Fetal rat brain injury: effect of transient maternal hypoxemia.

OBJECTIVE: To determine whether transient acute maternal hypoxemia during the end of pregnancy may cause neuronal damage in fetal rat brains. STUDY DESIGN: Nine pregnant rats (4 study and 5 controls) at 16-17 gestational days were studied. The study rats were placed in a chamber and breathed a gas mixture of 11.8% oxygen, 4.95% CO2, and nitrogen for either 1 or 2 h, while the control animals breathed room air. Tail venous blood was collected and gases were evaluated at the beginning and conclusion of the exposure periods. After 72 h of recovery, at 19-20 days' gestation, the fetal cardiovascular systems were perfused with saline and formalin. The brains were embedded in paraffin, sectioned in coronal plane, and stained with hematoxylin and eosin Histologic assessment of sections was performed by a neuropathologist blinded to the protocol. Statistical analysis of the data was performed using analysis of variance and the chi-square test. RESULTS: Exposure to the gas mixture resulted in decreased maternal pO2 from 39.9 +/- 7.6 mm Hg in the control group to 28.8 +/- 2.0 mm Hg in the 2-hour hypoxia group (p < 0.05). No significant changes in maternal pH or pCO2 status have been noted. A total of 34 fetal rat brains served as controls and 26 brains as the hypoxia study group There was a significant increase in isolated neuronal damage, including necrosis and shrinkage of cells, with karyorrhexis (fragmentation and breakage of the nucleus) in the hippocampus, basal ganglia, thalamus, and hypothalamus in the hypoxemia rats as compared with controls. CONCLUSION: Transient maternal hypoxemia may cause neuronal necrosis in vulnerable regions of the fetal rat brain, including the hippocampus, basal ganglia, thalamus, and hypothalamus.

Effect of magnesium sulfate on excitatory amino acid receptors in the rat brain. I. N-methyl-D-aspartate receptor channel complex.

OBJECTIVE: Our purpose was to determine the effect of peripherally administered magnesium sulfate on the N-methyl-D-aspartate receptor channel complex in the rat central nervous system. STUDY DESIGN: Six rats were injected intraperitoneally with 270 mg/kg magnesium sulfate, followed by 27 mg/kg every 20 minutes for 4 hours. Controls (n = 6) received saline solution. Six rats received intraperitoneal injections of magnesium sulfate (270 mg/kg) every 4 hours for 24 hours and 6 received saline solution. Six rats received intraperitoneal magnesium sulfate (270 mg/kg) every 12 hours for 2 weeks and 6 received saline solution. Rats were subsequently perfused and killed and their brains dissected and frozen. Cryostate sections were taken, labeled in vitro by one of three ligands for autoradiography assay, and mounted on tritium-sensitive film for 4 weeks. The ligands were tritiated glutamate agonist, N-methyl-D-aspartate binding site; tritiated glycine agonist, glycine binding site; and tritiated MK-801 noncompetitive antagonist, channel site. Optical density measurements of binding of 11 brain regions on each section were performed with an image analyzing system. RESULTS: N-methyl-D-aspartate receptor binding in the hippocampus was higher than in all other brain regions in all three experiments. Systemic administration of magnesium sulfate for 24 hours resulted in reduced tritiated glutamate binding, whereas long-term administration (2 weeks) resulted in significantly decreased tritiated glycine binding in all brain regions sampled. Binding of tritiated MK-801 was significantly increased in both short- and intermediate-term administration of magnesium sulfate. CONCLUSIONS: These data suggest that short-term magnesium sulfate administration results in increased inhibition of the ion channel. This effect is also continued with prolonged treatment, along with decreased sensitivity of the N-methyl-D-aspartate receptor channel complex to its agonists glutamate and glycine. This proposed time-dependent, twofold effect may provide insight into the mechanisms of magnesium sulfate's central anticonvulsant effect.

Effect of magnesium sulfate on excitatory amino acid receptors in the rat brain. II. Kainate and alpha-amino-3-hydroxy-5-methylisoxazole-4-propionic acid receptors.

OBJECTIVE: Our purpose was to determine the effect of peripherally administered magnesium sulfate on kainate and alpha-amino-3-hydroxy-5-methylisoxazole-4-propionic acid receptors in the rat brain. STUDY DESIGN: Six rats were injected intraperitoneally with 270 mg/kg magnesium sulfate, followed by 27 mg/kg every 20 minutes for 4 hours. Controls (n = 6) received saline solution. Six rats received intraperitoneal injections of magnesium sulfate (270 mg/kg) every 4 hours for 24 hours and six received saline solution. Then 6 rats received intraperitoneal magnesium sulfate (270 mg/kg) every 12 hours for 2 weeks and six received saline solution. Rats were subsequently perfused and killed; their brains were dissected and frozen. Cryostat sections were labeled in vitro for autoradiography assay. The ligands used were tritiated kainate agonist, kainate binding site; tritiated alpha-amino-3-hydroxy-5-methylisoxazole-4-propionic acid agonist, and tritiated 6-cyano-7-nitroquinoxaline-2,3-dione antagonist, both at the alpha-amino-3-hydroxy-5-methylisoxazole-4-propionic acid binding site. RESULTS: Magnesium sulfate caused decreased binding of the agonist to the kainate receptor recognition site after both short-term and intermediate-term systemic administration, whereas long-term treatment resulted in increased binding. No significant consistent effect on the binding to the alpha-amino-3-hydroxy-5-methylisoxazole-4-propionic acid receptor agonist site was recorded after magnesium administration. The receptor antagonist showed an increased binding after short-term treatment. Long-term administration also resulted in increased binding of the antagonist, an effect that was limited to the hippocampus. CONCLUSIONS: These data suggest down-regulation of the kainate receptor population during short- and intermediate-term magnesium sulfate treatment. However, long-term inhibition by magnesium resulted in up-regulation of the receptor population. The results may also reflect an increased inhibitory effect of magnesium sulfate on the alpha-amino-3-hydroxy-5-methylisoxazole-4-propionic acid receptor.

Anticonvulsant Effects of Magnesium Sulfate in Hippocampal-Kindled Rats.

The objective of the present study was to determine whether magnesium sulfate has anticonvulsant actions in the hippocampal-kindled rat model of epilepsy. Fully kindled rats received acute intraperitoneal injections of magnesium sulfate (270 mg/kg), phenytoin (20 mg/kg) or saline in random order. Electrical seizure duration, behavioral seizure stage and duration of postictal EEG depression were examined 15, 30 and 60 min after injection. In an additional group of rats, kindled seizures were measured before and after chronic (2 h) intraperitoneal injections of magnesium sulfate versus saline. There was a significant decrease in electrical seizure duration (p < 0.01) and behavioral seizure stage (p < 0.01) with acute magnesium sulfate injections compared to saline injections. Phenytoin had no statistically significant effects on hippocampal-kindled seizures. Chronic magnesium sulfate treatment significantly reduced behavioral seizure stage at 2, 24, and 48 h postinjection (p < 0.05), but did not affect seizure duration. There was a significant time by treatment effect for magnesium sulfate on postictal EEG depression (p < 0.01). We conclude that in this model of hippocampal epilepsy-induced (kindled) rats, magnesium sulfate has significant anticonvulsant effects. Copyright 1995 S. Karger AG, Basel

Magnesium sulfate versus phenytoin for seizure prevention in amygdala-kindled rats.

OBJECTIVE: Magnesium sulfate is widely used for seizure prophylaxis in preeclampsia-eclampsia. However, its anticonvulsant effects in other types of seizures have not been proved. Diphenylhydantoin has been widely characterized as the "gold standard" of anticonvulsants. In this study we compared the anticonvulsant effects of therapeutic blood levels of magnesium sulfate and phenytoin in seizures generated in amygdala-kindled rats. STUDY DESIGN: Eighteen male rats had a bipolar electrode stereotaxically implanted into the central nucleus of the amygdala. After recovery an electrical seizure threshold was determined for each rat. Rats were stimulated twice daily at their seizure thresholds (i.e., kindling) until three consecutive generalized tonic-clonic seizures occurred. Kindled rats randomly received one of the following intravenous injections in a volume of 1.5 ml/kg: saline solution, magnesium sulfate (30, 60, or 90 mg/kg), or phenytoin (12.5, 25, or 50 mg/kg). Fifteen minutes after injection rats were stimulated at their seizure thresholds, and electrical and behavioral seizure activity was assessed. Statistical comparisons were made by analysis of variance and post hoc comparisons when appropriate. RESULTS: Magnesium sulfate had no effect on any of the seizure parameters assessed. Phenytoin significantly reduced seizure duration (p < 0.01), duration of postictal depression (p < 0.01), and behavioral seizure stage (p < 0.01). CONCLUSION: Amygdala-kindled seizures are more potently inhibited by phenytoin than by magnesium sulfate.

Magnesium is more efficacious than phenytoin in reducing N-methyl-D-aspartate seizures in rats.

OBJECTIVE: Although magnesium sulfate is one of the most commonly used agents for seizure prophylaxis in preeclampsia, its efficacy relative to other anticonvulsants is incompletely investigated. The underlying mechanisms of eclamptic seizures are unknown, and there is currently no universally accepted animal model for eclampsia. However, one commonly used method for studying the relative efficacy of antiepileptic drugs is through their effect on N-methyl-D-aspartate-induced seizures. Our aim was to compare the anticonvulsant effects of phenytoin and magnesium sulfate in an N-methyl-D-aspartate-induced seizure model. STUDY DESIGN: Twenty-one female rats were each stereotaxically implanted with a chronic indwelling bipolar recording electrode in the hippocampus and an injection cannula in the lateral cerebral ventricle. After 7 days animals were randomly given 90 mg/kg magnesium sulfate (n = 7), 50 mg/kg phenytoin, or saline solution (n = 7) intravenously. Fifteen minutes after the infusions animals were given 20 micrograms/microliters N-methyl-D-aspartate by direct intraventricular injection, and seizure activity was assessed for 20 minutes thereafter. All data were analyzed with the Mann-Whitney test. RESULTS: When compared with saline solution controls, total duration of seizure activity in animals treated with magnesium sulfate was significantly decreased (p < 0.05) and time to onset of seizure activity was significantly increased (p < 0.05). However, rats that received phenytoin did not show significant changes in these parameters. The post-N-methyl-D-aspartate seizure mortality rate was 50% in the saline solution controls and 29% in the phenytoin group, whereas none of the rats that received magnesium sulfate died. CONCLUSION: These results suggest that magnesium sulfate is a significantly more effective prophylactic agent than phenytoin for N-methyl-D-aspartate-induced seizures.

Magnesium sulfate treatment decreases N-methyl-D-aspartate receptor binding in the rat brain: an autoradiographic study.

OBJECTIVE: We determined the effect of peripherally administered magnesium sulfate on N-methyl-D-aspartate (NMDA) receptor binding capacity in various regions of the rat brain. METHODS: Three separate experiments were performed. 1) Six rats were injected intraperitoneally with 270 mg/kg of magnesium sulfate, followed by 27 mg/kg every 20 minutes for 4 hours; controls (n = 6) received saline. 2) Six rats received intraperitoneal injections of magnesium sulfate (270 mg/kg) every 4 hours for 24 hours, while six received saline. 3) Six rats received intraperitoneal magnesium sulfate (270 mg/kg) every 12 hours for a total of 2 weeks, and six received saline. Rats were subsequently perfused and sacrificed, and their brains were dissected, rinsed, and frozen. Cryostat sections were taken, labeled by in vitro [3H]-CGP 39653, assayed autoradiographically, and mounted on Ultrofilm for 4 weeks. Optical density measurements of binding on each section were performed using an image analyzing system. Eleven brain regions were sampled: 1, 2) frontal and occipital cortex; 3-7) hippocampus--CA-1, CA-3, stratum radiatum, stratum oriens, dentate gyrus; 8) thalamus; 9) hypothalamus; 10) caudate nucleus; and 11) cerebellum. RESULTS: The NMDA receptor binding density in the hippocampus was significantly higher than in all other brain regions in all three experiments. In experiment 1, there was no significant effect on NMDA receptor binding. However, prolonged systemic administration of magnesium sulfate for 24 hours resulted in significantly reduced [3H]-CGP binding in all brain regions sampled. After chronic magnesium sulfate administration (2 weeks), the [3H]-CGP binding was still reduced in the cortex and some regions of the hippocampus; however, there was no significant change in other regions. CONCLUSIONS: Peripheral treatment with magnesium sulfate results in a significant reduction in the NMDA receptor binding capacity in the rat brain. These results support the hypothesis that magnesium central activity is mediated, at least in part, via the NMDA receptor.

Stimulation and inhibition of N-methyl-D-aspartate receptors in rats: developing a seizure model.

OBJECTIVE: The objective of this study was to develop an experimental rat hippocampal seizure model based on the stimulatory effects of N-methyl-D-aspartate and to determine the inhibitory effects of MK-801 on N-methyl-D-aspartate-induced seizures. STUDY DESIGN: Two separate experiments were performed. In the first experiment chemitrode-implanted rats were injected intracranially with increasing doses (5, 10, 20, and 30 micrograms) of N-methyl-D-aspartate into the hippocampus. Various electrophysiologic and behavioral parameters were examined to determine the dose required to reliably elicit hippocampal seizure activity without having toxic effects on the rats. In the second experiment rats were given an intraperitoneal injection of MK-801 (0.5 or 1 mg/kg), followed 20 minutes later by an intracranial injection of N-methyl-D-aspartate (20 or 30 micrograms). The ability of MK-801 to suppress N-methyl-D-aspartate-induced seizure activity was assessed in this experiment. RESULTS: Intrahippocampal injection of 20 micrograms of N-methyl-D-aspartate produced the shortest electrical seizure latency (193 +/- 72 seconds, p < 0.01). At this dose seizure was achieved in 80% (four of five of the animals, and the highest numbers of electrical seizures per animal were produced (2.2 +/- 0.8, p < 0.05). The group that received 30 micrograms of N-methyl-D-aspartate had a shorter latency, a longer duration of behavioral seizure and a higher number of behavioral seizures (p < 0.05). However, this group suffered a 60% (three of five) mortality rate. The addition of MK-801 significantly decreased the number of seizures per animal and the total seizure duration (p < 0.05). MK-801 also reduced the latency period. CONCLUSION: Intracranial injection of 20 micrograms of N-methyl-D-aspartate produced reliable hippocampal seizure activity without mortality. MK-801 at a dose of 1 mg/kg injected intraperitoneally had significant inhibitory effects on this seizure model.

Central anticonvulsant effects of magnesium sulfate on N-methyl-D-aspartate-induced seizures.

OBJECTIVE: The objective of this study was to determine if magnesium sulfate's central anticonvulsant activity is effective against N-methyl-D-aspartate-induced seizures. STUDY DESIGN: In two separate experiments we investigated magnesium sulfate's ability to inhibit N-methyl-D-aspartate-induced hippocampal seizures in rats. In the first experiment magnesium sulfate was administered peripherally before an intracranial injection of 20 micrograms of N-methyl-D-aspartate. In the second experiment magnesium sulfate was injected intracranially concurrently with N-methyl-D-aspartate. The ability of magnesium sulfate to suppress N-methyl-D-aspartate-induced seizure activity under both conditions was assessed. RESULTS: Peripherally administered magnesium sulfate significantly increased the latency from the time of an N-methyl-D-aspartate injection to the first seizure both by acute injection and after 2 hours of sustained elevation of serum magnesium levels when compared with saline solution-injected controls (p < 0.01). The duration of the first seizure was also significantly reduced. Intracranially administered magnesium sulfate significantly (p < 0.01) increased the seizure latency period by 120%. Overall, central magnesium sulfate prevented seizure activity in 40% of the animals (p < 0.01). CONCLUSION: Magnesium sulfate has a central anticonvulsant action on N-methyl-D-aspartate-induced seizures in this rat model of hippocampal seizures.

Peripheral magnesium sulfate enters the brain and increases the threshold for hippocampal seizures in rats.

OBJECTIVES: Our objectives were to determine whether magnesium sulfate crosses the blood-brain barrier and whether it has central anticonvulsant action. STUDY DESIGN: In experiment 1 34 female Long-Evans rats were divided into six groups: control (n = 7); single magnesium sulfate injection and evaluation after 20 minutes in 3 conditions: normal rats (n = 7), sham-operated animals (n = 5), and after electrical stimulation by hippocampal electrode (n = 5); single injection and evaluation after 2 hours (n = 5); and prolonged (2 hours) serum magnesium elevation (n = 5). Serum, cerebrospinal fluid, and specific brain areas were analyzed for magnesium concentrations. In experiment 2 threshold for electrical seizure was measured in eight rats before and after intraperitoneal injections of magnesium sulfate versus saline solution. RESULTS: In experiment 1 there was a significant correlation between blood and cerebrospinal fluid magnesium concentrations (r = 0.80, p < 0.0001). Magnesium concentrations were increased in the cortex and hippocampus, with the largest changes occurring after two hours of sustained serum magnesium concentrations (p < 0.01). Induction of hippocampal seizure activity resulted in further elevations in cerebrospinal fluid magnesium concentrations but did not change brain concentrations. In experiment 2 magnesium sulfate increased the electrical threshold required to induce seizures by 34% (p = 0.01). CONCLUSIONS: Magnesium sulfate enters the cerebrospinal fluid and brain and has a central anticonvulsant effect.

Effect of eye removal on N-acetylaspartylglutamate immunoreactivity in retinal targets of the cat.

The endogenous brain dipeptide N-acetylaspartylglutamate (NAAG) has previously been demonstrated in the somata of retinal ganglion cells and the neuropil of retinal targets. In this paper we report that the NAAG immunoreactivity of the neuropil in the retinal targets is dependent on an intact optic pathway. Removal of one eye produced a marked decrease in the staining of the neuropil in layer A of the contralateral geniculate nucleus (LGN) and layer A1 of the ipsilateral LGN. There was also decreased staining in the superficial layers of the superior colliculus contralateral to the removal. These results suggest that NAAG is present in the terminals of retinal ganglion cells and is consistent with a role for NAAG in visual synaptic transmission.