Glycine potentiates the anticonvulsant action of diazepam and phenobarbital in kindled amygdaloid seizures of rats

The effect of glycine on the anticonvulsant activity of diazepam and phenobarbital in fully developed kindled amygdaloid seizures in rats was determined. Glycine alone had no significant effect on the seizure response, either when administered orally 1 hr prior to the seizure test or when given chronically in a 0.5 M solution as the source of water.

Administration of glycine (10 mmol/kg, oral) together with diazepam produced a significant reduction in both cortical epileptiform afterdischarge and the severity of seizures at doses of diazepam which had no significant effect on the seizures when administered alone. Glycine potentiated the effects of phenobarbital on the cortical afterdischarge but not the severity of the seizures. The observed potentiation of the effects of diazepam and phenobarbital suggests a glycinergic mechanism in the anticonvulsant action of these drugs which may be mediated in part by the inhibitory γ-aminobutyric acid (GABA) systems.     

The anticonvulsant effects of diazepam and phenobarbital in prekindled and kindled seizures in rats

The effects of various doses of diazepam (0.5–4 mg/kg) and phenobarbital (7.5–60 mg/kg) were determined on prekindled and kindled amygdaloid seizures in the same rats. Diazepam was ineffective against the prekindled focal seizures, but demonstrated profound and statistically significant control of the kindled seizures. In the kindled state, diazepam reduced the afterdischarge duration and seizure rank score to prekindled levels. Only the largest sedating dose of phenobarbital produced a reduction of both prekindled afterdischarge duration and seizure rank score. Against the kindled seizure, phenobarbital showed a marked and statistically significant increase in effectiveness in all but the smallest dose tested. The afterdischarge duration of kindled seizures was reduced to prekindled levels by 15–60 mg/kg of phenobarbital, while seizure rank score was reduced to prekindled levels by 30 and 60 mg/kg phenobarbital. The effects of two doses of diazepam (0.5 and 2.5 mg/kg) and phenobarbital (7.5 and 30 mg/kg) were tested against prekindled and kindled pentylenetetrazol (PTZ)-induced seizures. Preliminary work with 3 doses of pentylenetetrazol (30, 40 and 60 mg/kg) demonstrated that repeated doses of 30 mg/kg readily kindled seizures without the significant mortality seen with larger doses. Both diazepam and phenobarbital were less effective against seizures kindled with 30 mg/kg pentylenetetrazol compared to prekindled seizures. The comparative lack of effect that was seen with diazepam and phenobarbital against the pentylenetetrazol kindled seizure at doses associated with control of the kindled amygdaloid seizure may reflect an underlying difference in the pathogenesis of kindling between these seizure models. Further, the lack of suppression of the prekindled amygdaloid afterdischarge duration by large doses of diazepam, in contrast to large doses of phenobarbital, may also reflect differences between the mechanisms of action of these two drugs. This paradigm provides a model for testing the effectiveness of anticonvulsants during the progressive development of various epileptogenic seizures.     

Comparison of the anticonvulsant effects of two novel GABA uptake inhibitors and diazepam in amygdaloid kindled rats

Summary Two novel, specific inhibitors of GABA uptake, namely SKF 89976-A (N-[4,4-diphenyl-3-butenyl]-nipecotic acid) and SKF 100330-A (N-[4,4-diphenyl-3-butenyl]-guvacine) were tested for anticonvulsant effects in amygdaloid kindled female rats. The anticonvulsant effectiveness of the compounds was compared with that of diazepam. SKF 89976-A and SKF 100330-A produced dosedependent anticonvulsant effects on all seizure parameters measured in fully kindled rats, i.e. they inhibited seizure severity, increased seizure latency, and decreased the duration of motor seizures and EEG afterdischarges. ED 50s for inhibition of seizure severity were 4.6 and 15.1 mg/kg (0.014 and 0.045 mmol/kg) i.p. for SKF 100330-A and SKF 89976-A, respectively. For comparison, the ED 50 of diazepam was 1.9 mg/kg (0.0067 mmol/kg) i.p. Observation of behaviour indicated that the novel GABA uptake blockers exerted no side-effects in anticonvulsant doses, whereas diazepam produced sedative effects at all active dosage levels. The data demonstrate that SKF 100330-A and SKF 89976-A are potent, non-sedative anticonvulsant drugs in the kindling model of epilepsy, and these compounds thus may deserve interest as potential antiepileptic drugs with a very selective mechanism of action.     

The anticonvulsant effects of diazepam and phenobarbital in prek1ndled and kindled cortical seizures

The effects of various doses of diazepam (0.5–4 mg/kg) and phenobarbital (15–60 mg/kg) were determined on prekindled (focal) and kindled (generalized) cortical seizures in the same rats. Only high sedating doses of diazepam or phenobarbital reduced the elicited afterdischarge duration (ADD) and behavioral response in the prekindled focal cortical seizure. Against the kindled seizure, both diazepam and phenobarbital showed a marked and statistically significant increase in effectiveness in all but the smallest doses tested. The ADD of the kindled cortical seizures was reduced to prekindled lengths by diazepam (1–4 mg/kg) or phenobarbital (30–60 mg/kg). The increased anticonvulsant effectiveness found in this study is similar to previous findings with diazepam and phenobarbital against prekindled and kindled amygdaloid seizures, but stands in contrast to findings with prekindled and kindled pentylenetetrazol seizures.     

The effects of various anesthetics on amygdaloid kindled seizures

The effects of various doses of cataleptic anesthetics, gamma-butyrolactone (GBL), phencyclidine (PCP), and ketamine (KET), and the depressant anesthetics, pentobarbital (PB) and chloral hydrate (CH), on amygdaloid kindled seizures were tested in the rat. The seizure activity was monitored by behavioral observation and EEG recording. Anesthetic doses of the cataleptic anesthetics with the exception of KET had minimal effects on the afterdischarge duration (AD) and behavioral ranking (BR) of the elicited seizures. On the other hand, they were more inhibitory to the AD and BR than was the convulsant pentylenetetrazol (PTZ). The only cataleptic that induced spontaneous seizure activity at anesthetic doses was PCP, although KET induced epileptoid activity at supranesthetic doses. Ketamine, PB, and CH completely inhibited elicited seizure activity at anesthetic doses. In addition, rats were kindled by repetitive electrical stimulation during GBL-induced anesthesia or catalepsy. Although both these GBL groups averaged more stimulations to reach generalized seizures than the saline controls, GBL did not block the kindling process.     

The effects of pentylenetetrazol, bicuculline and strychnine on the development of kindled seizures

Modification of the rate of acquisition of the kindled amygdaloid seizure by the convulsants pentylenetetrazol, bicuculline and strychnine was studied. Injections of saline, 25 mg/kg of pentylenetetrazol, 2 mg/kg of bicuculline or 1 mg/kg of strychnine were given 15 min prior to the daily electrical stimulation of the amygdala. The drug doses selected were capable of producing some behavioral and electrical epileptoid activity prior to stimulation without inducing generalized seizures. To determine whether pentylenetetrazol or bicuculline accelerated the rate of development of the kindled amygdaloid seizure or merely augmented the expression of each seizure, a crossover design was implemented. The crossover studies involved switching animals during the acquisition phase (between stimulations 3–6) from prestimulation saline to drug or drug to saline injections. It was found that pentylenetetrazol markedly augmented the expression of seizures during kindling development but the results of the crossover studies showed a less dramatic acceleration in the actual rate of the development of the fully generalized kindled amygdaloid seizure. The bicuculline-treated animals showed little augmentation in the expression of seizures during the kindling acquisition phase and in the actual rate of development of the kindled amygdaloid seizure. The strychnine treated animals showed no augmentation in expression of the seizures nor in the rate of development. The effects of prestimulation injections of bicuculline (1, 2 and 3 mg/kg) and strychnine (0.5, 1 and 2 mg/kg) on fully developed kindled amygdaloid seizures were also evaluated. Pretreatment with bicuculline minimally increased seizure afterdischarge duration at the highest dose. When fully kindled animals were pretreated with strychnine, a paradoxical decrease in afterdischarge length and an increase in severity (tonic hindlimb extension) was seen with the largest dose tested. This study emphasizes the potential importance of crossover studies in evaluating pharmacological manipulations of the rate of acquisition of the kindled seizure.     

甘草多糖对戊四氮点燃癫痫模型大鼠的影响

目的 研究甘草多糖对戊四氮点燃癫痫大鼠的影响及机制.方法 通过戊四氮点燃构建癫痫大鼠模型,另取10只大鼠为对照组注射等量生理盐水.将癫痫大鼠随机分为模型组(n=20)及实验组(n=20).实验组大鼠于戊四氮注射前1 h腹腔注射甘草多糖溶液(50 mg·kg-1),对照组和模型组注射等量生理盐水,连续干预30 d.以苏木...     

The long-term effects of diazepam and pentylenetetrazol on the potentiated startle response

Stress desensitization is observed as a decrease in the disruptive effects of a stressor on behavior when the organism is repeatedly exposed to the stressor. For instrumental behavior, stress desensitization was also reported for rats preexposed to anxiogenic drugs; stress sensitization was reported for rats preexposed to an anxiolytic compound. The present study investigated whether similar effects are found in a noninstrumental task situation. First, rats received 12 daily injections of pentylenetetrazol (PTZ, 20 mg/kg, IP), diazepam (DZP, 5 mg/kg, IP) or saline. After each injection the effect of the drugs on the acoustic startle reflex was measured. No drugs were given during the remainder of the experiment. Eight days later rats were given 5 days of Pavlovian fear conditioning to establish a light as a shock signal. During the next 3 days, potentiation of the startle response by the light was measured. None of the drug treatments had an effect on startle potentiation, indicating that stress sensitivity is not affected by previous administration of PTZ and DZP in a noninstrumental task. An explanation for the different effects found for instrumental and noninstrumental tasks is suggested.     

Tolerance to the anticonvulsant effects of carbamazepine, diazepam, and sodium valproate in kindled rats.

We assessed the development of tolerance to the anticonvulsant effects of carbamazepine (CBZ), diazepam (DZP), and sodium valproate (VPA) on convulsions elicited by amygdala stimulation in kindled rats in three similar experiments. In each experiment, amygdala-kindled rats were assigned to a drug group or to a corresponding vehicle control group. The rats in the three drug groups received a total of 10 bidaily (one every 48 h) IP injections of CBZ (70 mg/kg), DZP (2 mg/kg) or VPA (250 mg/kg) at a dose that initially blocked the forelimb clonus elicited by an amygdala stimulation (400 microA, 60 Hz, 1 s) administered 1 h after the injection. The rats in the three vehicle control groups were similarly treated except that they received injections of the saline vehicle. The drug tolerance test occurred 48 h after the final tolerance-development trial; the rats from each drug group and the corresponding vehicle control group received an injection of the appropriate drug followed 1 h later by the administration of a convulsive stimulation. The drug tolerance test revealed almost total tolerance in each of the three drug groups but no tolerance in any of the three vehicle control groups. Such large tolerance effects are inconsistent with the less dramatic effects reported in previous studies; possible reasons for this inconsistency were considered.     

Behavioral comparison of pentylenetetrazol,clonidine, chlordiazepoxide and diazepam in infant rats

The effects of various doses of pentylenetetrazol, clonidine, chlordiazepoxide and diazepam on limb and head movement and behavioral seizure signs were examined in 4-, 8- and 16-day old rats tested at ambient temperatures of either 25 or 35°C. All 4 drugs produced intense behavioral activation at the 2 younger ages but there were marked differences among in the effects of test temperature on this activation and in the relationship between age and their activating effect. A “paradoxical” and intense behavioral energization was observed after the administration of either of the 2 benzodiazepines at 4, 8 but not 16 days, particularly at the lower test temperature. Clonidine and pentylenetetrazol were activating at all 3 ages but while clonidine had greater effect at the low test temperature, the opposite was the case after pentylenetetrazol. The effects of the benzodiazepines and clonidine were clearly distinct from those of pentylenetetrazol and this was the only drug to substantially elicit seizure signs. It is uncertain whether or not the benzodiazepines cause brain seizures in young animals. If so, then their behavioral manifestation is clearly different from that observed after pentylenetetrazol.     

Anticonvulsant drugs and their antagonism of kindled amygdaloid seizures in rats

The kindling phenomenon has become a useful animal model for studying the processes in the central nervous system involved in one type of epileptiform seizure response. A systematic, multipledose evaluation of 20 drugs known to have anticonvulsant properties and two new compounds (WB-CPI and SHK-I-11) is reported using a standard protocol with rats kindled by amygdaloid stimulation. Drugs which have a wide anticonvulsant spectrum of activity against various animal models of epilepsy were the most effective. The barbiturates, the benzodiazepines, a piperazine, an acetate derivative and WB-CPI (a significantly modified phenacylurea), were found to be the most effective in attenuating the induced seizures. Kindled amygdaloid seizures in the rat can be characterized as a model useful for the detection of a wide variety of anticonvulsants likely to be effective in many types of experimental seizures. The compound WB-CPI merits further neuropharmacological and psychopharmacological testing due to its activity in rats.     

Contingent tolerance to the anticonvulsant effects of carbamazepine, diazepam, and sodium valproate in kindled rats.

The effect of convulsive stimulation during periods of drug exposure on the development of tolerance to the anticonvulsant effects of carbamazepine (CBZ), diazepam (DZP), or sodium valproate (VPA) was studied in three similar experiments. In each experiment, amygdala-kindled rats were assigned to one of three groups: one group received a drug injection (CBZ, 70 mg/kg, IP; DZP, 2 mg/kg, IP; VPA, 250 mg/kg, gavage) 1 h before each of a series of 10 bidaily (one every 48 h) convulsive stimulations, a second group received the same dose of the drug 1 h after each of the 10 stimulations, and a third group served as a vehicle control. The drug tolerance test occurred in each experiment 48 h after the 10th tolerance-development trial; every rat received the appropriate dose of CBZ, DZP, or VPA 1 h before being stimulated. In each experiment, only the rats from the drug-before-stimulation group displayed a significant amount of tolerance to the drug's anticonvulsant effect. Thus the development of tolerance to the anticonvulsant effects of CBZ, DZP, and VPA was not an inevitable consequence of drug exposure; the development of tolerance was contingent upon the occurrence of convulsive stimulation during the periods of drug exposure. These results support the idea that functional drug tolerance is an adaptation to a drug's effects on ongoing patterns of neural activity, rather than to drug exposure per se.     

Effects of carnosine on amygdaloid kindled seizures in Sprague - Dawley rats

The effects of carnosine （ -alanyl -L -histidine） on amygdaloid kindled seizures were investigated in rats. Intraperitoneal injection of carnosine （500, 1000, 1500 mg/kg, i. p. ） significantly decreased seizure stage, afterdischarge duration and generalized seizure duration, and significantly prolonged generalized seizure latency of amygdaloid kindled seizures, in a dose - dependent, and time - related manner. The protective effect of carnosine （ 1500 mg/kg） was completely antagonized by histamine H1 -antagonists pyrilamine （2, 5 mg/kg, i. p. ） and diphenhydramine （5, 10 mg/kg, i. p. ）,     

The long-term effects of diazepam and pentylenetetrazol on behavioral sensitivity to a stressor

Rats were trained to bar press for sucrose reinforcement following daily injections of 20 mg/kg pentylenetetrazol (PTZ), 5 mg/kg diazepam (DZ), or saline. At the end of 12 days of this training, all injections were suspended for the remainder of the experiment. Five days later, the rats were given 10 days of Pavlovian fear conditioning (two trials per day) to establish a light as a shock signal. Next, the rats were returned to the bar press situation to test the capacity of the light to suppress responding. Rats previously treated with DZ showed stronger conditioned fear of the light than did rats originally trained following injections of either PTZ or saline. In contrast, bar pressing by PTZ-treated rats was less suppressed by light than was control performance. The results indicate that modification of the behavioral effects of environmental stressors can be a long-term consequence of drug treatments. DZ treatments had the long-term effect of increasing behavioral disruption by a stressor, while treatment with PTZ reduced the stressor's negative behavioral impact. These findings appear compatible with the idea that behavioral sensitivity to stressors is dependent, in part, on learning about the stimulus properties of internal states.     

Effect of cocaine on afterdischarge threshold in previously kindled rats

Previous experiments have yielded conflicting reports on the effect of cocaine on the afterdischarge (AD) threshold for electrical stimulation. The present study was designed to determine if differences in the type of stimulation used could account for these discrepancies. Male Long-Evans rats which had been previously kindled by stimulation of the olfactory bulb were used to determine the AD threshold of the olfactory bulb following the intraperitoneal injection of saline or 20 mg/kg cocaine hydrochloride. AD's were elicited by trains of square-wave pulses which varied in frequency and train duration. Cocaine significantly increased the amount of current required to produce AD's using stimulus trains with frequencies of 30–100 pulses/sec, while evidence of a decrease in AD threshold by cocaine was found at a frequency of 20 pulses/sec. The results suggest that cocaine has opposite effects on AD threshold at high and low frequencies of stimulation.     

戊四唑点燃大鼠海马、杏仁核一氧化氮合酶阳性神经元的变化

目的：探讨一氧化氮（NO）在戊四唑（PTZ）点燃发病机理中的作用,方法：每天注射PTZ建立大鼠点燃模型,运用组织化学方法观察海马,杏仁核一氧化氮合酶（NOS）阳性神经元变化,结果：点燃后海马,杏仁核内NOS阳性神经元明显增多,结论：NO促进了戊四唑点燃的形成。     

Combined effects of epileptic seizure and phenobarbital induced overexpression of P-glycoprotein in brain of chemically kindled rats

Background and purpose:

The multidrug resistance of epilepsy may result from the overexpression of P-glycoprotein, but the mechanisms are unclear. We investigated whether the overexpression of P-glycoprotein in the brains of subjects with pharmacoresistant epilepsy resulted from both drug effects and seizure activity.

Experimental approach:

Kindled rats were developed by injecting a subconvulsive dose of pentylenetetrazole (33 mg·kg⁻¹·day⁻¹, i.p.) for 28 days. Groups were then treated with an oral dose of phenobarbital (45 mg·kg⁻¹·day⁻¹) for 40 days. In accord with behavioural observations, P-glycoprotein activity in brain was assessed using brain-to-plasma concentration ratios of rhodamine 123. P-glycoprotein levels in the brain regions were further evaluated using RT-PCR and Western blot analysis. The distribution of phenobarbital in the brain was assessed by measuring phenobarbital concentrations 1 h following its oral administration.

Key results:

The kindling significantly increased P-glycoprotein activity and expression. Good associations were found among P-glycoprotein activity, expression and phenobarbital concentration in the hippocampus. Short-term treatment with phenobarbital showed good anti-epileptic effect; the maximum effect occurred on day 14 when overexpression of P-glycoprotein was reversed. Continuous treatment with phenobarbital had a gradually reduced anti-epileptic effect and on day 40, phenobarbital exhibited no anti-epileptic effect; this was accompanied by both a re-enhancement of P-glycoprotein expression and decreased phenobarbital concentration in the hippocampus. P-glycoprotein function and expression were also increased in age-matched normal rats treated with phenobarbital.

Conclusions and implications:

The overexpression of P-glycoprotein in the brain of subjects with pharmacoresistant epilepsy is due to a combination of drug effects and epileptic seizures.     

Tolerance to the anticonvulsant effects of phenobarbital, trimethadione, and clonazepam in kindled rats: cross tolerance to carbamazepine.

The kindled-convulsion model was used to assess the development of tolerance and cross tolerance to the anticonvulsant effects of antiepileptic drugs. In Experiment 1, tolerance developed to the anticonvulsant effects of bidaily (one every 48 h) IP injections of phenobarbital, trimethadione, and clonazepam on convulsions elicited 1 h after each injection in kindled rats by amygdala stimulation. In Experiment 2, kindled rats that were tolerant to the anticonvulsant effects of phenobarbital, trimethadione, or clonazepam received bidaily IP injections of carbamazepine, each followed 1 h later by a convulsive amygdala stimulation. There was a statistically significant transfer of tolerance from phenobarbital to carbamazepine, but not from either trimethadione or clonazepam to carbamazepine. Apparently, tolerance to anticonvulsant drugs is most likely to transfer between drugs that are effective against similar kinds of clinical and experimental seizures and have similar putative mechanisms of action.     

Physical dependence on morphine, phenobarbital and diazepam in rats by drug-admixed food ingestion.

To produce physical dependence on morphine, phenobarbital and diazepam in rats, these drugs were mixed with powder form of rat food in concentrations of 0.5 mg/g, 1 mg/g and 2 mg/g of food. One group of rats (the lower dose group) was continuously exposed for 1 week to two morphine-admixed foods with morphine to food ratios of 0.5 mg/g and 1 mg/g in a cage. The other group (the higher dose group) could choose between two morphine-admixed foods with morphine to food ratios of 1 mg/g and 2 mg/g. After 1 week, morphine-admixed foods were replaced with morphine free food for 2 days. Both groups of rats showed greatly reduced body weight and food intake after the first 24-48 hr withdrawal. The body weight decrease was greater for rats in the higher dose group. Control groups of morphine dependent rats were kept on the morphine added food diets and showed the same body weight increase as well as normal control rats during the course of these experiments. Physical dependence on phenobarbital and diazepam was produced using the same dosage schedules as with morphine. Both the lower and higher dose groups showed significant decrease in body weight due to withdrawal after 1 week of drug-food exposure. Levallorphan (0.5, 1, 3 and 5 mg/kg, s.c.) administered to morphine dependent rats had dose-dependent effects on the intensity of abstinence symptoms (e.g., diarrhea, piloerection and wet shakes phenomena), maximal decrease in body weight and duration of decreased body weight. Cross-physical dependence between phenobarbital and diazepam was demonstrated by this method.     

Behavioral analysis of the effects of 15 anticonvulsants in the amygdaloid kindled rat

One hundred-four Wistar rats were implanted with a bipolar electrode aimed at the right basolateral amygdala, and stimulated bipolarly twice daily (interval between stimulations 30 min) with a 150 A, 4-s train of biphasic square-wave 1-ms pulses (100 pulses/s). Seventy-four animals developed a stimulation-induced clonic convulsion and, after further testing, 41 animals satisfied the stability criteria for entrance into the drug study. These criteria were three convulsions in response to three stimulations on the tenth stimulation day, and a convulsion after each stimulation for a period of 1 week on the drug-testing schedule. In the standardized test system, kindled rats were given two control stimulations; drug was then administered IP and, 30 min and 24 h later, further stimulation was given. After each stimulation the presence or absence of ipsilateral turning, closing of the ipsilateral eye, rearing up, falling over, facial clonus, forepaw clonus, hind paw clonus, hind paw tonus, and wet-shaking was assessed and the duration of forepaw clonus was measured. ID₅₀ values (mg/kg) for inhibition of forepaw clonus 30 min after injection are: Clonazepam, 0.28; diazepam, 0.28; chlordiazepoxide, 0.86; carbamazepine, 5.0; meprobamate, 11.2; mesuximide, 17.5; depamide, 22.4; diphenylhydantoin, 31.5; phenacemide, 31.6; flunarizine, 42.8; mephenytoin, 64.7; valproate sodium, 64.7; acetazolamide, 80.0; trimethadione, 120.0; and primidone, 142.2. The ED₅₀ doses for inhibition of the other seizure components were significantly correlated (P<0.01) with their ID₅₀ value for forepaw clonus. These results suggest that the kindled rat is an appropriate and sensitive model with which to assess anticonvulsant drugs.Part of these results were presented at the Epilepsy International Symposium, Vancouver, Canada, September 10–14, 1978 (Ashton and Wauquier, 1978)