リボゾーム封入GABAのキンドリングけいれんに対する抑制効果

SUPPRESSIVE EFFECT OF GABA-CONTAINING LIPOSOMES ON KINDLED CONVULSION

　抄録　γ-アミノ酪酸（GABA）は血液脳関門（BBB）を通過し難いため，その末梢投与によっては抗けいれん作用を期待することはできない。一方，リポゾーム（LIPO）は天然の脂質を素材としたマイクロカプセルで，BBBを通過することが確認されている。本研究では，このLIPOにGABAを封入し（GABA-L），その腹腔内投与時の抗けいれん作用をラットの扁桃核キンドリングを用いて検討した。更に，3H-GABA-Lおよび3H-GABAをラットの腹腔内に投与して脳のradioactivityを求めた。GABA，およびLIPO単独では抗けいれん作用はみられなかったが，GABA-Lでは強い抗けいれん作用がみられた。また，3H-GABA-L投与後の脳のradioactivityは3H-GABA単独のそれと有意の差はなかったが，25％強の高い値を示した。以上の結果から，末梢投与GABA-Lの抗けいれん作用はGABA-Lにより脳内に搬入されたGABAにより発現されたものであること，更に，LIPO封入によるGABAの作用増強が関与していることを考察し，LIPOの臨床応用の可能性について言及した。

γ-aminobutyric acid (GABA), a major neuro-transmitter in the mammalian brain, may be natural mediator of defence against epileptic activi-ties. When given peripherally, however, GABA itself can cross the blood-brain barrier (BBB) in-sufficiently. Liposomes (LIFO), being composed of lipid bilayers in which various compounds can be entrapped, have been shown to cross the BBB. Recently, Loeb et al. demonstrated that intraperi-toneal injection of GABA entrapped within LIPO (GABA-L) significantly suppresses penicillin- or isoniazid-induced seizures in rats. In the present study, we examined the effect of intraperitoneallyadministered GABA-L on the kindled amygdala (AM) seizures in addition to the comparative uptake of radioactivity by brain following intraperitoneal administration of 3H-GABA-L or 3H-GABA in rats.

Expt. I: Aniticonvulsant effect of GABA-L

Twelve male Wistar rats weighing 250-300 g were used. Bipolar electrodes made of twisted stainless steel wire 0.2 mm in diameter were im-planted into the left AM. All the animals were kindled at the left AM until a stable kindled seizure was evoked for at least five successive days. Subsequently, the stimulus intensity was gradually reduced and the last intensity to induce the kindled seizure was designated as the generalized seizure triggerring threshold (GST).

GABA was dissolved in deionized water at a concentration of 0.5 g/ml. GABA-L and LIPO were prepared as follows. 80μmoles L-a-phos-phatidylcholine, 22.8μmoles stearylamine and 11.4 μmoles cholesterol were dissolved in chloroform. After chloroform was removed under a stream of nitrogen gas, dried thin film of phospholipids was formed. The thin film was then dispersed in 8 ml deionized water, containing 4 g GABA or without containing the amino acid, at about 50℃ by mechanically shaking. All the animals were tested with GABA, GABA-L and LIPO. All the drugs were intraperitoneally administered at a dose of 8 ml/kg, i. e., 4 g/kg GABA, one hour before the GST stimulation

Both GABA and LIPO had no influence on the kindled seizure induced by the GST stimulation. In contrast, GABA-L showed strong anticonvulsant activities, i. e., after-discharge generation was to-tally suppressed in 9/12 of the animals when the AM was stimulated at the GST, although the remaining 3 animals were less affected.

Expt. 2: Comparative uptake of radioactivity by brain following ³H-GABA-L and 3H-GABA injection.

Eight male Wistar rats weighing 150g were divided into two groups, i. e. A (N=4) and B (N=4). A and B groups received intraperitoneal injection of 125 μCi of ³H-GABA and ³3H-GABA-L, respectively. One hour after the injection, the animals were killed by decapitation. Their brains were solubilized using tolen, and the radioactivity was calculated in liquid scintillation counter.

The radioactivity in B group showed about 125% of that in A group, although there was no sig-nificant difference between the two groups.

The result of Expt. 1 is consistent with that of Loeb et al., who demonstrated an inhibitory effect of peripherally injected GABA-L on penicilline- orisoniazid-induced seizures in rats. Since GABA-L can probably pass the BBB, it is suggested that GABA-L, after entering the brain, protects con-vulsive seizures induced by the three models of epilepsy. This assumption receives further supportfrom the result of Expt. 2. The use of drugs based on specific neurotransmission mechanisms may provide improved clinical therapy for epilep-sy.