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.

The adenosine binding enhancer, PD 81,723, inhibits epileptiform bursting in the hippocampal brain slice.

The effects of adenosine and the adenosine binding enhancer, PD 81,723, on low magnesium-induced bursting in the in vitro hippocampal slice were examined. Extracellular recordings were obtained from the CA3 pyramidal cell layer while electrically stimulating in the stratum radiatum under low magnesium perfusion. Adenosine (6-100 microM) reduced the duration of epileptiform bursting in a dose-related manner, which was reversible upon washout of adenosine. Application of PD 81,723 (50-100 microM) also resulted in a dose-dependent reduction in the duration of the triggered burst, which was irreversible. These results demonstrate anticonvulsant activity of adenosine and the adenosine binding enhancer, PD 81,723, in the low magnesium model of epilepsy.

Adenosinergic modulation of the EEG and locomotor effects of the A2 agonist, CGS 21680.

The present study in rats was designed to identify the respective roles of A1 and A2 adenosine receptor activation in the anticonvulsant and behavioral actions of adenosine. Intracaudate injections of the highly selective A2 agonist, CGS 21680, did not affect caudate seizures. However, seizure threshold was increased in the presence of CGS 21680 after blockade of the A1 receptor with CPX, or following activation of the A1 receptor with R-PIA or NECA. Additionally, CGS 21680 led to a dose-related inhibition of locomotor activity when injected into the caudate. These results implicate the involvement of the A2 adenosine receptor in the locomotor depressant actions of adenosine and also suggest possible A2 anticonvulsant effects may depend upon the activation of the A1 receptor.

The A2-selective adenosine analog, CGS 21680, depresses locomotor activity but does not block amygdala kindled seizures in rats.

The actions of the highly selective A2 adenosine agonist, CGS 21680, in modulating kindled seizures and locomotor activity were examined. I.c.v. injections of CGS 21680 into the lateral cerebral ventricle in fully kindled rats were found to prolong the period of postictal EEG depression and reduce postictal spiking in a dose-dependent manner, while not affecting the behavioral seizure stage or afterdischarge duration. CGS 21680 injections also lead to a dose-related inhibition of locomotor activity in rats exposed to an open field apparatus compared to rats receiving control injections of saline. These observations implicate the involvement of the A2 adenosine receptor in postictal phenomena and the locomotor depressant actions of adenosine, but do not indicate a direct anticonvulsant activity following A2 activation.

Anticonvulsant effects of magnesium sulfate on hippocampal seizures: therapeutic implications in preeclampsia-eclampsia.

OBJECTIVE: The objective of this study was to determine whether magnesium sulfate has central anticonvulsant effects. STUDY DESIGN: In three experiments we investigated the anticonvulsant properties of magnesium sulfate on the hippocampus because of its high density of N-methyl-D-aspartate receptors and link to clinical epilepsy. Seizure activity was elicited in the hippocampus of rats or in in vitro hippocampal brain slices. The effects of magnesium on seizure activity were determined. RESULTS: (1) Intraperitoneal magnesium sulfate (270 or 360 mg/kg) failed to block hippocampal seizures but reduced electroencephalographic amplitude and seizure duration. (2) Injection of magnesium sulfate directly into a seizure focus blocked seizures and elevated seizure thresholds. (3) Low magnesium levels produced epileptiform activity in an in vitro hippocampal brain slice preparation; the epileptiform activity was reversed by increasing the magnesium concentration. CONCLUSIONS: Magnesium sulfate has central anticonvulsant activity on hippocampal seizures, implicating the N-methyl-D-aspartate receptor in eclamptic seizures and in the therapeutic efficacy of magnesium sulfate.

Functional activity of the adenosine binding enhancer, PD 81,723, in the in vitro hippocampal slice.

The adenosine receptor binding enhancer, PD 81,723, enhances the inhibitory effects of exogenously applied adenosine in a dose-dependent manner in hippocampal brain slices. Extracellular recordings were obtained from the CA1 cell layer while electrically stimulating the stratum radiatum. Application of 1, 10 or 32 microM PD 81,723 in the presence of adenosine resulted in a dose-dependent reduction in the amplitude of the population spike which could be partially reversed by theophylline. In addition, hippocampal slices exposed to adenosine showed greater paired-pulse facilitation compared to control and this facilitation was significantly enhanced by the presence of PD 81,723. PD 81,723 had no effect when administered alone, but required the presence of adenosine. These results demonstrate that in addition to enhancing adenosine receptor binding, PD 81,723 also enhances the functional activity of adenosine in the hippocampal slice.

Cardiorespiratory function is altered by picomole injections of 5'-N-ethylcarboxamidoadenosine into the nucleus tractus solitarius of rats.

A limited occipital craniotomy was conducted on urethane-anesthetized, spontaneously breathing rats to expose the caudal medulla in the region of the obex. Microinjections of 5'-N-ethylcarboxamidoadenosine (NECA), an adenosine analog, were made into the medial region of the caudal nucleus tractus solitarius (NTS) at the level of the caudal tip of the area postrema, an area of the NTS in which there is known to be a functional co-existence of cardiovascular and respiratory-related neuronal elements. Cardiorespiratory responses were subsequently recorded for a 60 min test period. Microinjections of NECA, in the dose range of 0.35-350 pmol per rat, produced significant dose-related reductions in respiratory rate which were accompanied by dose-dependent increases in tidal volume and these pronounced effects on respiration persisted throughout the test period. In contrast, the effects of NECA microinjections on cardiovascular parameters in this region of the NTS were bidirectional and elicited considerably more complex responses during the test period. During the initial period (2-5 min) following injection, NECA elicited significant hypotension (at lower doses) and pressor responses (at higher doses) in addition to significant bradycardia (at lower doses) whereas by the end of the 60 min test period, almost all doses of NECA had resulted in hypertension and tachycardia. Multivariate analysis of variance (MANOVA) and correlation statistics indicated that the effects of NECA on blood pressure during the initial 2-5 min were dose-dependent and unlikely related to depression of respiratory frequency. A further examination of the data by MANOVA indicated that the pharmacological effects of NECA during the 60 min test period exhibited a highly significant and specific dose-dependent and time-related response pattern for the respiratory, but not the cardiovascular, parameters. Taken together, these manifold response patterns suggest that the respiratory effects of NECA may be mediated by different intrinsic mechanisms in the NTS than are the cardiovascular effects of NECA. At the end of the 60 min test period following the administration of NECA, the respiratory rate remained profoundly depressed. In view of previous studies showing that microinjections of cyclic AMP analogs, forskolin, isoproterenol and adenosine into the same NTS sites elicit a similar depression of respiration, the results with NECA in the present study further support the notion that cyclic AMP may serve as a second messenger in NTS respiratory control regions and these respiratory depressant effects may be mediated by a single adenosine receptor subtype.(ABSTRACT TRUNCATED AT 400 WORDS)

Respiratory effects of 5'-ethylcarboxamidoadenosine, an analog of adenosine, following microinjections into the nucleus tractus solitarius of rats.

Microinjections of 5'-ethylcarboxamidoadenosine (NECA), an adenosine analog, were made into respiratory-related regions of the nucleus tractus solitarius (NTS) of spontaneously breathing rats and cardiorespiratory parameters were recorded during a 60-min test period. Microinjections of NECA, in the dose range of 0.35-350 picomol per rat, produced significant dose-related reductions in respiratory rate which were accompanied by correlative increases in tidal volume. At the end of the 60-min test period following the administration of NECA, the respiratory rate remained profoundly depressed, whereas blood pressure and heart rate were not significantly affected compared to preinjection control values. The data from this study suggest that adenosine may exert modulatory influences in brainstem respiratory control regions.