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P.5.020 Targeted transport of PEGylatedimmunoliposomes to brain astrocytes
Poster Sessions barrier (detected through its occurrence in the blood). Rats with intracerebral transplants displayed significant levels of this brain-specific protein in the blood, as detected on the 7th and 14th postoperative days. In rats with wellconsolidated grafts without pronounced reactive gliosis, the GFAP concentration reached 253.99+79.30ng/ml by the 4th week and then decreased to 8.2+3.3 ng/ml by the 8th to 12th weeks after transplantation. In rats with poorly consolidated grafts characterized by the death of the transplanted neurons and gliosis in the graft and surrounding tissues, the GFAP concentration increased to 476.4+ 111.0 ng/ml within 4 weeks and remained elevated (235.0+44.8ng/ml) for a further 12 weeks. Thus, serum GFAP concentration correlated well with the extent of reactive gliosis, which allows the real functional state of the intracerebral graft to be assessed and prognosed in living animals.
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Targeted transport of PEGylated immunoliposomes to brain astrocytes
V.P. Chekhonin, Y.A. Zhirkov, O.I. Gurina, I.A. Rjabukhin, T.B. Dmitrieva. Serbsky National Research Center for
Social and Forensic Psychiatry, Moscow, Russia PEGylated (stealth) immunoliposomes directed against human gliofibrillary acidic protein (GFAP) were prepared by coupling the thiolated monoclonal anti-GFAP antibodies, D4, with a maleimide derivative of the phosphatidyl ethanolamine of the liposomal membrane. Depending on the initial protein-to-lipid ratios, the immunoliposomes prepared (with a diameter of about 70nm) were coupled with 60 to 240 molecules of the antibodies. In vitro experiments with cultures of embryonic rat brain astrocytes demonstrated a specific binding of these immunoliposomes, which could be inhibited by the preincubation of the cells with free non-coupled D4 but not with irrelevant antibodies. Administered intravenously into rats, the immunoliposomes displayed a kinetic behaviour typical of the PEGylated liposomes: after 24 h, approximately 20% of the doses injected were still present in the blood; the elimination rate constants were 0.050.09 h -], the elimination halflifes were 8-15 h. With such a systemic longevity, as well as with such a specificity, these immunoliposomes, although incapable of penetrating the unimpaired blood-brain barrier (BBB), should be useful in delivering pharmacological agents to glial brain tunaours (which continue to express GFAP) or to other pathological loci in the brain with a partially disintegrated BBB.
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s211
Influence of M- and N-cholinoblockers on principal manifestations of experimental parkinsonism induced by Nivalin and neurotoxine MPTP
N.A. Losev, N.S. Sapronov. Institute Experimental
Medicine, Russian Academy of Medical Sciences, St. Petersburg 197376, Russia The aim of the investigation was the study of the action of M- and N-cholinoblockers (ACI-IBs) on experimental parkinsonism imitating hyperfunction of cholinergic mechanisms in CNS. In male mice the injection of acetylcholine esterase inhibitor Nivalin (Galantamine, 10-15mg/kg) was followed by a decrease of locomotor activity (in 1-1.5min), the appearance of rigidity (in 4-5 min), tremor and muscular fasciculation of legs and body registered with myograph. In experiments M-ACHB metamizyl (methylbenactyzine, 2mg/kg) and N-ACHB Eterofen [2-(diethylamino) buthylester difenilacetic acid] (30mg/kg) were used. It was established that Eterofen injection led to disappearance of muscular fasciculation and to intensification of tremor; after subsequent introduction of metamizyl tremor was also deleted. When metamizyl was introduced firstly tremor was abolished and muscular fasciculation increased, however subsequent injection of Eterofen resulted in abolition of fasciculation. The results show that blockade of M-cholinoreactive biosystems is accompanied with increased activity of N-cholinoreactive systems and on the contrary. Therefore in this case reciprocal interactions between M- and N-cholinergic mechanisms take place. Neurotoxine 1-methyl-4-phenyl- 1,2,3,6-tetrahydropiridine (MPTP) was administered to 3 groups of animals in doses 20.0, 30.0 and 40.0 mg/kg i.p. daily during 20 days. MPTP-induced syndrome like tremor, hypokinesia and rigidity was inhibited by metamizyl (2.0-5.0mg/kg) and phenamine (10.0mg/kg), but it was potentiated by ganglerone (10.0-15.0 mg/kg) - N-cholinoblocker - and haloperidol (1.0 mg/kg) - dopaminoblocker. Thus, the data indicated interaction between cholinergic and dopaminergic neurotransmitter systems. Effects of metamizyl - M-cholinoblocker - and phenamine dopaminemimetic - were additional, but opposite to the effects of eterofen and ganglerone - N-cholinoblockers and haloperidol - dopaminoblocker. It is possible to believe that in the treatment of Parkinson patients, who are characterized with hyperfunction of M-cholinergic and deficit of dopaminergic mechanisms in the structures of the striapallidar system, it is advisable together with the blockade of M-cholinoreceptors to stimulate N-cholinoreceptors in neurons of compact substance nigra
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Targeted transport of PEGylated immunoliposomes to brain astrocytes
V.P. Chekhonin, Y.A. Zhirkov, O.I. Gurina, I.A. Rjabukhin, T.B. Dmitrieva. Serbsky National Research Center for
Social and Forensic Psychiatry, Moscow, Russia PEGylated (stealth) immunoliposomes directed against human gliofibrillary acidic protein (GFAP) were prepared by coupling the thiolated monoclonal anti-GFAP antibodies, D4, with a maleimide derivative of the phosphatidyl ethanolamine of the liposomal membrane. Depending on the initial protein-to-lipid ratios, the immunoliposomes prepared (with a diameter of about 70nm) were coupled with 60 to 240 molecules of the antibodies. In vitro experiments with cultures of embryonic rat brain astrocytes demonstrated a specific binding of these immunoliposomes, which could be inhibited by the preincubation of the cells with free non-coupled D4 but not with irrelevant antibodies. Administered intravenously into rats, the immunoliposomes displayed a kinetic behaviour typical of the PEGylated liposomes: after 24 h, approximately 20% of the doses injected were still present in the blood; the elimination rate constants were 0.050.09 h -], the elimination halflifes were 8-15 h. With such a systemic longevity, as well as with such a specificity, these immunoliposomes, although incapable of penetrating the unimpaired blood-brain barrier (BBB), should be useful in delivering pharmacological agents to glial brain tunaours (which continue to express GFAP) or to other pathological loci in the brain with a partially disintegrated BBB.
•
s211
Influence of M- and N-cholinoblockers on principal manifestations of experimental parkinsonism induced by Nivalin and neurotoxine MPTP
N.A. Losev, N.S. Sapronov. Institute Experimental
Medicine, Russian Academy of Medical Sciences, St. Petersburg 197376, Russia The aim of the investigation was the study of the action of M- and N-cholinoblockers (ACI-IBs) on experimental parkinsonism imitating hyperfunction of cholinergic mechanisms in CNS. In male mice the injection of acetylcholine esterase inhibitor Nivalin (Galantamine, 10-15mg/kg) was followed by a decrease of locomotor activity (in 1-1.5min), the appearance of rigidity (in 4-5 min), tremor and muscular fasciculation of legs and body registered with myograph. In experiments M-ACHB metamizyl (methylbenactyzine, 2mg/kg) and N-ACHB Eterofen [2-(diethylamino) buthylester difenilacetic acid] (30mg/kg) were used. It was established that Eterofen injection led to disappearance of muscular fasciculation and to intensification of tremor; after subsequent introduction of metamizyl tremor was also deleted. When metamizyl was introduced firstly tremor was abolished and muscular fasciculation increased, however subsequent injection of Eterofen resulted in abolition of fasciculation. The results show that blockade of M-cholinoreactive biosystems is accompanied with increased activity of N-cholinoreactive systems and on the contrary. Therefore in this case reciprocal interactions between M- and N-cholinergic mechanisms take place. Neurotoxine 1-methyl-4-phenyl- 1,2,3,6-tetrahydropiridine (MPTP) was administered to 3 groups of animals in doses 20.0, 30.0 and 40.0 mg/kg i.p. daily during 20 days. MPTP-induced syndrome like tremor, hypokinesia and rigidity was inhibited by metamizyl (2.0-5.0mg/kg) and phenamine (10.0mg/kg), but it was potentiated by ganglerone (10.0-15.0 mg/kg) - N-cholinoblocker - and haloperidol (1.0 mg/kg) - dopaminoblocker. Thus, the data indicated interaction between cholinergic and dopaminergic neurotransmitter systems. Effects of metamizyl - M-cholinoblocker - and phenamine dopaminemimetic - were additional, but opposite to the effects of eterofen and ganglerone - N-cholinoblockers and haloperidol - dopaminoblocker. It is possible to believe that in the treatment of Parkinson patients, who are characterized with hyperfunction of M-cholinergic and deficit of dopaminergic mechanisms in the structures of the striapallidar system, it is advisable together with the blockade of M-cholinoreceptors to stimulate N-cholinoreceptors in neurons of compact substance nigra