Picrotoxin Research Articles

GABA A receptors in deep cerebellar nuclei play important roles in mouse eyeblink conditioning

The neural circuitry of eyeblink conditioning in rabbits has been studied in detail, however, the basic knowledge of eyeblink conditioning in mice remains limited. In the present study, we examined the role of the deep cerebellar nuclei (DCN) in mice in delay eyeblink conditioning and rotor rod test performance by using the γ-aminobutyric acid A (GABA A) receptor agonist muscimol (MSC) and the GABA A receptor antagonist picrotoxin (PTX). Bilateral injections of MSC and PTX into the DCN significantly impaired motor coordination in the rotor rod test, however the performance recovered within 24 h after the injections. Bilateral injection of MSC and PTX significantly impaired learned eyeblink responses (LER) during the acquisition test. MSC-injected mice could not acquire LER, however, PTX-injected mice acquired LER latently, suggesting the distinctive effect of these drugs in DCN. Bilateral injection of MSC and PTX also impaired the retention of acquired LER during a 7-day performance test. Furthermore, ipsilateral injections of MSC and PTX impaired the acquired LER as much as bilateral injection of them. Contralateral MSC injections also impaired the expression of LER, but contralateral PTX injections only partially impaired eyeblink conditioning. These results suggest that GABA A receptors in bilateral DCN play important roles in both the acquisition and the expression of mouse eyeblink conditioning, and that GABA A receptors not only in ipsilateral but also in contralateral DCN are critical for the expression of LER.

Stoichiometric Pore Mutations of the GABA AR Reveal a Pattern of Hydrogen Bonding with Picrotoxin

Picrotoxin (PTX) is a noncompetitive antagonist of many ligand-gated ion channels, with a site of action believed to be within the ion-conducting pore. In the A-type gamma-aminobutyric acid receptor, a threonine residue in the second transmembrane domain is of particular importance for the binding of, and ultimate inhibition by, PTX. To better understand the relationship between this residue and the PTX molecule, we mutated this threonine residue to serine, valine, and tyrosine to change the structural and biochemical characteristics at this location. The known subunit stoichiometry of the A-type gamma-aminobutyric acid receptor allowed us to create receptors with anywhere from zero to five mutations. With an increasing number of mutated subunits, each amino acid substitution revealed a unique pattern of changes in PTX sensitivity, ultimately encompassing sensitivity shifts over several orders of magnitude. The electrophysiological data on PTX-mediated block, and supporting modeling and docking studies, provide evidence that an interaction between the PTX molecule and three adjacent uncharged polar amino acids at this position of the pore are crucial for PTX-mediated inhibition.

Antiabsence Effects of Safranal in Acute Experimental Seizure Models: EEG and Autoradiography

We examined the effect of safranal, a constituent of Crocus sativus, in acute experimental animal models of generalized absence seizures. the effect of acute systemic administration of safranal on latency to seizure onset as well as spike and wave discharches (SWD) duration following pharmacologically-induced absence seizures was investigated in wildtype mice. We further characterized its effects on the GABAergic system through the regional modification of [3H] flunitrazepam, a benzodiazepine agonist binding site and [3H] CGP54626A, a GABAB receptor antagonist binding site in mouse brain. The systemic administration of safranal resulted in a significant and dose-dependent attenuation in experimental absence seizures elicited by either gamma-butyrolactone (GBL), baclofen (BAC) or low doses of GABAA receptor antagonists; pentylenetetrazole (PTZ), picrotoxin (PTX) and bicuculline (BMC). After a single intraperitoneal administration of safranal (291 mg/kg), no changes in baseline electrocorticographic (ECoG) recording were observed, however, a significant decrease in [3H] flunitrazepam binding was seen in the cortex (33.16%, p<0.001), hippocampus (27.36%, p<0.01) and thalamus (29.91%, p<0.01) of mouse brain, while the [3H] CGP54626A binding did not show any modification in the same brain regions. These data indicate that there is an antiabsence seizure property in safranal and its effect may be due to modifications on the benzodiazepine binding sites of the GABAA receptor complex.

The Lateral Hypothalamus: A Locus for GABAA Receptor Modulation of Feeding in the Rat

The goal of the present study was to determine whether the lateral hypothalamus (LH) is a specific site for feeding produced by injection of GABAA receptor (GABAAR) antagonists. Two cannula‐mapping studies were performed in which the GABAAR antagonist picrotoxin (PIC) was injected into LH or surrounding sites. In the first experiment, adult male rats with indwelling unilateral guide cannulas in the lateral preoptic area, dorsal medial hypothalamus, internal capsule/entopeduncular nucleus, thalamus, ventral tegmental area or LH were injected with 0.3ul aCSF vehicle or PIC (10 ng, 20 ng or 80 ng/0.3 ul veh) and food intake was measured 30 min to 4 h post‐injection. We found that, of the six sites tested, only LH injection of 80 ng PIC elicited significant feeding, with mean intakes of 5 g in 30 min versus under 1 g in surrounding sites. In the second experiment animals with unilateral cannulas in the tuberal LH (tLH) or sites 1 mm anterior or posterior to the tLH were injected with aCSF or PIC. We found that 80 ng PIC elicited eating only in tLH, with an intake of 4.2 g in 30 min, whereas little or no feeding was elicited in either anterior or posterior sites. Finally, to determine if LH is a site where natural feeding can be suppressed by activation of GABAAR, we injected the GABAAR agonists, muscimol or isoguvacine, through bilateral cannulas targeted at LH. Muscimol suppressed spontaneous feeding at the onset of the nocturnal phase of the photoperiod and also feeding in 24 h fasted rats. Isoguvacine similarly suppressed nocturnal eating but was ineffective in fasted rats. Collectively, these data provide evidence that LH is a major locus at which blockade of GABAA receptors can disinhibit eating and where activation of these receptors can suppress natural eating.

Contribution of synaptic inhibition to the control of inspiratory hypoglossal motoneuronal (IHMN) activity in vivo

We examine the role of endogenous γ‐aminobutyric acid (GABA)ergic and glycinergic inputs on IHMN discharge activity in decerebrate, vagotomized, paralyzed, and mechanically ventilated dogs during hypercapnic hyperoxia. The GABAA receptor antagonist bicuculline (BIC) (200 μM), the GABAA receptor antagonist picrotoxin (PIC) (5 mM), or the glycine receptor antagonist strychnine (STR) (200 μM) was picoejected onto extracellularly‐recorded single IHMNs using multibarrel micropipettes at increasing dose‐rates until no further change in neuronal discharge frequency was observed. Presence of GABAA and glycine receptors and effectiveness of the antagonist doses were confirmed by picoejection of GABA (2 mM) or glycine (2mM) with subsequent block by the respective antagonists. At maximally effective BIC dose‐rates, the peak frequency of 20 IHMNs increased by 29 ± 30%. BIC also increased the slope of the control inspiratory phase discharge pattern by 25 ± 28%. Neither PIC (n=17, 5 ± 19%), nor STR (n =24, 3 ± 15%) had a significant effect on discharge activity. These data suggest that the basal activity of IHMNs is attenuated by a BIC‐sensitive tonic GABAergic inhibition, but phasic GABAergic inhibition and glycine receptors, although present, are not involved in controlling the activity of these neurons. Support: VA Medical Research Funds &amp; NIH R01 GM59234.

The role of GABAergic transmission in the dentate gyrus on acquisition, consolidation and retrieval of an inhibitory avoidance learning and memory task in the rat

The hippocampal GABAergic interneurons are responsible for controlling the input of large principal cell populations, and they thereby determine the oscillatory discharge patterns and synaptic plasticity in the hippocampus. Such oscillations within neuronal systems serve various complex functions, such as perception, cognition, plasticity and memory. The aim of this study is to define the function of GABAergic synaptic transmission in the dentate gyrus (DG) of the hippocampus in the different stages of inhibitory avoidance (IA) learning and memory in the rat. Two cannulae were implanted above the hippocampal DG. Then the rats were trained on a step-through IA learning task. Each rat received intra-DG injection of picrotoxin (PTX) or saline before training, after training or before the retrieval test. The results show that post-training injection of PTX impaired the IA memory. On the other hand, pre-training and pre-retrieval injection of PTX had no significant effect on the IA activity. Therefore, it seems that GABAergic transmission in the DG is involved in the consolidation step (but not in the acquisition and retrieval steps) of the IA task by controlling the input to the principal cells.

Anticonvulsant effect of Rhus chirindensis (Baker F.) (Anacardiaceae) stem-bark aqueous extract in mice

Extracts of Rhus chirindensis stem-bark are used extensively in South African traditional medicines for the treatment, management and/or control of an array of human ailments, including childhood convulsions and epilepsy. In this study, we investigated the anticonvulsant activity of the plant's stem-bark aqueous extract (RCE, 50–800 mg/kg i.p.) against pentylenetetrazole (PTZ)-, picrotoxin (PCT)- and bicuculline (BCL)-induced seizures in mice. Phenobarbitone and diazepam were used as reference anticonvulsant drugs for comparison. Single intraperitoneal injections of PTZ (90 mg/kg), PCT(10 mg/kg) or BCL (30 mg/kg) produced tonic-clonic seizures. Like the standard antiseizure drugs used, Rhus chirindensis stem-bark aqueous extract (RCE, 100–800 mg/kg i.p.) significantly delayed ( p < 0.05–0.001) the onset of, and antagonized pentylenetetrazole-induced seizures. The plant's stem-bark aqueous extract (RCE, 100–800 mg/kg i.p.) also profoundly antagonized picrotoxin-induced seizures, but only weakly antagonized bicuculline-induced seizures. Although the data obtained in the present study do not provide conclusive evidence, it would appear that RCE produces its antiseizure effect by enhancing GABAergic neurotransmission and/or action in the brain. The results of this laboratory animal study indicate that RCE possesses anticonvulsant activity in the mammalian experimental model used, and thus suggest that the plant may be used as a natural supplementary remedy in the management, control and/or treatment of childhood convulsions and epilepsy. In conclusion, the findings of this study lend pharmacological credence to the suggested folkloric, ethnomedical uses of Rhus chirindensis in the management of childhood convulsions and epilepsy in some rural communities of South Africa.

Long-term dark rearing induces permanent reorganization in retinal circuitry

Recent data challenged the assumption that light has little effect on retina development. Here, we report evidence that dark rearing permanently changes the synaptic input to GCs. A reduced spontaneous postsynaptic currents (SPSCs) frequency was found in retinal GCs from rats born and raised in the dark for three months. Glutamate antagonists (CNQX and AP-5) reversibly reduced SPSCs frequency in control and dark-reared (DR) retinae. The GABA antagonist picrotoxin (PTX) reduced SPSCs frequency in control retinas, but increased SPSCs frequency in DR, mainly by presynaptic action on excitatory currents. In DR animals exposed to normal cyclic light for 3 months, SPSCs frequency remained lower then in control rats and increased following PTX, suggesting that long-term dark rearing induces permanent modifications of the retinal circuitry. Our results strongly support the idea that light stimulation plays a role in establishing normal synaptic input to GCs.

The pattern of ATP‐sensitive K+ channel subunits, Kir6.2 and SUR1 mRNA expressions in DG region is different from those in CA1‐3 regions of chronic epilepsy induced by picrotoxin in rats

ATP-sensitive K(+) (K(ATP)) channel's function is a key determinant of both excitability and viability of neurons. In the present report, in situ hybridization histochemistry and Western blot were used to examine whether picrotoxin (PTX)-kindling convulsions involved the changes in distribution of K(ATP) channels. The data demonstrated that the formation of kindling state was associated with a decreased amount of Kir6.2 mRNAs and proteins both in cerebral cortex and dentate gyrus (DG) as well as with a decreased amount of (regulatory subunit) SUR 1 mRNAs in DG. In contrast, resulting from a PTX re-induced seizure insult, both subunits were transiently up-modulated but not exactly paralleled between them and among different regions. In DG, Kir6.2 mRNAs increased toward normal levels at 12 h, followed a gradual decrease from 1 day to 3 days, being distinct from that detected in CA1-3 regions in which no significant change was shown. Further, SUR1 mRNAs markedly increased at 12 h, decreased significantly at 1 day, and even went down to a faint level at 3 days which was similar to that of CA1-3 regions, and there was no significant change in CA1-3 regions of SUR1 mRNAs. Also, at 7 days after a PTX re-treatment, Kir6.2 proteins increased significantly in the cortex, CA1, CA3 and DG (increasing 49.52%, 39.36%, 33.41%, and 54.79%, respectively) as well, SUR1 proteins increased significantly in DG (increasing 3.42 times), as compared with kindling rats without PTX retreatments (P < 0.05). These results indicated that K(ATP) channels in brain particularly in DG are likely related to enhanced seizure susceptibility and dynamic controls of seizure propagation of chronic epilepsy induced by PTX in rats.

Processing of Odor Mixtures in theDrosophilaAntennal Lobe Reveals both Global Inhibition and Glomerulus-Specific Interactions

To understand how odor information is represented and processed in the antennal lobe (AL) of Drosophila melanogaster, we have optically recorded glomerular calcium responses to single odors and odor mixtures from olfactory sensory neurons (OSNs) and projection neurons (PNs). Odor mixtures offer a good tool to analyze odor processing because experimental results can be tested against clear predictions. At the level of the OSNs, the representation of odor mixtures could be predicted from the response patterns of the components in most cases. PN responses to mixtures, however, provide evidences of interglomerular inhibition. Application of picrotoxin (PTX), an antagonist of GABA(A)-like receptors, enhanced odor responses, modified their temporal course, and eliminated mixture suppression at the PN level. Our results can be best explained by postulating the existence of at least two local networks in the fly AL: a glomerulus specific network, which includes excitatory and inhibitory connections and a PTX sensitive inhibitory global network that acts on all glomeruli with proportional strength to the global AL input.

Anticonvulsant property of Sutherlandia frutescens R. BR. (variety Incana E. MEY.) [Fabaceae] shoot aqueous extract

Aerial parts of Sutherlandia frutescens R. BR. [family: Fabaceae] are extensively used in South African traditional medicines for the treatment, management and/or control of an array of human ailments, including childhood convulsions and epilepsy. In this study, we examined the anticonvulsant property of the plant's shoot aqueous extract (SFE, 25–400 mg/kg i.p.) against pentylenetetrazole (PTZ)-, picrotoxin (PCT)- and bicuculline (BCL)-induced seizures in mice. Phenobarbitone and diazepam were used as reference anticonvulsant drugs for comparison. Like the reference antiseizure drugs used, S. frurescens shoot aqueous extract (SFE, 50–400 mg/kg i.p.) significantly delayed ( p < 0.05–0.001) the onset of, and antagonized, pentylenetetrazole (PTZ)-induced seizures. The plant's shoot aqueous extract (SFE, 50–400 mg/kg i.p.) also profoundly antagonized picrotoxin (PCT)-induced seizures, but only weakly antagonized bicuculline (BCL)-induced seizures. Although the data obtained in the present study do not provide conclusive evidence, it appears that S. frurescens shoot aqueous extract (SFE) produces its antiseizure effect directly by acting like GABA, or indirectly by enhancing GABAergic neurotransmission and/or action in the brain. The results of this laboratory animal study suggest that the herb may be used as a natural supplementary remedy in the management, control and/or treatment of childhood convulsions and epilepsy. In conclusion, the findings of this study indicate that S. frurescens shoot aqueous extract possesses anticonvulsant activity, and thus lend pharmacological credence to the suggested folkloric, ethnomedical uses of the herb in the management or treatment of childhood convulsions and epilepsy in some rural communities of South Africa.

Anticonvulsant activity of Hypoxis hemerocallidea Fisch. &amp; C. A. Mey. (Hypoxidaceae) corm (‘African potato’) aqueous extract in mice

Extracts of Hypoxis hemerocallidea Fisch. & C. A. Mey. (Hypoxidaceae) corm (popularly known as 'African potato') are extensively used in South African traditional medicines for the treatment, management and/or control of an array of human ailments, including childhood convulsions and epilepsy. This study examined the anticonvulsant activity of the plant's corm aqueous extract (APE, 50-800 mg/kg i.p.) against pentylenetetrazole (PTZ)-, picrotoxin (PCT)- and bicuculline (BCL)-induced seizures in mice. Phenobarbitone and diazepam were used as reference anticonvulsant drugs for comparison. Like the reference antiseizure drugs used, Hypoxis hemerocallidea corm aqueous extract (APE, 100-800 mg/kg i.p.) significantly delayed (p < 0.05-0.001) the onset of, and antagonized, pentylenetetrazole (PTZ)-induced seizures. The plant's corm aqueous extract (APE, 100-800 mg/kg i.p.) also profoundly antagonized picrotoxin (PCT)-induced seizures, but only weakly antagonized bicuculline (BCL)-induced seizures. Although the data obtained in the present study do not provide conclusive evidence, it would appear that 'African potato' aqueous extract (APE) produces its antiseizure effect by enhancing GABAergic neurotransmission and/or action in the brain. The results of this laboratory animal study indicate that APE possesses anticonvulsant activity in the mammalian experimental model used and, therefore, tend to suggest that the herb may be used as a natural supplementary remedy in the management, control and/or treatment of childhood convulsions and epilepsy. In conclusion, the findings of this study indicate that Hypoxis hemerocallidea corm aqueous extract possesses anticonvulsant activity, and thus lend pharmacological credence to the suggested folkloric, anecdotal ethnomedical uses of the herb in the management of childhood convulsions and epilepsy in some rural communities of South Africa.

A proposed structural basis for picrotoxinin and picrotin binding in the glycine receptor pore

Picrotoxin, an antagonist of structurally-rated GABA(A) receptors (GABA(A)Rs) and glycine receptors (GlyRs), is an equimolar mixture of picrotoxinin (PTXININ) and picrotin (PTN). These compounds share a common structure except that PTN contains a slightly larger dimethylmethanol in place of the PTXININ isopropenyl group. Although the homomeric alpha1 GlyR is equally sensitive to both compounds, we show here that homomeric alpha2 and alpha3 GlyRs, like most GABA(A)Rs, are selectively inhibited by PTXININ. As conservative mutations to pore-lining 6' threonines equally affect the sensitivity of the alpha1 GlyR to both compounds, we conclude that PTXININ and PTN bind to 6' threonines by hydrogen bonding with exocyclic oxygens common to both molecules. In contrast, substitution of the 2' pore-lining glycine by serine selectively reduces PTN sensitivity, whereas the introduction of 2' alanines selectively increases PTXININ sensitivity. These results define the orientation of PTXININ and PTN binding in the alpha1 GlyR pore and allow us to conclude that the relatively reduced sensitivity of PTN at GABA(A)Rs and alpha2 and alpha3 GlyRs is due predominantly to its larger size and reduced ability to form hydrophobic interactions with 2' alanines.

Regional differences in GABAergic modulation for TEA-induced synaptic plasticity in rat hippocampal CA1, CA3 and dentate gyrus

Tetraethylammonium (TEA), a K +-channel blocker, reportedly induces long-term potentiation (LTP) of hippocampal CA1 synaptic responses, but at CA3 and the dentate gyrus (DG), the characteristics of TEA-induced plasticity and modulation by inhibitory interneurons remain unclear. This study recorded field EPSPs from CA1, CA3 and DG to examine the involvement of GABAergic modulation in TEA-induced synaptic plasticity for each region. In Schaffer collateral-CA1 synapses and associational fiber (AF)-CA3 synapses, bath application of TEA-induced LTP in the presence and absence of picrotoxin (PTX), a GABA A receptor blocker, whereas TEA-induced LTP at mossy fiber (MF)-CA3 synapses was detected only in the absence of GABA A receptor blockers. MF-CA3 LTP showed sensitivity to Ni 2+, but not to nifedipine. In DG, synaptic plasticity was modulated by GABAergic inputs, but characteristics differed between the afferent lateral perforant path (LPP) and medial perforant path (MPP). LPP-DG synapses showed TEA-induced LTP during PTX application, whereas at MPP-DG synapses, TEA-induced long-term depression (LTD) was seen in the absence of PTX. This series of results demonstrates that TEA-induced DG and CA3 plasticity displays afferent specificity and is exposed to GABAergic modulation in an opposite manner.

Effects of GABAergic inhibition on neocortical long-term potentiation in the chronically prepared rat

Long-term potentiation (LTP) is a form of activity-dependent synaptic plasticity that is a candidate cellular mechanism for some forms of learning and memory. Although GABAergic synaptic inhibition plays a critical role in regulating of synaptic plasticity, there is still little known about the GABAergic modulation on LTP induction in chronic preparation. In the present study we examined the effect of GABA(A) agonist, diazepam (DZM), and antagonist, picrotoxin (PTX) on the induction of LTP in the somatosensory cortex of freely moving rats for a long-term period. In adult rats a bipolar stimulating and recording electrode were implanted into corpus callusom and somatosensory cortex, respectively. Two weeks after the surgery, evoked field potential responses were recorded before, during (12 days), and after (1 month) induction period of LTP by high-frequency stimulation. The LTP characteristics were compared between control, DZM and PTX groups during the time course of recording in each rat. Administration of DZM prior to train, blocked the induction of neocortical LTP, while the PTX increased the development of LTP making the highest differential levels of LTP about 12 days after the initiation of LTP induction. Our findings suggest that the augmentation of LTP by PTX can be explained by an interaction between excitatory and inhibitory pathways. Suppression of neocortical inhibitory inputs by PTX causes enhancement in LTP induction. These results suggest that GABAergic system has an important role in synaptic plasticity and long-term modification of somatosensory cortex in freely moving rat.

Mechanisms for Picrotoxinin and Picrotin Blocks of α2 Homomeric Glycine Receptors

Contrary to its effect on the gamma-aminobutyric acid type A and C receptors, picrotoxin antagonism of the alpha1 homomeric glycine receptors (GlyRs) has been shown to be non-use-dependent and nonselective between the picrotoxin components picrotoxinin and picrotin. Picrotoxin antagonism of the embryonic alpha2 homomeric GlyR is known to be use-dependent and reflects a channel-blocking mechanism, but the selectivity of picrotoxin antagonism of the embryonic alpha2 homomeric GlyRs between picrotoxinin and picrotin is unknown. Hence, we used the patch clamp recording technique in the outside-out configuration to investigate, at the single channel level, the mechanism of picrotin- and picrotoxinin-induced inhibition of currents, which were evoked by the activation of alpha2 homomeric GlyRs stably transfected into Chinese hamster ovary cells. Although both picrotoxinin and picrotin inhibited glycine-evoked outside-out currents, picrotin had a 30 times higher IC50 than picrotoxinin. Picrotin-evoked inhibition displayed voltage dependence, whereas picrotoxinin did not. Picrotoxinin and picrotin decreased the mean open time of the channel in a concentration-dependent manner, indicating that these picrotoxin components can bind to the receptor in its open state. When picrotin and glycine were co-applied, a large rebound current was observed at the end of the application. This rebound current was considerably smaller when picrotoxinin and glycine were co-applied. Both picrotin and picrotoxinin were unable to bind to the unbound conformation of the receptor, but both could be trapped at their binding site when the channel closed during glycine dissociation. Our data indicate that picrotoxinin and picrotin are not equivalent in blocking alpha2 homomeric GlyR.

Anticonvulsant properties of the total alkaloid fraction of Rauvolfia ligustrina Roem. et Schult. in male mice

Rauvolfia ligustrina Roem et. Schult (Apocynaceae), commonly known as paratudo and arrebenta-boi is a small tree found in Brazilian Northeastern. Previous studies have demonstrated depressant and anticonvulsant properties of the ethanol extract of Rauvolfia ligustrina. The aim of the present study was the determination of the lethal dose 50% (LD50) and the effects of total alkaloid fraction (TAF) of the aerial parts of R. ligustrina in animal models of convulsion. It was found that the acute toxicity of TAF was 127.8 (112.5-145.2) mg/kg (i.p.) in mice. TAF (20 mg/kg, ip) significantly increased (p < 0.05) the latencies of clonic seizures induced by pentylenetetrazol (PTZ) and picrotoxin (PIC). However, TAF did not protect the animals in maximal electroshock (MES) induced seizures. These results suggest that TAF of R. ligustrina possesses anticonvulsant properties.

Anticonvulsant effect of Sclerocarya birrea (A. Rich.) Hochst. subsp. caffra (Sond.) Kokwaro (Anacardiaceae) stem-bark aqueous extract in mice

Sclerocarya birrea (A.Rich.) Hochst. subsp. caffra (Sond.) Kokwaro (family: Anacardiaceae) is traditionally used in South African folk medicine for the treatment, management and control of a variety of human ailments, including childhood convulsions and epilepsy. In the present study, we have investigated the anticonvulsant activity of the plant’s stem-bark aqueous extract (SBE) against pentylenetetrazole (PTZ)-, picrotoxin (PCT)-, and bicuculline (BCL)-induced seizures in mice. Phenobarbital and diazepam were used as reference anticonvulsant drugs for comparison. Like the reference anticonvulsant agents used, S. birrea SBE (100–800 mg/kg p.o.) significantly (P 0.05). The plant extract also depressed the central nervous system. In conclusion, the findings of the present experimental animal study indicate that S. birrea SBE possesses anticonvulsant activity, thus lending pharmacological support to the suggested folkloric, ethnomedical uses of the plant extract in the management, control, and treatment of childhood convulsions and epilepsy in some rural communities of South Africa.

Anticonvulsant effect of Persea americana Mill (Lauraceae) (Avocado) leaf aqueous extract in mice

Various morphological parts of Persea americana Mill (Lauraceae) (avocado) are widely used in African traditional medicines for the treatment, management and/or control of a variety of human ailments, including childhood convulsions and epilepsy. This study examined the anticonvulsant effect of the plant's leaf aqueous extract (PAE, 50-800 mg/kg i.p.) against pentylenetetrazole (PTZ)-, picrotoxin (PCT)- and bicuculline (BCL)-induced seizures in mice. Phenobarbitone and diazepam were used as reference anticonvulsant drugs for comparison. Like the reference anticonvulsant agents used, Persea americana leaf aqueous extract (PAE, 100-800 mg/kg i.p.) significantly (p < 0.05-0.001) delayed the onset of, and antagonized, pentylenetetrazole (PTZ)-induced seizures. The plant's leaf extract (PAE, 100-800 mg/kg i.p.) also profoundly antagonized picrotoxin (PCT)-induced seizures, but only weakly antagonized bicuculline (BCL)-induced seizures. Although the data obtained in the present study do not provide conclusive evidence, it would appear that 'avocado' leaf aqueous extract (PAE) produces its anticonvulsant effect by enhancing GABAergic neurotransmission and/or action in the brain. The findings of this study indicate that Persea americana leaf aqueous extract possesses an anticonvulsant property, and thus lends pharmacological credence to the suggested ethnomedical uses of the plant in the management of childhood convulsions and epilepsy.

Mechanisms of anabolic androgenic steroid inhibition of mammalian ɛ‐subunit‐containing GABAA receptors

GABAergic transmission regulates the activity of gonadotrophin-releasing hormone (GnRH) neurons in the preoptic area/hypothalamus that control the onset of puberty and the expression of reproductive behaviours. One of the hallmarks of illicit use of anabolic androgenic steroids (AAS) is disruption of behaviours under neuroendocrine control. GnRH neurons are among a limited population of cells that express high levels of the epsilon-subunit of the GABAA receptor. To better understand the actions of AAS on neuroendocrine mechanisms, we have characterized modulation of GABAA receptor-mediated currents in mouse native GnRH neurons and in heterologous cells expressing recombinant alpha2beta3epsilon-receptors. GnRH neurons exhibited robust currents in response to millimolar concentrations of GABA and a picrotoxin (PTX)-sensitive, bicuculline-insensitive current that probably arises from spontaneous openings of GABAA receptors. The AAS 17alpha-methyltestosterone (17alpha-MeT) inhibited spontaneous and GABA-evoked currents in GnRH neurons. For recombinant alpha2beta3epsilon-receptors, 17alpha-MeT inhibited phasic and tonic GABA-elicited responses, accelerated desensitization and slowed paired pulse response recovery. Single channel analysis indicated that GABA-evoked events could be described by three open dwell components and that 17alpha-MeT enhanced residence in the intermediate dwell state. This AAS also inhibited a PTX-sensitive, spontaneous current (open probability, approximately 0.15-0.2) in a concentration-dependent fashion (IC50 approximately 9 microm). Kinetic modelling indicated that the inhibition induced by 17alpha-MeT occurs by an allosteric block in which the AAS interacts preferentially with a closed state and promotes accumulation in that state. Finally, studies with a G302S mutant epsilon-subunit suggest that this residue within the transmembrane domain TM2 plays a role in mediating AAS binding and modulation. In sum, our results indicate that inclusion of the epsilon-subunit significantly alters the profile of AAS modulation and that this allosteric inhibition of native GnRH neurons should be considered with regard to AAS disruption of neuroendocrine control.