GABAergic excitotoxicity injury of the immature hippocampal pyramidal neurons' exposure to isoflurane.

BACKGROUND: Certain anesthetics exhibit neurotoxicity in the brains of immature but not mature animals. γ-Aminobutyric acid (GABA), the primary inhibitory neurotransmitter in the adult brain, is excitatory on immature neurons via its action at the GABAA receptor, depolarizing the membrane potential and inducing a cytosolic Ca2+ increase ([Ca2+]i), because of a reversed transmembrane chloride gradient. Recent experimental data from several rodent studies have demonstrated that exposure to isoflurane during an initial phase causes neuronal excitotoxicity and apoptosis. GABAA receptor-mediated synaptic voltage-dependent calcium channels' (VDCCs) overactivation and Ca2+ influx are involved in these neural changes. METHODS: We monitored [Ca2+]i using Fluo-4 AM fluorescence imaging. Using whole-cell patch clamp techniques, IVDCC (voltage-dependent calcium channel currents) were recorded from primary cultures of rat hippocampal neurons (5-day culture) exposed to isoflurane. To further investigate the neurotoxicity of high cytosolic-free calcium after isoflurane in a dose- and time-dependent manner, the possibility of increased caspase-3 levels was evaluated by Western blot and quantitative real-time polymerase chain reaction. Statistical significance was assessed using the Student t test or 1-way analysis of variance followed by the Tukey post hoc test. RESULTS: Under control conditions, isoflurane enhanced the GABA-induced [Ca2+]i increase in a dose-dependent manner. Dantrolene and nicardipine markedly inhibited this enhancement mediated by isoflurane. Moreover, in Ca2+-free media, pretreatment with isoflurane did not show any influence on the caffeine-induced increase of [Ca2+]i. Similarly, using whole-cell recording, isoflurane increased the peak amplitude of IVDCC in the cultured neurons from rat hippocampus by depolarization pulses. Isoflurane (0.25, 0.5, 0.75, and 1 minimum alveolar concentration [MAC]) potentiated IVDCC peak current amplitude by 109.11%±9.03%, 120.56%±11.46%, 141.33%±13.87%, and 146.78%±15.87%, respectively. To analyze variation in protein levels, the effect of treatments with isoflurane on caspase-3 activity was dose- and time-dependent, reaching a maximal caspase-3 activity after exposure to 1 MAC for 6 hours (P<0.001). However, in the mRNA levels, hippocampal caspase-3 mRNA levels began to be significantly increased in isoflurane-treated developing rat hippocampal neurons after 6 hours of exposure to 0.25 MAC isoflurane (P<0.001). CONCLUSIONS: Isoflurane-mediated enhancement of GABA-triggered [Ca2+]i release results from membrane depolarization with subsequent activation of VDCCs and further Ca2+-induced Ca2+ release from the ryanodine-sensitizing Ca2+ store. An increase in [Ca2+]i, caused by activation of the GABAA receptor and opening of VDCCs, is necessary for isoflurane-induced calcium overload of immature rat hippocampal neurons, which may be involved in the mechanism of an isoflurane-induced neurotoxic effect in the developing rodent brain.

Isoflurane enhances spontaneous Ca(2+) oscillations in developing rat hippocampal neurons in vitro.

BACKGROUND: During the nervous system development, spontaneous synchronized Ca(2+) oscillations are thought to possess integrative properties because their amplitude and frequency can influence the patterning of neuronal connection, neuronal differentiation, axon outgrowth, and long-distance wiring. Accumulating studies have confirmed that some drugs such as volatile anesthetic isoflurane produced histopathologic changes in the central nervous system in juvenile animal models. Because the hippocampus plays an important role in learning and memory, the present work was designed to characterize the Ca(2+) oscillations regulated by volatile anesthetic isoflurane in primary cultures of developing hippocampal neurons (5-day-cultured). METHODS: Primary cultures of rat hippocampal neurons (5-day-cultured) were loaded with the Ca(2+) indicator Fluo-4AM (4 microM) and were studied with a confocal laser microscope. RESULTS: Approximately 22% of 5-day-cultured hippocampal neurons exhibited typical Ca(2+) oscillations. These oscillations were dose-dependently enhanced by isoflurane (EC50 0.5 MAC, minimum alveolar concentration) and this effect could be reverted by bicuculline (50 microM), a specific gamma-aminobutyric acid (GABA(A)) receptor antagonist. CONCLUSION: Unlike its depressant effect on the Ca(2+) oscillations in adult neurons in previous researches, isoflurane dose-dependently enhanced calcium oscillations in developing hippocampal neurons by activating GABA(A) receptors, a major excitatory receptor in synergy with N-methyl-d-aspartate receptors at the early stages of development. It may be involved in the mechanism of an isoflurane-induced neurotoxic effect in the developing rodent brain.

[Material selection and structural design of simulated space module for field].

OBJECTIVE: To select the suitable material for the structure of simulated space cabin to meet the special requirements which the unitary metallic material cannot do. METHOD: The structural material was selected through comparison between the mechanical properties of fiber reinforced plastics (FRP) and a few conventional metallic materials. The content and arrangement of the fibers in the composite material were suitably designed according to load condition and structural shape of the cabin. RESULT: High strength and high stiffness, light weight, anti-fatigue and shock proof were achieved for the whole module structure. It meets the medical and hygienic standard for hazardous gases. CONCLUSION: The structural design of fiber glass reinforced plastics composite module was proved to be successful. It reduced the weight of the module body, and increased the strength and toughness of the whole module.