Glutamic acid decarboxylase inhibition and ultrastructural changes by the convulsant drug allylglycine

doi:10.1016/0006-2952(69)90018-5

Biochemical Pharmacology

Volume 18, Issue 1, January 1969, Pages 137-142, IN9-IN10, 143

https://doi.org/10.1016/0006-2952(69)90018-5 Get rights and content

Abstract

The effects in vivo and _vitro of the convulsant drug allylglycine on the activity of glutamic acid decarboxylase (GAD), aminobutyrate aminotransferase, glutamine synthetase and aspartate- and alanine-aminotransferases of the rat cerebral cortex were studied. The most significant result was the finding of an inhibition of GAD, during the period of convulsion, which was even greater by addition in vitro of the drug. This inhibition is not by way of the cofactor, pyridoxal phosphate. Preincubation of the homogenate with allylglycine enhanced the inhibition, while preincubation in buffer-substrate produced a protective effect.

The inhibition of GAD was correlated with a decrease of 40 per cent in the concentration of γ-aminobutyric acid in the cerebral cortex. In the convulsant rat, ultra-structural alterations of some nerve endings of the cerebral cortex were observed. After cell fractionation, such altered nerve endings were preferentially found in the GAD-rich (nonaminergic) fraction of isolated nerve endings. The possible mechanism of the convulsions induced by allyglycine is discussed and a specific effect on GAD-rich inhibitory nerve endings is postulated.

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Fit for purpose application of currently existing animal models in the discovery of novel epilepsy therapies
2016, Epilepsy Research
Citation Excerpt :
Recently, Leclercq et al. (2015) studied the pharmacology of seizures induced by allylglycine in mice and larval zebrafish and proposed the use of this convulsant in zebrafish as a convenient and high-thoughput model of treatment-resistant seizures (Table 2). Allylglycine is an inhibitor of the GABA-synthesizing enzyme glutamic acid decarboxylase (GAD) and long known to cause seizures via decreasing synaptic GABA levels (Alberici et al., 1969). Experiments performed in zebrafish larvae revealed behavioral ASD activity profiles against allylglycine-induced seizures highly analogous to those obtained in mice (Leclercq et al., 2015).
Animal seizure and epilepsy models continue to play an important role in the early discovery of new therapies for the symptomatic treatment of epilepsy. Since 1937, with the discovery of phenytoin, almost all anti-seizure drugs (ASDs) have been identified by their effects in animal models, and millions of patients world-wide have benefited from the successful translation of animal data into the clinic. However, several unmet clinical needs remain, including resistance to ASDs in about 30% of patients with epilepsy, adverse effects of ASDs that can reduce quality of life, and the lack of treatments that can prevent development of epilepsy in patients at risk following brain injury. The aim of this review is to critically discuss the translational value of currently used animal models of seizures and epilepsy, particularly what animal models can tell us about epilepsy therapies in patients and which limitations exist. Principles of translational medicine will be used for this discussion. An essential requirement for translational medicine to improve success in drug development is the availability of animal models with high predictive validity for a therapeutic drug response. For this requirement, the model, by definition, does not need to be a perfect replication of the clinical condition, but it is important that the validation provided for a given model is fit for purpose. The present review should guide researchers in both academia and industry what can and cannot be expected from animal models in preclinical development of epilepsy therapies, which models are best suited for which purpose, and for which aspects suitable models are as yet not available. Overall further development is needed to improve and validate animal models for the diverse areas in epilepsy research where suitable fit for purpose models are urgently needed in the search for more effective treatments.
Cross-species pharmacological characterization of the allylglycine seizure model in mice and larval zebrafish
2015, Epilepsy and Behavior
Citation Excerpt :
Comparison of results between mice and zebrafish showed that seizure onset occurred in both species after a similar latency time. This latency period was described previously in mice (44–240 min) and in rats (120–150 min) by Horton and Meldrum [41] and Alberici et al. [64], respectively. A wide variation in seizure onset between test animals was also reported by these authors.
Treatment-resistant seizures affect about a third of patients suffering from epilepsy. To fulfill the need for new medications targeting treatment-resistant seizures, a number of rodent models offer the opportunity to assess a variety of potential treatment approaches. The use of such models, however, has proven to be time-consuming and labor-intensive. In this study, we performed pharmacological characterization of the allylglycine (AG) seizure model, a simple in vivo model for which we demonstrated a high level of treatment resistance. (d,l)-Allylglycine inhibits glutamic acid decarboxylase (GAD) – the key enzyme in γ-aminobutyric acid (GABA) biosynthesis – leading to GABA depletion, seizures, and neuronal damage. We performed a side-by-side comparison of mouse and zebrafish acute AG treatments including biochemical, electrographic, and behavioral assessments. Interestingly, seizure progression rate and GABA depletion kinetics were comparable in both species. Five mechanistically diverse antiepileptic drugs (AEDs) were used. Three out of the five AEDs (levetiracetam, phenytoin, and topiramate) showed only a limited protective effect (mainly mortality delay) at doses close to the TD₅₀ (dose inducing motor impairment in 50% of animals) in mice. The two remaining AEDs (diazepam and sodium valproate) displayed protective activity against AG-induced seizures. Experiments performed in zebrafish larvae revealed behavioral AED activity profiles highly analogous to those obtained in mice. Having demonstrated cross-species similarities and limited efficacy of tested AEDs, we propose the use of AG in zebrafish as a convenient and high-throughput model of treatment-resistant seizures.
Glutamate decarboxylase of the parasitic arthropods Ctenocephalides felis and Rhipicephalus microplus: Gene identification, cloning, expression, assay development, identification of inhibitors by high throughput screening and comparison with the orthologs from Drosophila melanogaster and mouse
2013, Insect Biochemistry and Molecular Biology
Citation Excerpt :
However, early attempts to assess this question were hampered by the lack of specificity of PALP-addressing GAD inhibitors (Gammon et al., 1986, 1987). GAD inhibition by hydrazines, hydroxylamines, mercaptopropionic acid, allylglycine and other compounds is known to generate convulsions in mammals (Killam and Bain, 1957; Killam, 1957; Balzer et al., 1960; Roberts and Simonsen, 1963; Alberici et al., 1969; Lamar 1970, Horton and Meldrum, 1973). Therefore, it is essential that any GAD-inhibiting pesticide or parasiticide possesses arthropod specificity, a property so far unknown for established GAD inhibitors (Tunnicliff, 1990).
Glutamate decarboxylase (l-glutamate 1-carboxylyase, E.C. 4.1.1.15, GAD) is the rate-limiting enzyme for the production of γ-aminobutyric acid (GABA), the major inhibitory neurotransmitter in vertebrates and invertebrates. We report the identification, isolation and characterization of cDNAs encoding GAD from the parasitic arthropods Ctenocephalides felis (cat flea) and Rhipicephalus microplus (cattle tick). Expression of the parasite GAD genes and the corresponding Drosophila melanogaster (fruit fly) GAD1 as well as the mouse GAD₆₅ and GAD₆₇ genes in Escherichia coli as maltose binding protein fusions resulted in functional enzymes in quantities compatible with the needs of high throughput inhibitor screening (HTS). A novel continuous coupled spectrophotometric assay for GAD activity based on the detection cascade GABA transaminase/succinic semialdehyde dehydrogenase was developed, adapted to HTS, and a corresponding screen was performed with cat flea, cattle tick and fruit fly GAD. Counter-screening of the selected 38 hit substances on mouse GAD₆₅ and GAD₆₇ resulted in the identification of non-specific compounds as well as inhibitors with preferences for arthropod GAD, insect GAD, tick GAD and the two mouse GAD forms. Half of the identified hits most likely belong to known classes of GAD inhibitors, but several substances have not been described previously as GAD inhibitors and may represent lead optimization entry points for the design of arthropod-specific parasiticidal compounds.
l-Allylglycine dissociates the neural substrates of fear in the periaqueductal gray of rats
2010, Brain Research Bulletin
The dorsal (dPAG) and ventral (vPAG) regions of the periaqueductal gray are well known to contain the neural substrates of fear and anxiety. Chemical or electrical stimulation of the dPAG induces freezing, followed by a robust behavioral reaction that has been considered an animal model of panic attack. In contrast, the vPAG is part of a neural system, in which immobility is the usual response to its stimulation. The defense reaction induced by the stimulation of either region is accompanied by antinociception. Although GABAergic mechanisms are known to exert tonic inhibitory control on the neural substrates of fear in the dPAG, the role of these mechanisms in the vPAG is still unclear. The present study examined defensive behaviors and antinociception induced by microinjections of an inhibitor of γ-aminobutyric acid synthesis, l-allylglycine (l-AG; 1, 3, and 5 μg/0.2 μl), into either the dPAG or vPAG of rats subjected to the open field and tail-flick tests. Passive or tense immobility was the predominant behavior after l-AG (1 or 3 μg) microinjection into the vPAG and dPAG, respectively, which was replaced with intense hyperactivity, including jumps or rearings, after injections of a higher dose (5 μg/0.2 μl) into the dPAG or vPAG. Moreover, whereas intra-dPAG injection of 3 μg l-AG produced intense antinociception, only weak antinociception was induced by intra-vPAG injections of 5 μg l-AG. These findings suggest that GABA mechanisms are involved in the mediation of antinociception and behavioral inhibition to aversive stimulation of the vPAG and exert powerful control over the neural substrates of fear in the dPAG to prevent a full-blown defense reaction possibly associated with panic disorder.
Chronic administration of DL-allyl-glycine into the neostriatum, disorganises the firing modes of the nigral dopaminergic neurons in the rat
1997, Neuroscience Letters
Nigral dopaminergic (DA) neurons have been reported to fire according to three modes: very regular (pacemaker 42%) irregular (random 46%) and bursty (12%). The switch from simple spiking mode (pacemaker or random) to bursty firing would correspond to an increase in DA release necessary for the performance of a new motor act. As nigral DA cells are impinged upon by a high percentage of GABAergic afferents we blocked striatal GABAergic output neurons by chronic administration into the neostriatum of allyl-glycine, a glutamic acid decarboxylase (GAD) inhibitor. After treatment, rats presented hyperkinesia and hypertonia on the injected side and recordings showed a drastic change in the percentage distribution of nigral DA cell discharge patterns; 85% were `random', 12% `pacemaker' and 3% bursty. Such a disturbance, by impeding adapted DA release, may account for the hyperkinetic and dystonic disorders observed.
Allylglycine-induced seizures in male and female rats
1991, Physiology and Behavior
The possibility that a sex difference in γ-aminobutyric acid (GABA) activity might be an underlying cause of the previously reported sex differences in picrotoxin-induced seizures was investigated. Male and female rats were injected (IP) with the GABA antagonist, L-allylglycine (100, 150, 200, or 250 mg/kg), and observed for behavioral signs of five categories of seizures induced by picrotoxin: myoclonic, focal, generalized tonic extension, generalized clonic, and generalized seizures with tonic and clonic components. The latency to the first occurrence of each seizure category was scored. Of the five categories of seizures investigated, only focal and generalized tonic extension seizures were observed after allylglycine in the doses tested. Female rats were significantly more susceptible than male rats to allylglycine-induced focal (p<0.007) and generalized tonic extension (p<0.05) seizures. The results suggest that male-female differences in presynaptic GABA activity may have relevance for sex differences in the occurrence of focal and tonic extension seizures. Differences in the seizure profiles associated with allylglycine and picrotoxin suggests that pre- and postsynaptic antagonism of GABA activity may have different consequences for specific seizure categories and the sex differences associated with those categories.

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Glutamic acid decarboxylase inhibition and ultrastructural changes by the convulsant drug allylglycine

Abstract

Biochem. Pharmac.

Biochem. Pharmac.

J. biol. Chem.

Life Sci.

Biochem. Pharmac.

Life Sci.

J. Neurochem.

J. Neurochem.

J. Neurochem.