The effects of purinoceptor subtypes on hippocampal extracellular serotonin levels were determined by using in vivo microdialysis. Perfusion with adenosine-5'-triphosphate (ATP) for 20 min produced concentration-dependent changes in hippocampal extracellular serotonin levels, which consisted of an initial rise phase, with levels increasing to 309% of control with 100 microM ATP, followed by a later rebound reduction phase, with levels decreasing to 6% of control. The P2X1-7 active P2 purinoceptor agonist, 2-methylthioATP (2-MeSATP: 100 microM) increased the extracellular serotonin level drastically (638%), while the P2X1,3 active P2 purinoceptor agonist, alpha, beta-methylene-L-ATP (alpha, beta-meATP: 100 microM) produced a small increase (132%) in the serotonin level. The P2X1,2,3,5,7 active P2 purinoceptor antagonist, suramin (100 microM), reduced the basal serotonin level (86%) and the ATP-evoked initial rise phase (from 309 to 254%) without affecting the late reduction phase. The adenosine A1 receptor antagonist, 8-cyclopentyl-1,3-dimethylxanthine (CPT: 50 microM) potentiated the rising phase (167%) and abolished the subsequent ATP-evoked reduction phase. Perfusion with CPT and an adenosine A2 receptor antagonist, 3,7-dimethyl-1-propargylxanthine (DMPX: 10 microM), reduced the ATP-evoked initial rise (to 181%) and abolished the late reduction phases of serotonin release. These results indicate that ATP-evoked hippocampal serotonin release is composed of an initial rise phase and a later reduction phase. The ATP-evoked initial rise phase might be produced by an activation of P2X purinoceptor function, whereas the late reduction phase was modulated by the activation of adenosine A1 receptor function by adenosine, metabolized from ATP in the synaptic cleft.
Abstract: To identify the major risk factors for the increased incidence of congenital malformations in offspring of mothers being treated for epilepsy with antiepileptic drugs (AEDs) during pregnancy and, to determine the relative teratogenic risk of AEDs, we prospectively analyzed 983 offspring born in Japan, Italy, and Canada. The incidence of congenital malformations in offspring without drug exposure was 3.1%, versus an incidence with drug exposure of 9.0%. The highest incidence in offspring exposed to a single AED occurred with primidone (PRM; 14.3%), which was followed by valproate (VPA; 11.1%), phenytoin (PHT; 9.1%), carbamazepine (CBZ; 5.7%), and phenobarbital (PB; 5.1%). The VPA dose and level positively correlated with the incidence of malformations. This study first determined a cut-off value of VPA dose and level at 1000 mg/day and 70 microg/ml, respectively, to avoid the occurrence of malformations. The incidence of malformations increases as the number of drugs increases, and as the total daily dose increases. Specific combinations of AEDs such as VPA + CBZ and PHT + PRM + PB produced a higher incidence of congenital malformations. The incidence of malformations was not associated with any background factors studied except for the presence of malformations in siblings. These results indicate that the increased incidence of congenital malformations was caused primarily by AEDs, suggesting that malformations can be prevented by improvements in drug regimen, and by avoiding polypharmacy and high levels of VPA (more than 70 microg/ml) in the treatment of epileptic women of childbearimg age.
Abstract: To study the mechanisms of antiepileptic action of zonisamide (ZNS), we determined the effects of ZNS on extracellular, total levels and re-uptake activity of serotonin (5-HT) in rat striatum and hippocampus. After acute administrations, plasma ZNS concentrations associated with anticonvulsive action (effective concentrations) increased the total levels of 5-HT, its metabolise (5-hydroxyindoleacetic acid: 5-HIAA) and precursor (5-hydroxytryptophan: 5-HTP). After chronic administration of ZNS, effective plasma concentrations also increased the extracellular and total levels of 5-HT, 5-HIAA, and 5-HTP. On the other hand, after both acute and chronic administrations of ZNS, a supra-effective ZNS concentration either decreased or did not affect the total levels of these substances. Therefore, the stimulatory effects of ZNS on the 5-HT system were reduced by an increase in ZNS concentration to supra-effective concentrations. ZNS concentrations of 30-1000 microM did not affect hippocampal 5HT re uptake activity in vitro. These results suggest that ZNS has biphasic effects on the 5-HT system, in that effective concentrations of ZNS enhance and supra-effective concentrations of ZNS reduce the function of the 5-HT system. These biphasic effects of ZNS on the 5-HT system may be involved in the mechanisms of action of the antiepileptic and psychotropic effects, and side effects of ZNS.
Abstract: To clarify the effects of zonisamide (ZNS) on neurotransmission and intracellular Ca2+ mobilization, both Ca2+ and K+ evoked hippocampal releases of dopamine (DA) and serotonin (5-HT) were determined by in vivo microdialysis, and K+ (25 and 50 mM) evoked elevation of intracellular Ca2+ level was determined by fluorescence microscopy in vitro. Therapeutic concentrations of ZNS had different effects on Ca2+ and K+ evoked release of monoamine. ZNS stimulated Ca2+ evoked monoamine release, while ZNS inhibited K+ evoked monoamine release. ZNS inhibited K+ evoked elevation of hippocampal intracellular Ca2+ levels in a concentration dependent manner. These results suggest that ZNS inhibits the depolarization induced by neuronal excitation, whereas ZNS might enhance the N-type Ca2+ channel activity or N-type Ca2+ channel related exocytosis mechanisms.
Abstract: Two hundred and one school refusers (< or =18 years old), excluding schizophrenia, were treated at the Department of Neuropsychiatry, Hirosaki University Hospital between April 1975 and March 1995. Of 56 cases of school refusal, 31 did not improve for more than 2 years (group P), and 25 cases had improved in the degree of school refusal and social impairment (group B). The remaining cases were excluded from the analysis for several reasons. There was no significant difference between groups B and P in age at the first psychiatric evaluation as well as the age of onset of school refusal. The duration from school absence to the first evaluation of group P was significantly longer than that of group B. The duration of school refusal significantly and positively correlated with the duration from school absence to the first evaluation. Introversion and nervousness prolonged the duration of school refusal. The non-presence of volition for school attendance, and a low frequency of school attendance during the 1 month prior to the first evaluation influenced the prolongation of school refusal. The 'duration from school absence to the first evaluation', the 'patient's character', the 'non-presence of volition for school attendance' and the 'frequency of school attendance' influenced the prolongation of school refusal. The introduction of treatment within 10 months of the onset of school refusal is an important factor in preventing the prolongation of the school refusal.
Abstract: To clarify the effects of adenosine receptor subtypes (A1, A2 and A3) on hippocampal serotonin (5-HT) release and 5-HT reuptake activity, hippocampal extracellular 5-HT levels were determined in vivo by microdialysis in freely moving rats. Selective 5-HT reuptake inhibitor (SSRI) fluoxetine and DU24565 increased extracellular 5-HT levels. Adenosine and A1 receptor agonist, 2-chloro-N6-cyclopentyl-adenosine (CCPA), decreased extracellular 5-HT levels, whereas non-selective antagonist, caffeine, and A1 antagonist, 8-cyclopentyl-1,3-dimethylxanthine (CPT) increased them. When 5-HT reuptake activity was inhibited by DU24565 and fluoxetine, the effects of CPT and CCPA on 5-HT level were enhanced. A2A receptor agonist, CGS21680, A2 receptor agonist, PD125944, A2 receptor antagonist, 3,7-dimethyl-1-propargylxanthine (DMPX), and A3 receptor agonist, N6-2-(4-aminophenyl)ethyladenosine (APNEA) did not affect 5-HT levels; however, when A1 receptor was blocked by CPT, 5-HT levels were increased by adenosine, CGS21680 and PD125944, and decreased by DMPX and APNEA. Under conditions of A1 receptor blockade, pretreatment with DU24565 or fluoxetine, enhanced the stimulatory effects of CGS21680 and PD125944 as well as inhibitory effects of DMPX on 5-HT level, whereas the inhibitory effect of APNEA was abolished. These results indicate that the stimulatory effects of A2 receptor and inhibitory effects of A3 receptor on hippocampal extracellular 5-HT levels are masked or abolished by the inhibitory effects of A1 receptor. In addition, hippocampal serotonergic transmission might be modulated by hippocampal presynaptic adenosine receptor subtypes, and hippocampal 5-HT reuptake activity might be modulated by hippocampal A3 receptor.
Abstract: The effects of magnesium ion (Mg2+), adenosine A1 and A2 receptor ligands on basal extracellular levels of striatal dopamine, hippocampal dopamine and serotonin were investigated by using in vivo microdialysis in freely moving rats. An increase in extracellular levels of Mg2+ ([Mg2+]o) reduced extracellular striatal and hippocampal monoamine levels, in a concentration dependent manner. The extracellular monoamine level was increased by perfusion with a non-selective adenosine receptor antagonist, caffeine, a selective adenosine A1 receptor antagonist, 8-cyclopentyl-1,3-dimethylxanthine (CPT), and an antiepileptic drug, carbamazepine, whereas adenosine, a selective adenosine A1 receptor agonist, 2-chloro-N6-cyclopentyladenosine (CCPA) reduced extracellular monoamine levels. The former and latter were reduced and enhanced by an increase in [Mg2+]o, respectively. Neither a selective adenosine A2 agonist, N6-[2-(3,5-dimethoxyphenyl)-2-(methylphenyl) ethyl]adenosine (PD125944), nor an antagonist, 3,7-dimethyl-1-propargylxanthine (DMPX), affected extracellular monoamine levels. Under the conditions of adenosine A1 receptor blockade by CPT, adenosine, PD125944 and carbamazepine increased extracellular monoamine levels, whereas caffeine and DMPX decreased them. These stimulatory effects of adenosine, PD125944 and carbamazepine were enhanced by an increase in [Mg2+], whereas the inhibitory effects of caffeine and DMPX were reduced by Mg2+, in a concentration dependent manner. It is concluded that an activation of adenosine A1 and A2 receptors decrease and increase extracellular levels of striatal dopamine, hippocampal dopamine and serotonin, respectively. These whereas the stimulatory effects of adenosine A2 receptor on extracellular levels of monoamine are masked by the inhibitory effects of adenosine A1 receptor. In addition, an increase in [Mg2+]o enhances and reduces the effects of adenosine receptor agonists and antagonists on extracellular monoamine levels, respectively.
Abstract: GAGA factor is known to remodel the chromatin structure in concert with nucleosome-remodeling factor NURF in a Drosophila embryonic S150 extract. The promoter region of the Drosophila fushi tarazu (ftz) gene carries several binding sites for GAGA factor. Both the GAGA factor-binding sites and GAGA factor per se are necessary for the proper expression of ftz in vivo. We observed transcriptional activation of the ftz gene when a preassembled chromatin template was incubated with GAGA factor and the S150 extract. The chromatin structure within the ftz promoter was specifically disrupted by incubation of the preassembled chromatin with GAGA factor and the S150 extract. Both transcriptional activation and chromatin disruption were blocked by an antiserum raised against ISWI or by base substitutions in the GAGA factor-binding sites in the ftz promoter region. These results demonstrate that GAGA factor- and ISWI-mediated disruption of the chromatin structure within the promoter region of ftz activates transcription on the chromatin template.
Abstract: 1. To elucidate the mechanisms regulating the release of striatal dopamine and its precursor, 3,4-dihydroxyphenylalanine (DOPA), we determined the effects of various Ca2+ channel antagonists, an N-type Ca2+ channel antagonist, omega-conotoxin GVIA, a P-type Ca2+ channel antagonist, omega-agatoxin IVA, and a Q-type Ca2+ channel antagonist, omega-conotoxin MVIIC, on the basal and Ca2+- and K+-evoked release of striatal dopamine and DOPA, by use of in vivo microdialysis. 2. Omega-conotoxin GVIA strongly inhibited striatal basal dopamine release (IC50 = 0.48 nM), whereas this toxin only weakly modulated basal striatal DOPA release (IC50 = 9.55 nM). Neither omega-agatoxin IVA nor omega-conotoxin MVIIC affected the basal striatal release of dopamine and DOPA. 3. Omega-conotoxin GVIA strongly inhibited Ca2+-evoked striatal dopamine release (IC50 = 0.40 nM), whereas Ca2+-evoked striatal DOPA release only was weakly modulated (IC50 = 10.51 nM). Neither omega-agatoxin IVA nor omega-conotoxin MVIIC affected the Ca2+-evoked release of striatal dopamine and DOPA. 4. Both omega-agatoxin IVA and omega-conotoxin MVIIC inhibited the K+-evoked release of striatal dopamine (IC50 of omega-agatoxin IVA = 2.65 nM; IC50 of omega-conotoxin MVIIC = 12.54 nM) and DOPA (IC50 of omega-agatoxin IVA = 0.15 nM; IC50 of omega-conotoxin MVIIC = 3.05 nM), whereas omega-conotoxin GVIA had no effect on the K+-evoked release of striatal dopamine and DOPA. 5. An increase in the extracellular Ca2+ and K+ concentrations (Ca2+- and K+-evoked stimulation) did not affect tyrosine hydroxylase activity in vivo. 6. These findings suggest that striatal DOPA release is neurotransmitter-like and that, unlike the mechanisms of striatal dopaminergic transmission, this striatal DOPA transmission is at least partly regulated by voltage-sensitive Ca2+ channels.
Abstract: 1. Multiple components of hippocampal glutamate release were examined by study of Ca2+- and K+-evoked hippocampal extracellular glutamate release using an in vivo microdialysis glutamate biosensor in urethane-anaesthetized rats. In addition, the effects of the antiepileptic drugs, carbamazepine (CBZ) and zonisamide (ZNS) perfused through the probe on glutamate release were assessed. 2. Basal glutamate levels were below detection limits (approximately 0.1 microM). An increase in extracellular KCl (from 2.7 to 50 and 100 mM) increased extracellular hippocampal glutamate levels to 9.2+/-1.4 and 20.0+/-2.6 microM, respectively, calculated from the area under curve (AUC) for 60 min. 3. This KCl-evoked glutamate release consisted of three components: an initial transient rise, a late gentle rise, and late multiple phasic transient rises. 4. An increase in or removal of extracellular CaCl2 levels respectively enhanced and reduced the 50 mM KCl-evoked hippocampal glutamate release (AUC for 60 min) from 9.2+/-1.4 to 12.4+/-2.1 and 5.8+/-0.9 microM. 5. Perfusion with 100 microM CBZ or 1 mM ZNS inhibited both the 50 mM KCl-evoked hippocampal glutamate release (AUC for 60 min) from 9.2+/-1.4 to 5.5+/-1.1 and to 5.8+/-1.3 microM, respectively, as well as the stimulatory effects of Ca2+ on KCl-evoked hippocampal glutamate release. 6. These results suggest that both CBZ and ZNS may reduce epileptiform events by inhibiting excitatory glutamatergic transmission.
Abstract: To establish the mechanism of action of the antiepileptic and psychotropic effects of carbamazepine (CBZ), its effects on serotonin (5-HT) transmission, metabolism and re-uptake activity in the rat hippocampus were studied. After acute and chronic administrations of 25 mg/kg CBZ, the plasma concentration of CBZ was found to be within the therapeutic range, whereas both acute and chronic administrations of 50 and 100 mg/kg CBZ resulted in a supratherapeutic plasma concentration. Acute administration of the therapeutic dose of CBZ resulted in an increase in the hippocampal extracellular and total level of 5-HT, its metabolite, 5-hydroxydoleacetic acid (5-HIAA) and its precursor, 5-hydroxytryptophan (5-HTP). The acute administration of 50 mg/kg CBZ resulted in an increase in the hippocampal levels of extracellular 5-HT and 5-HIAA as well as in the total levels of 5-HTP, whereas hippocampal levels of extracellular 5-HTP, total 5-HT and 5-HIAA remained unaffected. CBZ at a dose of 100 mg/kg decreased the levels of all of these substances. After chronic administration, 25 mg/kg/day CBZ increased hippocampal total levels of 5-HT, 5-HTP and 5-HIAA, whereas 100 mg/kg/day CBZ decreased all of these total levels. CBZ at a dose of 50 mg/kg/day decreased total levels of 5-HT, however neither total levels of 5-HIAA nor 5-HTP were affected. Both therapeutic and supratherapeutic plasma concentrations of CBZ inhibited 5-HTP accumulation, and did not affect 5-HT re-uptake activity in vitro. These results suggest that a therapeutic concentration of CBZ enhances 5-HT turnover and transmission, whereas a supratherapeutic concentration of CBZ inhibits 5-HT turnover and transmission without affecting 5-HT re-uptake activity. These effects of CBZ on serotonergic systems may be, at least partially, involved in the mechanisms of action of CBZ.
Abstract: To clarify the effects of carbamazepine (CBZ) on dopamine (DA) release and their metabolism, the extracellular and total levels of DA, its metabolites (DOPAC and HVA) and precursor, 3,4-dihydroxyphenylalanine (DOPA) in the striatum and hippocampus were studied. DA re-uptake and DOPA accumulation in the striatum and hippocampus, and monoamine oxidase (MAO) activities were also determined. After acute and chronic administrations of CBZ, the plasma concentration of CBZ associated with therapeutic activity increased the extracellular and total levels of all substances determined, whereas supratherapeutic concentration of CBZ decreased extracellular and total levels of all substances. Neither therapeutic nor supratherapeutic concentrations of CBZ affected MAO-A nor -B activities, nor DA re-uptake. DOPA accumulation caused by NSD1015 was inhibited by therapeutic and supratherapeutic concentrations of CBZ. These results suggest that a therapeutic concentration of CBZ enhances DA turnover, whereas a supratherapeutic concentration of CBZ inhibits DA turnover. These effects of CBZ on dopaminergic systems may be, at least partially, involved in the mechanisms of action of CBZ.
Abstract: To clarify the effects of adenosine receptor subtypes (A1, A2, and A3) on hippocampal serotoninergic function, hippocampal extracellular serotonin (5-HT) levels were determined by in vivo microdialysis in freely moving rats under various conditions. Both adenosine and an adenosine A1 receptor agonist, 2-chloro-N6-cyclopentyladenosine, decreased extracellular 5-HT levels, whereas an adenosine A1 receptor antagonist, 8-cyclopentyl-1,3-dimethylxanthine (CPT), and caffeine increased these levels. A selective A2A receptor agonist (CGS-21680), an adenosine A2 receptor agonist (PD-125944), an adenosine A2 receptor antagonist, 3,7-dimethyl-1-propargylxanthine (DMPX), and an adenosine A3 receptor agonist, N6-2-(4-aminophenyl)ethyladenosine (APNEA), did not affect extracellular 5-HT levels. When the adenosine A1 receptor was blocked by CPT, the hippocampal extracellular 5-HT level was increased by adenosine, CGS-21680, and PD-125944, and decreased by caffeine, DMPX, and APNEA. When both adenosine A1 and A2 receptors were blocked by CPT and DMPX, the extracellular 5-HT level was decreased by adenosine, caffeine, and APNEA. The hippocampal extracellular 5-HT level was not affected by administration of APNEA alone, but was decreased by this agent when the adenosine A1 receptor was blocked, irrespective of whether the adenosine A2 receptor was functional. These inhibitory effects of adenosine, caffeine, and APNEA on extracellular 5-HT levels, during both adenosine A1 and A2 receptor blockade, were inhibited by selective 5-HT reuptake inhibitors. These results indicate that the stimulatory effects of the adenosine A2 receptor and the inhibitory effects of the A3 receptor on hippocampal extracellular 5-HT levels are masked by the inhibitory effects of the adenosine A1 receptor.
Abstract: Of the epileptic patients who were treated for > or = 5 years until the end of 1990 and had more than four seizures in 1990, 63 patients had been treated without interruption until the end of 1995. We analyzed their clinical courses from 1990 to 1995 prospectively. More than half the subjects were diagnosed with temporal lobe epilepsy. Twenty cases had presumed etiology, and 32 had neuropsychiatric complications. Of the subjects whose seizures were not controlled with conventional antiepileptic drugs (AED), 11 cases demonstrated significant improvement when new AED; that is, lamotrigine, vigabatrin, clobazam, topiramate, tiagabine or CGP33101 were added. However, 10 patients did not respond to new AED. Presumed etiology, neuropsychiatric complications, multiple epileptic foci in EEG and abnormalities on head CT or MRI were characteristics of the patients whose seizures were resistant to new AED.
Abstract: To clarify the mechanisms of the antiepileptic activity of phenytoin (PHI), the effects of PHT on extracellular and total levels of monoamines (dopamine and serotonin), in rat striatum and hippocampus were studied. The plasma concentrations of PHT associated with therapeutic activity did not affect striatal and hippocampal extracellular levels of monoamines, whereas supratherapeutic concentrations of PHT decreased striatal and hippocampal extracellular levels of monoamines, in a concentration dependent manner. Toxic concentrations of PHT produced generalized seizures 'paradoxical intoxication' and an initial drastic decrease in striatal and hippocampal extracellular levels of monoamines before seizure onset, whereas the extracellular monoamines levels increased after seizures. In addition, the therapeutic concentrations of PHT did not affect monoamine turnover, whereas supratherapeutic concentrations of PHT inhibited monoamine turnover. These results suggest that monoaminergic transmission may not be involved in the antiepileptic mechanism of action of PHT, and that dysfunction of monoaminergic transmission can produce generalized tonic-clonic convulsions. Thus, the present study suggests that 'Paradoxical Intoxication' induced by toxic concentrations of PHT, at least partially, can be mediated by hypo-monoaminergic function in the brain.
Abstract: The effects of carbamazepine and caffeine on adenosine receptor subtypes were determined using in vivo microdialysis in an attempt to elucidate their different psychotropic mechanisms of action. Adenosine and a selective adenosine A1 receptor agonist decreased the striatal extracellular dopamine level, whereas caffeine, carbamazepine and a selective adenosine A1 receptor antagonist increased it, but neither an adenosine A2 receptor agonist nor an antagonist affected it. Under conditions of adenosine A1 receptor blockade, adenosine, carbamazepine and a selective adenosine A2 receptor agonist increased the striatal extracellular dopamine level, whereas caffeine and a selective adenosine A2 receptor antagonist decreased it. These results suggest that adenosine A1 receptor stimulation reduces the striatal extracellular dopamine level, and that adenosine A2 receptor stimulation under conditions of adenosine A1 receptor blockade increases it. Therefore, caffeine is an antagonist of both adenosine A1 and A2 receptor subtypes, and carbamazepine is an adenosine A1 receptor antagonist as well as an adenosine A2 receptor agonist. These properties support the hypothesis that the central actions of both carbamazepine and caffeine result from effects on both adenosine A1 and A2 receptors.
Abstract: To clarify differences in the operating mechanisms of adenosine receptor subtypes (A1 and A2), striatal extracellular dopamine levels under various conditions were determined by in vivo microdialysis. Adenosine (50 microM) as well as the selective A1 agonist, 2-chloro-N6-cyclopentyladenosine (CCPA; 1 microM) decreased striatal extracellular dopamine levels, while the selective adenosine A1 antagonist, 8-cyclopentyl-1,3-dimethylxanthine (CPT; 50 microM) and caffeine (100 microM) increased striatal extracellular dopamine levels. A selective A2a agonist, 2-[4-(2-carboxyethyl)phenethylamino]-5'-N-ethylcarboxamideadeno sine (CGS21680; 10 microM), a selective A2 agonist, N6-[2-(3,5-dimethoxyphenyl)-2-(methylphenyl)ethyl]adenosine (DPMA; 5 microM) and a selective A2 antagonist, 3,7-dimethyl-1-propargylxanthine (DMPX; 10 microM), did not affect extracellular dopamine levels. When the A1 receptor was blocked by CPT, extracellular dopamine levels were increased by adenosine and DPMA, decreased by caffeine as well as DMPX, and unaffected by CGS21680. These results indicate that the stimulatory effects of the A2 receptor on striatal extracellular dopamine levels are masked by the inhibitory effects of the A1 receptor.
Abstract: The effects of adenosine and magnesium ion (Mg2+) on striatal dopamine release were studied in awake rats by in vivo microdialysis. The mean striatal basal levels of dopamine release at Mg2+ free perfusate were 56.95 +/- 5.30 fmol/sample (for 20 min). By varying the Mg2+ levels in perfusate from 0 mmol/L to 1, 10 or 40 mmol/L, the dopamine release was inhibited by Mg2+ in a level-dependent manner. Perfusion with modified Ringer's solution containing zero Mg2+ and from 5 to 50 mumol/L adenosine, non-selective adenosine agonist, as well as 0.1 mumol/L 2-chloro-N6-cyclopentyladenosine (CCPA), selective adenosine A1 agonist, showed no effect on dopamine release. However, from 5 to 50 mumol/L adenosine and from 0.1 to 1 mumol/L CCPA plus Mg2+ (1 and 40 mumol/L) perfusion decreased the dopamine release. This inhibitory effect of adenosine and CCPA on striatal dopamine release was enhanced by an increase in extracellular Mg2+ levels. Levels of 50 mumol/L of 8-cyclopentyl-1,3-dimethylxanthine (CPT), a selective adenosine A1 receptor antagonist, in perfusate increased the dopamine release under conditions both with and without Mg2+. This stimulatory effect of CPT on striatal dopamine release was reduced by an increase in extracellular Mg2+ levels. As a result, CPT antagonized the inhibitory effects of adenosine and CCPA on dopamine release under conditions of the presence and absence of Mg2+. These results suggest that the inhibition of striatal dopamine release by adenosine was mediated by adenosine A1 receptor. This inhibition was intensified by Mg2+. This study also revealed that the concentrations of Mg2+, which ranged from physiological to supraphysiological, reduced the striatal dopamine release; furthermore it was found that the physiological concentration of Mg2+ potentiated the effects of adenosine agonists, but inhibited adenosine antagonist. Thus, the present study, using in vivo microdialysis preparations, suggests Mg2+ inhibits the calcium ion channels and enhances the adenosinergic function in the central nervous system.
Abstract: Effects of zonisamide (ZNS) on extracellular dopamine (DA), its precursor 3,4-dihydroxyphenylalanine (DOPA), its metabolites 3,4-dihydroxyphenylacetic acid (DOPAC) and homovanillic acid (HVA) levels in the striatum as well as hippocampus of freely moving rats were studied. Intracellular DA, DOPA, DOPAC and HVA levels, as well as DOPA accumulation as an index of tyrosine hydroxylase activity in the rat brain in vivo, DA re-uptake in the striatum and hippocampus, and monoamine oxidase (MAO) activities were also determined. Acute administrations of therapeutic ZNS doses (20 and 50 mg/kg) increased striatal extracellular DOPA levels, intracellular striatal and hippocampal DOPA levels, and stimulated DOPA accumulation in both brain regions. ZNS also increased striatal and hippocampal intracellular as well as extracellular DA and HVA levels, but decreased those of DOPAC levels. Chronic (3 weeks) administrations of therapeutic ZNS doses (20 and 50 mg/kg/day) increased intracellular DA, DOPA, DOPAC and HVA levels in striatum and hippocampus. ZNS-induced changes were greater in intracellular levels than in extracellular levels. Acute and chronic supratherapeutic ZNS dose (100 mg/kg) administration decreased intracellular levels of all substances detectable in both brain regions, and inhibited DOPA accumulation. Both subtypes of MAO (type A and type B) activities were weakly inhibited by ZNS. ZNS showed no effect on DA re-uptake in striatum nor in hippocampus. These results suggest that therapeutic ZNS doses increase DOPA accumulation as well as both intracellular and extracellular DA, DOPA and HVA levels. However, such doses also decrease extracellular and intracellular DOPAC levels by enhancing DA synthesis and/or by selectively inhibiting MAO-B activities. In addition, chronic therapeutic ZNS dose administration enhances DA synthesis, which results in increased intracellular DA, its precursor and its metabolites levels. On the other hand, both acute and chronic supratherapeutic ZNS dose administrations inhibit DA turnover. These ZNS effects on DA metabolism are at least partly involved in the mechanisms of action of ZNS.
Abstract: We have developed a sensitive immunoradiometric assay for PTH-related peptide (PTHrP) using a monoclonal antibody against PTHrP(1-34) and a polyclonal antibody against PTHrP(50-83), with recombinant human PTHrP(1-87) as the standard. The detection limit of the immunoradiometric assay was 0.5 pmol/L, and plasma PTHrP(1-87) concentrations in 110 healthy subjects were 0.8 +/- 0.01 pmol/L, with the upper limit of the normal range being 1.1 pmol/L. Increased circulating PTHrP(1-87) concentrations were demonstrated in all 46 cancer patients with hypercalcemia, but not in patients with primary hyperparathyroidism, chronic renal failure, or hypoparathyroidism. Normalization of serum calcium levels after resection of tumors was shown to correlate well with that of plasma PTHrP(1-87) concentrations in 2 cancer patients. High circulating PTHrP(1-87) levels were also demonstrated in 12 out of 13 hypercalcemic patients with adult T-cell leukemia/lymphoma and in 7 out of 8 hypercalcemic patients with non-Hodgkin's lymphoma especially of B-cell type. These results suggest that PTHrP is a major humoral factor responsible for the hypercalcemia frequently associated with adult T-cell leukemia/lymphoma and also with B-cell lymphoma.
Abstract: The effects of zonisamide and carbamazepine on extracellular and intraneuronal levels of monoamine, its precursor and metabolites were determined in rat striatum and hippocampus. DA re-uptake and monoamine oxidase activities were also determined. Acute administrations of therapeutic doses of zonisamide and carbamazepine increased the extracellular and intraneuronal levels of monoamine, however, supratherapeutic doses of zonisamide and carbamazepine decreased extra- and intracellular monoamine levels. Chronic administrations of therapeutic doses of zonisamide and carbamazepine increased intraneuronal levels of monoamine, while supratherapeutic doses of zonisamide and carbamazepine decreased. Monoamine oxidase type B activity was inhibited by zonisamide, and the IC50 values of inhibition by zonisamide for type B monoamine oxidase were 660 microM, however, carbamazepine did not affect monoamine oxidase activity. Zonisamide and carbamazepine showed no effect on monoamine re-uptake within therapeutically relevant concentrations. These results suggest that therapeutic doses of zonisamide and carbamazepine enhance monoamine turnover, while supratherapeutic doses of zonisamide and carbamazepine inhibit monoamine turnover. Those effects of zonisamide and carbamazepine on monoamine metabolism are at least partly involved in the mechanisms of action of zonisamide and carbamazepine.
Abstract: Oxetanocin-G (OXT-G), a potent antiviral agent, is a novel nucleoside isolated from the culture filtrate of Bacillus megaterium. We investigated the antiviral effect of oral administration of OXT-G for five days on woodchuck hepatitis virus (WHV), in vivo, using 12 woodchucks. Woodchucks were randomized into each of four treatment groups according to the dose of OXT-G. Two out of six woodchucks treated with 1.0 or 2.0 mg/kg/day of OXT-G died. After treatment with OXT-G, serum levels of WHV-DNA significantly decreased in all woodchucks. However, the antiviral effect was only partial and levels of serum WHV-DNA returned after the cessation of treatment. The amount of viral replicative intermediates was decreased in livers of woodchucks treated with OXT-G. Although further study of the toxicity of this compound would be essential before studies in man can be carried out, OXT-G has potent antiviral activity against WHV and may deserve evaluation as an antiviral agent in the treatment of chronic hepatitis B infections in humans.
Abstract: The relationships between the plasma concentrations of mianserin and desmethylmianserin at 18 h after initial dosing and those at steady state were studied in 19 depressed patients receiving 30 mg of mianserin at bedtime. Significant linear relationships were observed for mianserin, desmethylmianserin, and mianserin plus desmethylmianserin. The present study thus suggests that the plasma concentrations of these compounds after an initial dose of mianserin can be used for the prediction of an optimal dose.
Abstract: The effects of zonisamide (3-sulfamoylmethyl-1,2-benzisoxazole), a novel anticonvulsant, on extracellular levels of monoamine and its metabolite in the striatum and hippocampus, and Ca2+ dependent monoamine release in the striatum of freely moving rats were studied by microdialysis. Zonisamide increased dopamine, homovanillic acid and 5-hydroxyindoleacetic acid, and decreased 3,4-dihydroxyphenylacetic acid in the rat striatum. However, zonisamide showed no effect on Ca2+ dependent dopamine release in the rat striatum. In the hippocampus, zonisamide increased dopamine, homovanillic acid, serotonin and 5-hydroxyindoleacetic acid and decreased 3,4-dihydroxyphenylacetic acid. The present results suggest that zonisamide facilitates dopaminergic and serotoninergic neurotransmission but does not affect Ca2+ dependent dopamine release within therapeutic plasma concentrations.
Abstract: Adrenocorticotropin labeled with 125I at Tyr23 [(125I-Tyr23]ACTH) was prepared by radioiodination of ACTH1-39 followed by reverse phase HPLC purification. When incubated with bovine adrenal cortical membranes, the radioligand bound specifically to a 40-kDa membrane protein as revealed by affinity labeling. This result indicates that the bovine adrenal ACTH receptor, whose identification and characterization have proved difficult, has an Mr of about 40,000.
Abstract: Immobilized asparaginase was prepared by embedding asparaginase (which is effective for remission in children with leukemia) into fibrin polymer formed by fibrinogen-fibrin conversion in the presence of thrombin. The immobilized asparaginase film did not dissolve in 6 mol/1 urea, suggesting that blood coagulation factor XIII participates in the cross-linking between fibrins and between fibrin and asparaginase.
