Bicuculline-induced Seizures Research Articles

Electroencephalographic evidence for a dose-related biphasic effect of morphine on bicuculline-induced seizures in the rat

The effects of systemic pretreatment with morphine sulfate (MS) on electroencephalographic (EEG) manifestations of seizures due to bicuculline (BIC) were studied in freely moving rats. Electrodes were placed on frontal and occipital cortices, into the hippocampus and into a region of the prepiriform cortex, area tempestas (AT). BIC (0.1–0.25 mg/kg i.v.) elicited dose-dependent changes in electrocortical activity, consisting of 3 stages: stage 1, slight increase in periods of desynchronization; stage 2, trains of 4–6 Hz spike-and-wave complexes and of 2–4 Hz slow waves; stage 3, ‘grand mal’ seizures. Isolated spikes and hypersynchronous high voltage spikes occurred in the hippocampus during stage 2 and stage 3, respectively. In the AT, EEG abnormalities were observed during stage 3 only. During the recovery period, cortical synchronization occurred associated with the presence of occasional synchronous spikes in both deep areas. After pretreatment with MS (2,12, and 25 mg/kg), stage 3 EEG seizures occurred with doses of BIC (0.12 or 0.18 mg/kg i.v.) that in the absence of MS produced only stage 1 or 2. In contrast, after pretreatment with a low dose of MS (0.5 mg/kg) the seizures induced by a maximally effective dose of BIC (0.25 mg/kg) were markedly attenuated. Thus, the effect of MS on BIC-induced seizure activity appears to be biphasic, potentiating seizures at doses at or above 2 mg/kg and inhibiting seizures at lower doses.

Effects of reflex stimuli on blood flow to the adrenal medulla.

The goal of this study was to examine changes in blood flow to the adrenal medulla during reflex stimuli that release catecholamines. Adrenal blood flow was measured with microspheres during exercise in conscious dogs and during bicuculline-induced seizures in anesthetized dogs. In awake dogs, blood flow to the adrenal cortex and adrenal medulla was 310 +/- 48 and 1,613 +/- 258 (SE) ml.min-1.100 g-1, respectively. Blood flow to the cortex and medulla was not affected by moderate exercise. Anesthesia (pentobarbital sodium) reduced blood flow to the adrenal cortex and medulla to 158 +/- 12 and 243 +/- 24 ml.min-1.100 g-1, respectively. In anesthetized dogs, seizures markedly increased blood flow to the adrenal medulla (282 +/- 68 to 1,257 +/- 128 ml.min-1.100 g-1), and this response was blocked by sectioning the greater splanchnic nerve. Vasodilator responses of the adrenal medulla to adenosine were not impaired by adrenal denervation. Blood flow to the adrenal cortex was not affected by seizures but increased in response to intravenous adenosine. These findings suggest that 1) anesthesia decreases adrenal blood flow, particularly to the adrenal medulla, and 2) reflex stimuli that release catecholamines can produce selective increases in blood flow to the adrenal medulla.

Cerebral superoxide anion generation during seizures in newborn pigs.

Cerebral superoxide anion generation during bicuculline-induced seizures was measured in newborn pigs. Using two closed cranial windows inserted over the parietal cortices, superoxide dismutase (SOD)-inhibitable nitroblue tetrazolium (NBT) reduction was determined during 20 min of seizure activity induced by bicuculline, 5 mg/kg i.v. A modest increase in SOD-inhibitable NBT reduction was observed in piglets subjected to bicuculline-induced seizure activity (2.4 +/- 0.6 pmol/mm2 in 20 min) when compared to control piglets (0.4 +/- 0.3 pmol/mm2 in 20 min). Pretreatment with indomethacin (5 mg/kg i.v.) reduced SOD-inhibitable NBT reduction during seizures to the control level (0.5 +/- 0.4 pmol/mm2 in 20 min). We conclude that small quantities of superoxide anion radical are produced by newborn pig brain during bicuculline-induced seizures and that cyclooxygenase metabolism of arachidonic acid appears to be a major source.

Trigeminovascular fibers increase blood flow in cortical gray matter by axon reflex-like mechanisms during acute severe hypertension or seizures.

Cerebral blood flow was measured and compared in 10 symmetrical brain regions following unilateral trigeminal ganglionectomy (n = 13), sham operation (n = 6), or trigeminal root section (rhizotomy) (n = 8) in cats. Multiple determinations were obtained in anesthetized and paralyzed animals using radiolabeled microspheres during (i) normocapnia-normotension, (ii) hypercapnia (5% CO2/95% room air), (iii) angiotensin-induced acute severe hypertension (190 greater than mean arterial blood pressure less than 210 mmHg), or (iv) bicuculline-induced seizures. Flow was symmetrical in all brain regions at rest and during increases induced by hypercapnia in the three groups. During severe hypertension or seizures, marked elevations developed bilaterally (approximately 93% and approximately 130%, respectively). In ganglionectomized animals, increases due to hypertension or seizures were attenuated by 28-32% on the denervated side within cortical gray matter regions corresponding to the anterior, middle, and posterior cerebral arteries. Flow was symmetrical within all brain regions in sham-operated animals and in the rhizotomy group, despite comparable increases in regional cerebral blood flow induced by angiotensin. Hence, the trigeminal nerve mediates blood flow adaptations during severe hypertension and seizures. Furthermore, since trigeminal cell bodies and peripheral axons are destroyed or degenerate following ganglionectomy but not following rhizotomy, local "axon reflex-like" mechanisms mediate these increases in cerebral blood flow.

Role of prostanoids in cerebrovascular responses during seizures in piglets

We examined the role of prostanoids to bicuculline-induced seizures in newborn pigs. Using the closed cranial window method and radioimmunoassay for prostanoids, we found that pial arterioles dilated during seizures; this dilation was associated with increased cortical periarachnoid cerebrospinal fluid (CSF) levels of prostaglandin E2, 6-ketoprostaglandin F1 alpha, prostaglandin F2 alpha, and thromboxane B2. Using radioactive microspheres, we found cerebral blood flow (CBF) increased dramatically during seizures in piglets. To examine the effects of pharmacological blockade of cyclooxygenase, we determined CBF in piglets pretreated with indomethacin (5 mg/kg) and in a corresponding group of piglets made hypothermic to be sure preseizure CBF values were similar. In the hypothermic group, CBF increased during seizure, but the increase in CBF during seizure in the indomethacin group was reduced dramatically in all brain areas. We conclude that 1) bicuculline-induced seizures result in pial arteriolar dilation and increased CBF; 2) CSF levels of prostanoids increase during seizures; and 3) pretreatment with indomethacin severely blunts increases in CBF during seizures.

Effects of progabide on bicuculline-induced epileptic seizures in developing rats.

The effects of progabide, a direct gamma-aminobutyric acid (GABA) receptor agonist, on bicuculline-induced seizures have been tested in developing rats, ages 7-28 days, to study the correlation between the antiepileptic effectiveness of this drug and the level of functional maturation of the GABAergic system. The incidence, latency of appearance, and behavioral characteristics of the epileptic manifestations, their evolution toward status epilepticus, and the percentage of recovery from status epilepticus have been evaluated in rats that had received a single injection (treatment) or three successive daily administrations (pretreatment) of progabide. The results have been compared with those obtained in a control group of animals in which only bicuculline had been injected. In rats ages 7-14 days the treatment appears to be substantially ineffective in protecting animals against bicuculline seizures and their consequences, probably because of the substantial immaturity of the GABAergic system at birth and during the first days of life. At this age, repetitive administrations of progabide cause a protective anticonvulsant action more remarkable than the single injection, particularly when using the higher doses of the substance. In 15-28-day-old rats, the treatment significantly reduces the lethality from status epilepticus but does not substantially modify the incidence of seizures, their latency of appearance, or their evolution toward status epilepticus. As in younger animals, in these rats also pretreatment is more effective than treatment against bicuculline seizures, whatever dose of progabide is used. At this age, therefore, the anticonvulsant properties of progabide appear to be more remarkable than in the previous age, probably because of a higher level of development of the GABAergic system, according to biochemical data on the GABAergic system ontogenesis.

Effects of bilateral injection of GABA into the substantia nigra on spontaneous behavior and measures of analgesia

Bilateral injection of gamma-aminobutyric acid (GABA, 10–300 μg) into the substantia nigra (pars reticulata) of rats produced stereotyped sniffing and had an analgesic-like effect on the hot-plate but not on the tail-flick test. These effects of GABA (30 μg) were suppressed by simultaneous administration of a sub-convulsant dose of bicuculline methiodide (100 ng). Significant increases in locomotion occurred when GABA (300 μg) was injected along with the inhibitor of GABA-transaminase, d, l/-gamma-vinyl-GABA (GVG; 5 μg) and the inhibitor of the uptake of GABA, 1-2,4-diaminobutyric acid (DABA; 5 μg). No other behavioral effects were observed following injection of GABA into the nigra, either alone or in combination with GVG and DABA. Bilateral injection of bicuculline (100–600 ng) into the nigra had strong convulsant actions. When injected simultaneously with bicuculline, GABA reduced bicuculline-induced seizures. These results are discussed in terms of their relevance to understanding the mechanisms that underlie the behavioral effects produced by injection of muscimol into the nigra.

Regional cerebral glucose utilization during vasopressin-induced barrel rotations or bicuculline-induced seizures in rats.

Regional cerebral metabolic rates of glucose (rCMRglu) were measured in conscious rats grouped according to three treatments: control, bicuculline (5.5 mg/kg s.c.), and intracerebroventricular arginine vasopressin (0.5 micrograms/5 microliter). Rats in the latter group were pre-exposed to the peptide 48 h earlier to render them 100% susceptible to a motor output termed barrel rotation (BR). Marked increases in rCMRglu occurred in all brain areas investigated after bicuculline. Increases after intracerebroventricular arginine vasopressin were smaller and occurred in fewer brain areas. Opposite to bicuculline, arginine vasopressin reduced rCMRglu in hippocampus and auditory cortex. The data confirm marked stimulation of rCMRglu during bicuculline-induced seizures and provide initial data for an endogenous peptide that causes BR. Generalized seizures and BR may share some neural substrates, but they produce distinct changes in rCMRglu. The rCMRglu changes are compatible with the interpretation that BR is initiated by brainstem/cerebellar areas that process visual-vestibular information, with subsequent involvement of higher brain structures.

In vivo 31P and in vitro 1H nuclear magnetic resonance study of hypoglycemia during neonatal seizure.

To examine the hypothesis that hypoglycemia has an adverse effect on brain energy state during seizure, neonatal dogs were subjected to bicuculline-induced seizure while hyperglycemic, normoglycemic, or hypoglycemic. Cerebral blood flow increased and remained elevated in all animals subjected to seizure, regardless of blood or brain glucose concentration. In vivo 31P nuclear magnetic resonance spectroscopy disclosed a small (10-20%) decrease in adenosine triphosphate levels and a greater (20-40%) decline in phosphocreatine levels in animals experiencing seizure, irrespective of whether they were hyper-, normo-, or hypoglycemic. In vitro analysis of brain extracts with 1H nuclear magnetic resonance spectroscopy disclosed a significant elevation of lactate in all seizing animals. There were differences in brain alanine, glycine, and beta-hydroxybutyrate levels between the hyperglycemia-seizure and hypoglycemia-seizure groups. Alternate substrates such as lactate, fatty acids, or amino acids may be used when neonatal seizure is complicated by hypoglycemia, thereby preventing further deterioration of brain metabolic state.

Interstitial fluid adenosine and sagittal sinus blood flow during bicuculline-seizures in newborn piglets.

Changes of interstitial fluid adenosine concentrations and effects of O2 supply on interstitial fluid adenosine were studied by the brain dialysis technique in the frontal cortex of newborn piglets subjected to bicuculline-induced seizures. The O2 supply was changed globally by changing MABP and locally by varying PO2 in the artificial CSF perfusing the dialysis cannula. Sagittal sinus blood flow (SSBF), cerebrovascular resistance (CVR), and CMRO2 were also examined in the same animals. Seizures increased interstitial fluid adenosine 7.9-fold (p less than 0.05) when ictal MABP was maintained at preictal level and perfusate PO2 was 24 mm Hg (group 1, n = 6). Interstitial fluid adenosine increased 11.8-fold (p less than 0.05) during seizures associated with moderate systemic hypotension and the low perfusate PO2 (group 2, n = 6). By contrast, seizures increased interstitial fluid adenosine three-fold (p less than 0.05) when perfusate PO2 was increased to 182 mm Hg and ictal MABP was maintained at preictal level (group 3, n = 8). When ictal MABP was elevated from the preictal level and the perfusate was rich in oxygen, seizures failed to increase interstitial fluid adenosine (group 4, n = 7). In groups 1 and 3, the increase in interstitial fluid adenosine during seizures was associated with significant increases in SSBF and CMRO2, as well as significant decreases in CVR. These data suggest that the increase in O2 supply during seizures in piglets did not match completely the increase in O2 demand and resulted in enhanced release of adenosine into the interstitial space.

Effects of bicuculline-induced seizures on benzodiazepine and adenosine receptors in developing rat brain

The effects of seizures induced by an acute administration of bicuculline have been investigated on the central benzodiazepine and adenosine receptors in developing rats and in adults. Generalized seizures rapidly increased the total number of both benzodiazepine binding sites and adenosine A 1 receptors, without changes in receptor affinity (K D). It was concluded that such a phenomenon may facilitate the anticonvulsant action of benzodiazepine and adenosine via receptor binding and that it could be an adaptative process to protect subjects against recurrent seizures, especially in newborns.

Differential sensitivity to bicuculline in three inbred mouse strains

We examined the inbred mouse strains DBA/2Ibg, C57BL/6Ibg and C3H/2Ibg for differences in susceptibility to bicuculline-induced seizures, as well as to bicuculline-induced epileptiform activity recorded in the CA1 pyramidal cell layer of hippoeampal slices. For susceptibility to seizure onset the strain rank order was (most to least susceptible):C3H=DBA>C57. The rank order for sensitivity to effects of bicuculline on tonic seizure latency and on hippoeampal epileptiform activity were identical: C3H=DBA>C57. It is suggested that mechanisms underlying the development of bicuculline-induced epileptiform events in the hippoeampal slice may be similar to those involved in the development of tonic seizures measured in vivo.

Cerebellar metabolism of phosphatidylethanolamine and its water-soluble precursors during bicuculline-induced convulsive seizures.

The incorporation of labeled ethanolamine into phosphatidylethanolamine (PE) and its water-soluble precursors, phosphoethanolamine and CDP-ethanolamine, is measured in rat cerebella during the course of bicuculline-induced convulsive seizures. The labeling of CDP-ethanolamine and phosphoethanolamine diminishes 6 min after the administration of both bicuculline and radioactive ethanolamine whereas that of PE is unaffected in these conditions. Time is very important to this effect; indeed, no differences of the labeling of PE water-soluble precursors can be found in rat cerebellum of normal and convulsing animals 12 min after the administration. The cerebellar pool of CDP-ethanolamine doubles after 6 min of convulsions, which means that unlabeled CDP-ethanolamine forms from a non-radioactive source, such as lipid, may be through the reversal of the ethanolamine phosphotransferase reaction. This effect disappears 12 min after the injection of the convulsant.

The effect of hypotension on brain energy state during prolonged neonatal seizure.

Prolonged seizures in the human neonate may be complicated by systemic hypotension. To examine the effect of systemic hypotension on brain metabolic state during seizure, neonatal dogs were made hypotensive (by exsanguination) during bicuculline-induced seizure. Measurement of regional cerebral blood flow showed that moderate hypotension did not impair cerebral perfusion during seizure. Measurement of brain energy state with in vivo 31P nuclear magnetic resonance spectroscopy disclosed a similar pattern of alteration of high energy phosphates in animals subjected to seizure or to the combination of seizure and hypotension. The additional metabolic stress imposed by moderate hypotension during seizure in the neonatal dog appears to be slight.

Time course for development of anticonvulsant tolerance and GABAergic subsensitivity after chronic diazepam

The time courses for development of neuronal and behavioral tolerance to diazepam (DZ) were estimated in rats continuously exposed to low levels of DZ for 3, 7, 14 or 21 days. Microiontophoretic sensitivity of dorsal raphe neurons to γ-aminobutyric acid (GABA) was initially facilitated after short-term exposure to DZ released from implanted capsules for up to 3 days but returned to control levels by 7 days postimplantation and continued to decrease thereafter. GABAergic sensitivity remained depressed for a minimum of 5 days following removal of DZ capsules. To obtain a behavioral measure of tolerance, the anticonvulsant activity of DZ against bicuculline-induced seizures was also assessed. Rats studied 3 days after capsule implantation showed a significant elevation in seizure threshold. Seizure liability returned to control levels ca. 7 days after chronic treatment was initiated. These results indicate that tolerance to anticonvulsant efficacy against bicuculline seizures are temporally related to the onset of reduced GABA sensitivity on dorsal raphe neurons during prolonged exposure to DZ.

ChemInform Abstract: Synthesis and Activity of 6‐Aryl‐3‐(hydroxypolymethyleneamino)pyridazines in Animal Models of Epilepsy.

AbstractAusgehend von den Aryl‐substituierten Chlorpyridazinen (I) und dem cyclischen Amin (II) werden die Verbindungen (III) synthetisiert.

Preferential blood flow to brainstem during generalized seizures in the newborn marmoset monkey

The effect of generalized seizures on local cerebral blood flow was studied autoradiographically in 21 immature marmoset monkeys, using either [ 123I]- or [ 131I]isopropyliodoamphetamine. Generalized convulsions were induced in ketamine-anesthetized and awake monkeys with bicuculline and continued for 4–59 min. During convulsions in marmosets less than 3 weeks of age, there was a striking rearrangement of blood flow in favor of the brainstem pontomedullary region. The ratios of blood flow in pons-medulla to blood flow in cerebral cortex, putamen, ventroposterior thalamic nuclei, lateral geniculate nuclei, cerebellum and hemispheric white matter increased1/12 to 2 timescompared to controls. In seizure animals 4–8 weeks of age, the redistribution of blood flow to brainstem did not occur. Although metabolic acidosis developed after 30 min of bicuculline-induced seizures, mean arterial blood pressure, temperature, arterial pO 2 and pCO 2 did not differ significantly from controls, indicating that hypoxemia, hypercapnia and hypotension cannot explain the altered cerebral blood flow pattern. The redistribution phenomenon could be explained by more hronounced vasodilatation in brainstem than many other brain regions during generalized seizures in newborn monkeys. Lack of significant vasodilatation in forebrain structures such as cerebral cortex could contribute to neuronal damage by limiting substrate supply at a time of increased metabolic activity.

Combined 1H and 31P nuclear magnetic resonance spectroscopic studies of bicuculline-induced seizures in vivo.

Using a 1.89-Tesla spectrometer, 1H and 31P nuclear magnetic resonance spectra were acquired from the brains of paralyzed rabbits ventilated with 30% oxygen in nitrous oxide. Intracellular pH and changes in lactate concentration in the cerebrum were monitored by nuclear magnetic resonance methods during and after bicuculline-induced seizures, together with the electroencephalogram, heart rate, and arterial blood pressure. During seizures lasting more than an hour, cerebral intracellular pH became acidic, the cerebral lactate level rose rapidly, and both changes persisted as long as 2 hours without signs of recovery. After less prolonged seizures, lactate elevations were no less persistent, despite nearly complete recovery of intracellular pH and the electroencephalogram.

Enhanced inositide turnover in brain during bicuculline-induced status epilepticus

Because brain inositides are enriched in the 1-stearoyl-2-arachidonoyl species, they form a likely source for the tetraenoic free fatty acids (FFA) and diacylglycerols (DG) that are accumulated during seizures. To study inositide turnover during bicuculline-induced seizures, rats were injected intraventricularly and bilaterally with 10–20 μCi 32P, mechanically ventilated and sacrificed by 6.5 KW head-focused microwave irradiation. Seizure activity was recorded by electroencephalography. Bicuculline-induced seizure activity resulted in: a) almost 50% increase in 32P labeling of phosphatidic acid (PA); phosphatidylinositol (PI) and phosphatidylinositol 4,5-bisphosphate (PIP 2) also increased (24% and 36%, respectively); b) no change in other lipids; and c) water-soluble phosphodiesteratic degradation products, analyzed by high voltage paper electrophoresis, increased 24% in the amount of radiotracer recovered as inositol 1,4-bisphosphate (IP 2) and by 44% in the amount recovered as inositol 1,4,5-trisphosphate (IP 3). These data indicate that during experimental status epilepticus the cerebral inositide cycle is accelerated: PIP 2 → ( (IP 3 → IP 2 → IP → I) + DG → PA → PI → PIP → PIP 2.

Glycerol as an indicator of lipid degradation in bicuculline-induced seizures and experimental cerebral ischemia.

Glycerol, the end product of phospholipid degradation, was measured in cat brains under pathophysiological conditions known to cause activation of lipolysis, namely, bicuculline-induced seizures, permanent focal cerebral ischemia (2 hr of middle cerebral artery occlusion), and global cerebral ischemia (15 min of complete cerebral ischemia with or without 2 hr of recirculation). In addition, ATP and lactate were measured in order to correlate the activation of lipid degradation with disturbances in the energy-producing metabolism. A highly significant increase in the tissue glycerol content was observed after 1 hr of bicuculline-induced seizures (from 0.29 +/- 0.07 mumol/g in control animals to 1.30 +/- 0.06 mumol/g in seizure animals; P less than 0.001) or after 15 min of complete cerebral ischemia (from 0.29 +/- 0.07 to 1.17 +/- 0.14 mumol/g; P less than 0.01). Furthermore, a close correlation was found between the increase in glycerol and the increase in lactate or decrease in ATP after permanent focal ischemia. In contrast, following recirculation after complete cerebral ischemia, restoration of the energy pool did not lead to a reduction of the glycerol formed during ischemia. It is concluded that glycerol is a useful indicator of lipid degradation under pathological conditions. Since glycerol formed during vascular occlusion is trapped in brain cells, presumably owing to low glycerol kinase activity, it can be used as a stable postischemic indicator of ischemia-induced lipid degradation.