Hideyuki Shiraishi

Regional excitatory and inhibitory amino acid concentrations in pentylenetetrazol kindling and kindled rat brain

We determined regional concentrations of excitatory and inhibitory amino acids in pentylenetetrazol (PTZ) kindling and kindled rat brains in order to investigate the mechanisms responsible for the PTZ kindling process and kindled state. Compared with control rats, PTZ kindling rats had significantly higher concentrations of aspartate in the striatum and the temporal cortex 24 h after the 14th injection of PTZ. Glutamate and GABA concentrations were also significantly higher in the brainstem of PTZ kindling rats 24 h after the 14th injection of PTZ. These findings suggest that the striatum and temporal cortex may participate in a pathway for propagation of the PTZ kindling process, and that the brainstem may be a primary site of PTZ kindling epileptogenesis or a part of the pathways for propagation. On the other hand, the concentrations of amino acids in PTZ kindled rats and controls did not differ 4 weeks after the 14th injection. This suggests that the alterations we detected in amino acid metabolism are not related to retention of the PTZ kindled state, and that some other mechanism for it must exist.

ACTH therapy in infantile spasms: Relationship between dose of ACTH and initial effect or long-term prognosis

The relationship between the dose of ACTH and the initial effect was investigated in 41 children with infantile spasms. More than 0.015 mg (0.6 IU)/kg/day of ACTH was needed for a good initial response of seizures and electroencephalographic abnormalities. The relationship between the dose of ACTH and long-term prognosis was investigated in 29 patients. There was no relationship between the daily or total ACTH dosage, provided the dose was greater than 0.015 mg (0.6 IU)/kg/day, and the outcome of seizures and electroencephalographic abnormalities; however, ACTH 0.04-0.06 mg (1.6-2.4 IU)/kg/day and a total ACTH dose of 1.1-1.5 mg (44-60 IU)/kg resulted in better mental development than smaller doses of ACTH. Side effects of ACTH increased with dosage. Too small or too large a dose of ACTH does not lead to better mental development. The proper dose of ACTH should be used with careful attention to potential side effects.

Involvement of the noradrenergic system in the seizures of epileptic El mice

We studied the role of the noradrenergic system in the seizures of epileptic El mice. To this end, the anticonvulsant activity of adrenergic drugs was tested with a scoring method, and the binding of [3H]dihydroalprenolol, [3H]prazosin and [3H]yohimbine was evaluated in whole brains and various brain regions from stimulated and unstimulated El mice, and their maternal ddy mice. The seizures of El mice were inhibited by noradrenaline, phenylephrine, oxymetazoline, clonidine and yohimbine in a dose-dependent manner. These preventive effects of alpha-adrenoceptor agonists were antagonized by pretreatment with alpha-adrenoceptor antagonists. The preventive effect of yohimbine was reversed by pretreatment with clonidine or alpha-methyl-p-tyrosine, although the latter drug did not affect the anticonvulsant effect of clonidine. The binding of [3H]dihydroalprenolol was the same in the three groups of mice. More [3H]prazosin was bound in the cerebellum and striatum, and there were more [3H]yohimbine binding sites in the whole brain, cerebral cortex, hippocampus and brainstem of stimulated and unstimulated El mice than in the same areas of ddy mice. These findings suggest that up-regulated alpha 1- and alpha 2-adrenoceptors are involved in the inhibition of the seizures of El mice.

123I-IMP SPECT findings in mitochondrial encephalomyopathies

We performed N-isopropyl-[123I]p-iodoamphetamine (IMP) single photon emission computed tomography (SPECT) in three patients with Leigh syndrome, two patients with mitochondrial encephalomyopathy, lactic acidosis and stroke-like episodes (MELAS), and two siblings with progressive external ophthalmoplegia (PEO). The SPECT images were compared with the findings on magnetic resonance imaging (MRI) and computed tomography (CT). All Leigh syndrome patients showed low accumulation areas (LAA) bilaterally in the frontal lobes and the basal ganglia. The frontal lobe LAA was seen even in an area without abnormalities on CT/MRI. Each MELAS patient showed a focal LAA. SPECT could also detect an old stroke-like lesion that was no longer shown by CT/MRI. However, SPECT did not show LAA in the basal ganglia, which showed calcification on CT or abnormal signal intensity on MRI. MRI in the 2 PEO patients showed lesions bilaterally in the basal ganglia in one, and in the internal capsules in the other. SPECT showed LAA not only in corresponding areas, but also in the occipital lobes, where no lesions were revealed by MRI. Thus, 123I-IMP SPECT was more sensitive than CT/MRI for detecting stroke-like lesions in MELAS patients, although it did not detect small lesions in the basal ganglia. LAA in the frontal lobes and occipital lobes may be SPECT findings characteristic of Leigh syndrome and PEO, respectively.

Age- and seizure-related changes in noradrenaline and dopamine in several brain regions of epileptic El mice

Noradrenaline (NA) and dopamine (DA) levels in six brain regions of stimulated and nonstimulated El (El[s] and El[ns]) mice and their maternal ddY mice were determined at various ages and various times after a convulsion. The NA levels in the striatum and hippocampus of 12-week-old El[s] and El[ns] mice were lower than in ddY mice, and remained lower in 23-week-old El[s] mice, but not in El[ns] mice. DA levels were lower in the striatum of El[s] mice than in El[ns] and ddY mice at 16 and 23 weeks of age. NA levels decreased during seizure in the striatum and hippocampus of El[s] mice, and returned to preconvulsive levels 1 hr after convulsion in the striatum and 30 min in the hippocampus. DA levels in the striatum of El[s] mice decreased during convulsion and increased from 1 to 10 min after convulsion. These changes suggest that the NAergic systems in the striatum and hippocampus and the DAergic system in the striatum have important roles in relation to seizure susceptibility in El mice.

Depth EEG in mutant epileptic El mice: demonstration of secondary generalization of the seizure from the hippocampus

Mutant epileptic E1 mice are thought to have focal epilepsy of hippocampal origin because glucose utilization is increased in the hippocampus (HPC) during seizures in these mice. However, direct electrographic evidence is still lacking for the notion. We recorded electroencephalograms (EEGs) using depth electrodes in E1 and non-epileptic ddY mice. All the mice were subjected to a conventional seizure-provoking maneuver during EEG recording; each mouse was placed on a mesh floor and observed for 3 min, and then tossed up in the air. When the E1 mice showed signs of abortive seizures or prodromal symptoms including squeaking, running and myoclonus, sporadic spikes or sharp waves were generated exclusively in the HPC. When generalized convulsions followed these prodromes, the sporadic discharges evolved into a burst of generalized spikes which again predominated in the HPC. We also observed the cerebral cortex, amygdaloid, caudate, centro-median thalamic and ventral postero-lateral thalamic nuclei, all of which were found to be only secondarily involved. These findings provide the first electrical evidence that E1 mice have a secondarily generalized seizure that has its initiating focus in the HPC.

Effect of penicillin on GABA-gated chloride ion influx

To investigate the mechanism of penicillin-induced convulsions, we have studied the effects of penicillin G (PC-G) on GABA-gated chloride ion influx in brain ‘microsac’ preparations of mice. In the presence of 10−4 M GABA, PC-G inhibited GABA-gated chloride ion influx in a dose-dependent manner. The dose-response curve for GABA in the presence of 10−3 M PC-G was shifted rightward and there was a decrease in maximum response. The inhibitory effects of PC-G were not reversed by RO 15-1788, an antagonist of benzodiazepine (BZ) receptors, but were reversed by washing the ‘microsac’ membranes. Therefore, PC-G probably exerts its proconvulsant effect by inhibiting GABA-gated chloride ion influx. However, it appears not to act through the BZ receptor of the GABA/BZ receptor complex.

GABA-gated chloride ion influx in brains of epileptic El mice

GABA-gated chloride ion influx was measured in brain “microsac” preparations of epileptic El mice. There was significantly greater sensitivity to GABA in stimulated El mice (which had 14–18 convulsions induced at weekly intervals) than in unstimulated El mice (which had not experienced convulsions) or ddY mice. GABA-gated chloride ion influx was significantly decreased 20 min after a single convulsion, and returned to the preconvulsion level 60 min after a convulsion. These findings suggest that the functional state of GABA-gated chloride channel in El mice is changed secondarily by single or repeated convulsions.

High doses of penicillin decreases [3H]flunitrazepam binding sites in rat neuron primary culture

Penicillin (PC) neurotoxicity (convulsions and encephalopathy) is considered to be due to GABAergic inhibition. The effects of penicillin G(PCG) on [3H]flunitrazepam (FNZ) binding in rat neuron-enriched primary cultures was examined to assess the role of the benzodiazepine (BDZ) receptor in the neurotoxicity. PCG application for 24 h induced a significant decrease in [3H]FNZ binding activity at 10(-3) M, and a decrease in available receptor number (Bmax) at 10(-2) M, without obvious cell damage. Pre-application of the BDZ receptor antagonist, Ro-15-1788, prevented the PC-induced decrease in [3H]FNZ binding activity. Therefore, PC seems to reduce the number of BDZ receptors through a direct effect on this receptor, which is a part of the major inhibitory system in mammalian brain; the GABAergic macromolecular receptor complex. This decrease in BDZ receptors may play a role in PC-induced neurotoxicity, especially encephalopathy.

Decreased benzodiazepine receptor binding in epileptic El mice: A quantitative autoradiographic study

Benzodiazepine receptors and subtypes were examined in El mice and normal ddY mice with a quantitative autoradiographic technique. Specific [3H]flunitrazepam binding in stimulated El mice, which had experienced repeated convulsions, was significantly lower in the cortex and hippocampus than in ddY mice and unstimulated El mice. In the amygdala, specific [3H]flunitrazepam binding in stimulated El mice was lower than in ddY mice. There was a tendency for the [3H]flunitrazepam binding in these regions in unstimulated El mice to be intermediate between that in stimulated El mice and that in ddY mice, but there was no significant difference between unstimulated El mice and ddY mice. [3H]Flunitrazepam binding displaced by CL218,872 was significantly lower in the cortex of stimulated El mice than in that of the other two groups, and in the hippocampus of stimulated than of unstimulated El mice. These data suggest that the decrease in [3H]flunitrazepam binding in stimulated El mice may be due mainly to that of type 1 receptor and may be the result of repeated convulsions.