Isolated trochlear nerve palsy caused by mixed dural-pial arteriovenous malformation of the anterior cranial fossa: a case report

The present study makes use of the photic induction of Fos in the suprachiasmatic nucleus (SCN) to explore the pharmacology of retinal input to this circadian pacemaker. Our results demonstrate that the GABAA antagonist bicuculline and the benzodiazepine agonist diazepam, both of which prevent light-induced phase shifts, do not inhibit photic induction of Fos expression in the hamster SCN. In contrast, the GABAB agonist, baclofen, prevents both light-induced phase shifts and inhibits photic induction of Fos expression in the SCN. One explanation of this difference may be that baclofen acts to prevent photic information from reaching the SCN while bicuculline and diazepam act within the SCN at a point 'downstream' from Fos induction.     

GABAB receptor stimulation phase-shifts the mammalian circadian clock in vitro

The mammalian circadian clock in the suprachiasmatic nucleus (SCN) generates 24-h rhythms in vitro. Here we show that the GABAB agonist baclofen resets the SCN pacemaker in vitro in a phase-dependent manner: advances were induced at zeitgeber time (ZT) 6, and delays were induced at ZT 22. Both effects were blocked the GABAB antagonist, 2-hydroxysaclofen, while the GABAA antagonist, bicuculline was ineffective. Thus, the SCN pacemaker is sensitive to resetting by GABAB stimulation.     

Presynaptic GABAB autoreceptor modulation of P/Q-type calcium channels and GABA release in rat suprachiasmatic nucleus neurons

GABA is the primary transmitter released by neurons of the suprachiasmatic nucleus (SCN), the circadian clock in the brain. Whereas GABAB receptor agonists exert a significant effect on circadian rhythms, the underlying mechanism by which GABAB receptors act in the SCN has remained a mystery. We found no GABAB receptor-mediated effect on slow potassium conductance, membrane potential, or input resistance in SCN neurons in vitro using whole-cell patch-clamp recording. In contrast, the GABAB receptor agonist baclofen (1-100 microM) exerted a large and dose-dependent inhibition (up to 100%) of evoked IPSCs. Baclofen reduced the frequency of spontaneous IPSCs but showed little effect on the frequency or amplitude of miniature IPSCs in the presence of tetrodotoxin. The activation of GABAB receptors did not modulate postsynaptic GABAA receptor responses. The depression of GABA release by GABAB autoreceptors appeared to be mediated primarily through a modulation of presynaptic calcium channels. The baclofen inhibition of both calcium currents and evoked IPSCs was greatly reduced (up to 100%) by the P/Q-type calcium channel blocker agatoxin IVB, suggesting that P/Q-type calcium channels are the major targets involved in the modulation of GABA release. To a lesser degree, N-type calcium channels were also involved. The inhibition of GABA release by baclofen was abolished by a pretreatment with pertussis toxin (PTX), whereas the inhibition of whole-cell calcium currents by baclofen was only partially depressed by PTX, suggesting that G-protein mechanisms involved in GABAB receptor modulation at the soma and axon terminal may not be identical. We conclude that GABAB receptor activation exerts a strong presynaptic inhibition of GABA release in SCN neurons, primarily by modulating P/Q-type calcium channels at axon terminals.     

GABAB receptor-mediated inhibition of GABAA receptor calcium elevations in developing hypothalamic neurons

In the CNS, gamma-aminobutyric acid (GABA) affects neuronal activity through both the ligand-gated GABAA receptor channel and the G protein-coupled GABAB receptor. In the mature nervous system, both receptor subtypes decrease neural excitability, whereas in most neurons during development, the GABAA receptor increases neural excitability and raises cytosolic Ca2+ levels. We used Ca2+ digital imaging to test the hypothesis that GABAA receptor-mediated Ca2+ rises were regulated by GABAB receptor activation. In young, embryonic day 18, hypothalamic neurons cultured for 5 +/- 2 days in vitro, we found that cytosolic Ca2+ rises triggered by synaptically activated GABAA receptors were dramatically depressed (>80%) in a dose-dependent manner by application of the GABAB receptor agonist baclofen (100 nM-100 microM). Coadministration of the GABAB receptor antagonist 2-hydroxy-saclofen or CGP 35348 reduced the inhibitory action of baclofen. Administration of the GABAB antagonist alone elicited a reproducible Ca2+ rise in >25% of all synaptically active neurons, suggesting that synaptic GABA release exerts a tonic inhibitory tone on GABAA receptor-mediated Ca2+ rises via GABAB receptor activation. In the presence of tetrodotoxin the GABAA receptor agonist muscimol elicited robust postsynaptic Ca2+ rises that were depressed by baclofen coadministration. Baclofen-mediated depression of muscimol-evoked Ca2+ rises were observed in both the cell bodies and neurites of hypothalamic neurons taken at embryonic day 15 and cultured for three days, suggesting that GABAB receptors are functionally active at an early stage of neuronal development. Ca2+ rises elicited by electrically induced synaptic release of GABA were largely inhibited (>86%) by baclofen. These results indicate that GABAB receptor activation depresses GABAA receptor-mediated Ca2+ rises by both reducing the synaptic release of GABA and decreasing the postsynaptic Ca2+ responsiveness. Collectively, these data suggest that GABAB receptors play an important inhibitory role regulating Ca2+ rises elicited by GABAA receptor activation. Changes in cytosolic Ca2+ during early neural development would, in turn, profoundly affect a wide array of physiological processes, such as gene expression, neurite outgrowth, transmitter release, and synaptogenesis.     

GABAergic effects on nucleus tractus solitarius neurons receiving gastric vagal inputs

Single units in the region of the medial nucleus tractus solitarius (NTS), responding to electrical stimulation of gastric vagal fibers, were recorded in an in vitro neonatal rat brainstem-gastric preparation. gamma-Aminobutyric acid (GABA) subreceptor agonists and antagonists were applied to the gastric and brainstem compartments of the bath chamber to evaluate the peripheral gastric and central brainstem GABAergic effects on NTS neuronal activity. The gastric effects of the GABAA receptor agonist muscimol and GABAB receptor agonist baclofen were evaluated on 55 tonic units that received the gastric vagal inputs. For approximately 58% (32 of 55) and 38% (21 of 55) of the units observed, muscimol (30 microM; IC50 = 2.0 microM) and baclofen (30 microM; IC50 = 1.5 microM) in the gastric compartment induced a concentration-dependent inhibition of 36.2 +/- 3.1% (mean +/- S.E.) and 31.0 +/- 2.9% of the control level of the NTS neuronal activity, respectively. The brainstem effects of muscimol and baclofen were tested on 51 units. For approximately 90% (46 of 51) and 78% (40 of 51) of the units tested, muscimol (30 microM; IC50 = 1.3 microM) and baclofen (30 microM; IC50 = 1.1 microM) in the brainstem compartment produced a concentration-dependent inhibition of 54.1 +/- 3.4% and 48.9 +/- 3. 5% of the control level, respectively. The remaining NTS units were not affected by these two GABA agonists. Bicuculline (10 microM) and saclofen (10 microM), the GABAA and GABAB subreceptor antagonists, competitively antagonized the gastric and brainstem effects by muscimol and baclofen, respectively. Our results demonstrated that both GABAA and GABAB receptors in the stomach and brainstem play an important role in activity modulation of the medial NTS neurons receiving gastric vagal inputs in neonatal rats.     

A comparative study of synchronised and conventional culture methods on the micronucleus dose-response curve

The involvement of GABAA and GABAB receptors in neural mechanisms responsible for the production of theta rhythms in hippocampal formation (HPC) slices is addressed in the present study. In a number of papers published in the last decade, we have demonstrated that theta-like activity can be successfully recorded in the limbic cortex maintained in vitro when the cholinergic agonists, acetylcholine, carbachol or muscarine, were added to the bath. Recently, we have also shown a strong GABAA modulation of the cholinergic-induced in vitro theta-like activity. This study presents a report of the first demonstration of in vitro theta-like field responses induced a consequence of simultaneously inhibiting hippocampal GABAA and GABAB receptors. HPC slices (350 microns) were maintained in a gas-liquid interface chamber (35 degrees C). Theta-like activity was induced in the presence of bath perfusion of bicuculline (GABAA antagonist) and 2-hydroxysaclophen (GABAB antagonist). This in vitro induced field response was antagonized both by muscimol (GABAA agonist) and baclophen (GABAB agonist). In addition, the experiments presented here revealed that bicuculline/2-hydroxysaclophen-induced in vitro theta-like activity also had a strong cholinergic M1 involvement: it was abolished by hemicholinium-3 (choline transport blocker) and pirenzepine (specific antagonist of M1 receptor), but not by gallamine (specific antagonist of M2 receptor). The results of the present study provided further evidence for a strong GABAergic/cholinergic interaction in the neural mechanism responsible for production of theta-like activity in the hippocampal formation slices.     

The effects of GABA and glycine on horizontal cells of the rabbit retina

Intracellular and patch-clamp recordings have been used to characterize GABA-activated channels in axonless horizontal cells (ALHC) of the rabbit retina. In our intracellular recordings on an everted eyecup preparation, GABA depolarized the horizontal cells (HC), diminished their light response amplitude and slowed the response rise time. Glycine showed similar effects on the HC light responses. In our whole cell patch-clamp recordings on dissociated ALHC, all HCs responded to 3 microM GABA but none to glycine, even at 100 microM. Dose-response relationship for GABA gave EC50 values around 10 microM and Hill slopes of 1.3. Whole-cell current-voltage (I-V) relationships of GABA-activated currents reversed close to the predicted Cl- equilibrium potential. Partial replacement of intracellular Cl- with isothetionate shifted the GABA reversal potential to a more negative value. Muscimol (30 microM), a GABAA agonist mimicked the effect of GABA, but baclofen (30 microM), a GABAB agonist and cis-aminocaprionic acid (30 microM), a GABAC agonist did not elicit any effect on ALHC. Responses to GABA were blocked by the GABAA receptor antagonist bicuculline (10 microM) and picrotoxin (100 microM). According to our results, we conclude that ALHC express GABA receptors coupled to ion channels, and they correspond to GABAA receptor subtypes.     

In vivo modulation of acetylcholine in the nucleus accumbens of freely moving rats: II. Inhibition by gamma-aminobutyric acid

Our earlier studies suggest dopamine and serotonin interact with acetylcholine (ACh) in the nucleus accumbens (NAC) as part of a system for motivation and reinforcement. The purpose of the present experiment was to characterize a possible link between GABA and acetylcholine in the nucleus accumbens using microdialysis in freely moving rats. Different doses of GABA, muscimol, baclofen, saclofen and bicuculline were locally infused into the nucleus accumbens through the microdialysis probe. GABA and its agonists dose-dependently decreased extracellular levels of acetylcholine in the nucleus accumbens. In contrast the GABAA antagonist, bicuculline, dose-dependently increased extracellular ACh while the GABAB antagonist, saclofen, was without effect. Co-infusion of bicuculline or saclofen was shown to block the decrease in recoverable ACh produced by muscimol or baclofen, respectively. The results demonstrate an inhibitory action of GABA on acetylcholine interneurones in the nucleus accumbens involving both GABAA and GABAB receptor subtypes. In addition a tonic inhibitory GABAergic tone is probably mediated through GABAA receptors.     

An AP-2 binding sequence within exon 1 of human and porcine choline acetyltransferase genes enhances transcription in neural cells

It has been established that GABAA and GABAB receptors can exist separately and/or co-exist in the membrane of dorsal root ganglion neurons. In our previous investigation it has been shown that co-existence of these two kinds of receptors is about 80% of the neurons examined (20/25). The present study was aimed to explore whether the activation of these two kinds of receptors could interact with each other using intracellular and whole-cell patch-clamp recordings. Baclofen, a specific GABAB receptor agonist, was found to exert negative modulatory effects on the responses mediated by GABAA receptor. In experiments with intracellular recording, GABA (0.3-1000 microM)- and muscimol (100-1000 microM)-induced depolarization was attenuated markedly and reversibly by preapplication of baclofen (100 microM) (15/21 and 17/21, respectively). In whole-cell patch-clamp recordings GABA (100 microM) and two specific GABAA receptor agonists, muscimol (10 microM) and isoguvacine (50 microM), activated currents were inhibited markedly by preapplication of baclofen 30 s or more and the inhibition was concentration dependent (1-100 microM baclofen) and reversible. The possible mechanisms underlying the inhibition by baclofen of the responses mediated by GABAA receptor and the physiological significance implicated are discussed.     

Transport activity of FhuA, FhuC, FhuD, and FhuB derivatives in a system free of polar effects, and stoichiometry of components involved in ferrichrome uptake

The role of GABA receptors in regulating the mesolimbic dopamine (DA) system and drug reinforced behaviors has not been well characterized. Using fast-cyclic voltammetry, the effects of specific GABA receptor modulation on DA release in the nucleus accumbens (NAcc) and heroin self-administration (SA) behavior was investigated. The GABAA agonist muscimol, administered either intravenously or directly into the ventral tegmental area (VTA), significantly increased DA release in the NAcc in 7 of the 10 rats tested. DA release decreased in the remaining three rats; both effects were blocked by pretreatment with the GABAA receptor antagonist bicuculline. In contrast, the GABAB agonist baclofen decreased, while 2-OH-saclofen (a GABAB antagonist) increased DA release in the NAcc. However, when VTA GABAB receptors were previously activated or inactivated by microinjections of baclofen or 2-OH-saclofen, systemic injections of muscimol caused an inhibition of NAcc DA release. These results suggest that GABAA receptors may be co-localized on both DA neurons and non-DA (GABAergic) interneurons in the VTA, with the effects of GABAA determined by the net effect of both direct inhibition and indirect disinhibition of DA neurons. Finally, although a DA releaser, muscimol was neither self-administered in drug naive rats, nor did it substitute for heroin in rats previously trained to self-administer heroin, suggesting that GABAA receptors appear to play a complex role in mediating drug reinforcement, depending upon the dynamic functional state of GABAA receptors on both tegmental DA and non-DA neurons.     

Presynaptic modulation by L-glutamate and GABA of sympathetic co-transmission in rat isolated vas deferens

1. The modulatory effects of L-glutamate and its structural analogues, and of gamma-aminobutyric acid (GABA), on sympathetic co-transmission were studied in the rat isolated vas deferens exposed to electrical field stimulation (EFS). 2. Application of exogenous L-glutamate caused a concentration-dependent (1 microM-3 mM) inhibition of the rapid twitch component of the biphasic EFS contraction. However, L-glutamate (1 microM-3 mM) had a minimal effect on the phasic contraction induced by exogenous adenosine 5'-triphosphate (ATP, 150 microM) and noradrenaline (50 microM). Unlike L-glutamate, D-glutamate had no effect on the EFS contraction. 3. The L-glutamate-induced inhibition of the EFS contractions was significantly attenuated by the glutamate decarboxylase (GAD) inhibitor 3-mercapto-propionic acid (150 microM) and was abolished in the presence of the GABA transaminase (GABA-T) inhibitor, 2-aminoethyl hydrogen sulphate (500 microM). 4. The L-glutamate-induced inhibition of the electrically evoked contraction was not affected by the adenosine A1-receptor antagonist, 8-cyclopentyl-1,3-dipropylxanthine (DPCPX)(30 nM), reactive blue 2 (30 microM) or the GABAA receptor antagonist bicuculline (50 microM). However, the GABAB receptor antagonist 2-hydroxysaclofen (50 microM) significantly inhibited the L-glutamate effect. 5. Similar to L-glutamate, GABA also caused a concentration-dependent (0.1-100 microM) inhibition of the EFS contractions. This GABA-induced inhibition was not affected by either the GABAA receptor antagonist bicuculline (50 microM) or reactive blue 2 (30 microM). However, a significant attenuation of the GABA-mediated effect was recorded with the GABAB receptor antagonist 2-hydroxysaclofen (50 microM). Contractions of the vas deferens induced by exogenous ATP and noradrenaline were not affected by GABA (0.1-100 microM). 6. The L-glutamate analogues, N-methyl-D-aspartate (NMDA) (1 microM-1 mM) and quisqualate (Quis 0.1 microM-0.3 mM) had no effect, whilst kainate (Kain, 1 microM-1 mM) caused an inhibition of the EFS-induced contractions. Effects of Kain could be abolished by the non-NMDA receptor antagonist 6-cyano-7-nitroquinoxaline-2,3-dioxine (CNQX, 10 microM). NMDA, Quis and Kain had no effect on the exogenous ATP- or noradrenaline-induced contractions. 7. It is concluded that the excitatory amino acid L-glutamate modulates the electrically evoked vas deferens contraction through conversion to the inhibitory amino acid GABA by a specific GABA transaminase. The GABA formed may then act on GABAB receptors and cause inhibition of the contraction through a presynaptic mechanism.     

Ontogenesis of presynaptic GABAB receptor-mediated inhibition in the CA3 region of the rat hippocampus

gamma-Aminobutyric acid-B(GABAB) receptor-dependent and -independent components of paired-pulse depression (PPD) were investigated in the rat CA3 hippocampal region. Intracellular and whole cell recordings of CA3 pyramidal neurons were performed on hippocampal slices obtained from neonatal (5-7 day old) and adult (27-34 day old) rats. Electrical stimulation in the hilus evoked monosynaptic GABAA postsynaptic currents (eIPSCs) isolated in the presence of the ionotropic glutamate receptor antagonists 6-cyano-7-nitroquinoxaline-2,3-dione (CNQX, 10 microM) and D(-)2-amino-5-phosphovaleric acid (-AP5, 50 microM) with 2(triethylamino)-N-(2,6-dimethylphenyl) acetamine (QX314) filled electrodes. In adult CA3 pyramidal neurons, when a pair of identical stimuli was applied at interstimulus intervals (ISIs) ranging from 50 to 1,500 ms the amplitude of the second eIPSC was depressed when compared with the first eIPSC. This paired-pulse depression (PPD) was partially blocked by P-3-aminoprophyl -P-diethoxymethylphosphoric acid (CGP35348, 0.5 mM), a selective GABAB receptor antagonist. In neonates, PPD was restricted to ISIs shorter than 200 ms and was not affected by CGP35348. The GABAB receptor agonist baclofen reduced the amplitude of eIPSCs in a dose-dependent manner with the same efficiency in both adults and neonates. Increasing the probability of transmitter release with high Ca2+ (4 mM)/low Mg2+ (0.3 mM) external solution revealed PPD in neonatal CA3 pyramidal neurons that was 1) partially prevented by CGP35348, 2) independent of the membrane holding potential of the recorded cell, and 3) not resulting from a change in the reversal potential of GABAA eIPSCs. In adults the GABA uptake blocker tiagabine (20 microM) increased the duration of eIPSCs and the magnitude of GABAB receptor-dependent PPD. In neonates, tiagabine also increased duration of eIPSCs but to a lesser extent than in adult and did not reveal a GABAB receptor-dependent PPD. These results demonstrate that although GABAB receptor-dependent and -independent mechanisms of presynaptic inhibition are present onGABAergic terminals and functional, they do not operate at the level of monosynaptic GABAergic synaptic transmission at early stages of development. Absence of presynaptic autoinhibition of GABA release seems to be due to the small amount of transmitter that can access presynaptic regulatory sites.     

Differential sensitivity of membrane-associated pyrophosphatases to inhibition by diphosphonates and fluoride delineates two classes of enzyme

The participation of GABAergic mechanisms in the regulation of circadian rhythmicity by the suprachiasmatic nuclei (SCN) has been suggested from different lines of evidence. Little is known, however, whether GABA synthesis, release, uptake or content within the SCN may show a circadian pattern. The present results show that the activity of the GABAergic system within the SCN region of the rat exhibits circadian rhythmicity, which is manifested by correlative changes of the GABA content and the glutamic acid decarboxylase activity under the light/dark cycle, and by changes in the GABA content in animals kept under constant darkness.     

Seizure sensitivity and GABAergic modulation of ethanol sensitivity in selectively bred FAST and SLOW mouse lines

FAST and SLOW selected mouse lines were bred for differences in locomotor response to low-dose ethanol. FAST mice exhibit an extreme stimulant response and SLOW mice exhibit locomotor depression at the same ethanol dose. We tested the hypothesis that gamma-aminobutyric acid (GABA) systems modulate ethanol's stimulant effects by examining convulsant responses to GABAA receptor ligands, and by assessing the effects of GABAA and GABAB ligands on locomotor activity in the presence and absence of EtOH. FAST mice were more sensitive to the convulsant effects of GABAA drugs, and to one of two non-GABAergic drugs also tested. FAST and SLOW mice differed in locomotor responses to two benzodiazepines, but not to other GABAA receptor ligands. Ethanol's stimulant effects were not selectively altered by bicuculline or picrotoxin. The selected lines differed in sensitivity to the locomotor depressant effects of the GABAB agonist, baclofen. Ethanol-stimulated activity of FAST mice was inhibited by baclofen, and this effect was reversed by administration of the GABAB antagonist, CGP-35348. These GABAB receptor mediated effects were replicated in DBA/2J inbred mice that exhibit extreme sensitivity to ethanol's stimulant effects. In summary, we found moderate to strong evidence that some sites on the GABAA receptor complex were altered as a consequence of selection of FAST and SLOW mice, but found little support for GABAA mediation of EtOH-stimulated activity. In contrast, we found moderate evidence for differential alteration of GABAB receptor function; however, GABAB receptor involvement in ethanol-stimulated activity was strongly supported by results in the selected lines and an inbred strain.     

GABAergic drugs and sexual motivation, receptivity and exploratory behaviors in the female rat

Female rats were allowed to pace sexual interactions in a bilevel chamber, where a sexually vigorous male was tethered to the bottom level. Exploratory behaviors (sniffing, rearing), locomotor activity (expressed as number of level changes and periods of inactivity) as well as items of sexual motivation (latency to descend to the male's level, approaches towards the male and genital exploration) were recorded. In addition, sexual receptivity was evaluated in a non-paced situation. A test for motor impairment was also performed. The GABA transaminase inhibitor gamma-acetylene GABA reduced exploratory behaviors at doses much lower than those needed to reduce receptivity. The GABA reuptake inhibitor SKF 100330A did not affect any behavior category at doses of 15 and 30 mg/kg, but had a sedative action at 60 mg/kg. This was shown as impaired motor coordination and an almost total absence of activity in the bilevel chamber. Receptivity was not impaired, however. The mixed GABAA/ GABAB agonist progabide reduced exploratory behaviors and receptivity without producing motor impairment at a dose of 400 mg/kg. The GABAA agonist THIP impaired motor coordination and reduced receptivity and exploratory behaviors at a dose of 10 mg/kg. A larger dose, 20 mg/kg, had a strong sedative action. Only a small proportion of the animals descended to the males level. The GABAB agonist baclofen reduced receptivity at a dose that had no effect on motor coordination or exploratory behaviors. None of the drugs had a specific effect on sexual motivation. Whenever behaviors reflecting motivation were reduced, there were also other behavioral effects indicative of sedation. These data show that GABA receptor agonists, particularly the GABAB agonist baclofen, reduce sexual receptivity at doses that have only slight effect on motor functions or exploratory behaviors. In contrast, non-specific enhancement of GABAergic activity by a transaminase or reuptake inhibitor have effects on motor functions and exploratory behaviors at doses much lower than those needed to reduce receptivity.     

Functional GABAA receptors on rat vagal afferent neurones

1. In the present study, in vitro electrophysiology and receptor autoradiography were used to determine whether rat vagal afferent neurones possess gamma-aminobutyric acid (GABA)A receptors. 2. GABA (1-100 microM) and isoguvacine (3-100 microM) caused a concentration-dependent depolarization of the rat isolated nodose ganglion preparation at room temperature. When applied to the tissue 20 min before the agonist, SR95531 (3 microM) and bicuculline (3 microM) caused a parallel shift to the right of the GABA and isoguvacine concentration-response curves, yielding shifts of 81 fold and 117 fold for SR95531 and 4 fold and 12 fold for bicuculline, respectively. 3. Baclofen (10 nM-100 microM) was unable to elicit a depolarization of the rat isolated nodose ganglion preparation at either room temperature or at 36 degrees C, whilst 5-aminovaleric acid (10 microM), a GABAB receptor antagonist, was unable to antagonize significantly the GABA-induced depolarization at either room temperature or at 36 degrees C. 4. [3H]-SR95531 (7.2 nM), a GABAA receptor-selective antagonist, bound topographically to sections of rat brainstem. Specific binding was highest in the medial nucleus tractus solitarius (NTS) and dorsal motor nucleus of the vagus nerve (DMVN). Binding was also observed in certain medullary reticular nuclei, in particular the parvocellular reticular nucleus. 5. Unilateral nodose ganglionectomy caused a reduction in GABAA binding site density in the medial NTS from 93 +/- 7 to 68 +/- 6 d.p.m./mm2. This procedure also caused a reduction in GABAA binding site density in the side of the NTS contralateral to the lesion, from 151 +/- 12 to 93 +/- 7 d.p.m./mm2. Sham surgery had no effect on the binding of [3H]-SR95531 in rat brainstem. 6. The present data provide evidence for the presence of GABAA receptors located on the soma and central terminals of rat vagal afferent neurones. Additionally, a population of GABAA receptors is evidenced postsynaptically in the rat NTS with respect to vagal afferent terminals. These data are discussed in relation to the functional pharmacology of GABA in this region of the NTS.     

GABA induces Ca2+ transients in astrocytes

By using the Ca(2+)-sensitive indictor Fura-2/AM, the cytosolic Ca2+ levels [Ca2+]i were measured in type 1 astrocytes in rat cortical astroglial primary cultures, after stimulation with GABA, muscimol (GABAA agonist), or baclofen (GABAB agonist). We report the first evidence that stimulation of both GABAA and GABAB receptors evokes Ca2+ transients in type I astrocytes. Two types of Ca2+ responses were seen: the single-phase curve, which was the most common, and the biphasic, which consisted of an initial rise that persisted at the maximal or submaximal level. Both types of Ca2+ responses appeared with some latency. The responses were obtained in astrocytes grown for 12-16 days in culture and the response frequencies for all three agonists were 18% of the total number of examined cells. However, when the astrocytes were grown in a mixed astroglial/neuronal culture the response frequencies for all three agonists increased to 35% of the total number of examined cells. In some cells, the responses after GABA stimulation were blocked to baseline levels after exposure to bicuculline (GABAA antagonist). In other cells, bicuculline only slightly reduced the GABA-evoked responses, and the addition of phaclofen (GABAB antagonist) did not potentiate this partial inhibition. However, the muscimol-evoked rises in [Ca2+]i were completely inhibited after exposure to bicuculline, while the responses after baclofen could only be partly blocked by phaclofen. GABA evoked rises in [Ca2+]i which alternatively were inhibited (mostly) or persisted in Ca(2+)-free buffer. The rises in [Ca2+]i persisted, but were reduced, in Ca(2+)-free buffer after stimulation with muscimol, but were inhibited after baclofen stimulation. The GABA uptake blockers guvacine, 4,5,6,7-tetrahydroisoxazolo(4,5-c)pyridin-3-ol and nipecotic acid were also able to reduce the GABA-evoked rises in [Ca2+]i. However, the L-type Ca2+ channel antagonist nifedipine failed to influence on the GABA-evoked Ca2+ transients. The results suggest that type 1 astrocytes in primary culture express GABA receptors which can elevate [Ca2+]i directly or indirectly via Ca2+ channels and/or via release from internal Ca2+ stores. The results also suggest that GABA can have intracellular Ca(2+)-mobilizing sites since the GABA-evoked responses were reduced after incubation with GABA uptake blockers.     

Imaging the induction and spread of seizure activity in the isolated brain of the guinea pig: the roles of GABA and glutamate receptors

1. The induction and spread of seizure activity was studied using imaging and electrophysiological techniques in the isolated whole brain of the guinea pig. We examined the role of GABA and glutamate receptor subtypes in controlling the spread of seizure activity across the olfactory cortex from a focus in the entorhinal cortex. Seizure spread was monitored by video imaging of intrinsic optical signals (reflectance changes) combined with multiple extracellular recordings. Both the unilateral and bilateral spread of seizure activity was monitored in different experiments. 2. Electrical stimulation of the lateral entorhinal cortex (10-15 V, 5 Hz, 5-10 s) evoked seizure activity that originated in the entorhinal cortex/hippocampus and later spread preferentially toward the posteromedial cortical amygdaloid nucleus ipsilaterally and bilaterally. The pattern of seizure spread in a given brain was highly reproducible. 3. The influence of gamma-aminobutyric acid (GABA) receptors on the spread of seizure activity was monitored at higher resolution on one side of the brain. Perfusion of a low concentration of the GABAA antagonist bicuculline methiodide (20 microM) resulted in spontaneous seizures that spread to the posteromedial cortical amygdaloid nucleus more rapidly than electrically evoked seizures [spread times: 5.5 +/- 3.7 s vs. 15.5 +/- 2.7 s, respectively (means +/- SE)]. Seizure spread was also more extensive in the presence of bicuculline involving the posterior perirhinal cortex and larger areas over the medial amygdala. Higher concentrations of bicuculline (100 microM) resulted in even more widespread propagation of spontaneous seizure activity throughout the olfactory cortex as well as to the perirhinal, insular, and occipital cortices. This concentration of bicuculline also further reduced the time required for seizure activity to spread from the entorhinal cortex to the posteromedial cortical amygdaloid nucleus (spread time = 2.3 +/- 1.7 s). The GABAB antagonist, CGP 35348 (200 microM), in contrast, had no significant effect of seizure induction or propagation. 4. The role of glutamate receptor subtypes in seizure propagation was studied by examining the bilateral spread of seizures. Perfusion of the kainate/alpha-amino-3-hydroxy-5-methyl-4-isoxazole proprionic acid (K/A) receptor antagonist (6-cyano-7-nitroquinoxaline-2,3-dione, CNQX, 20 microM) completely and reversibly suppressed stimulus-evoked seizure activity as detected electrophysiologically and optically. CNQX also reduced the magnitudes of field potentials recorded in the isolated brain in a reversible manner by an average of 70.8 +/- 2.21% of control. The N-methyl-D-aspartate (NMDA) receptor antagonist dibenzocyclohepteneimine (MK-801) did not significantly alter the magnitudes or shapes of field potentials recorded in the isolated brain nor did it significantly alter seizure activity measured optically or electrophysiologically. 5. Perfusion of the metabotropic glutamate receptor agonist [trans-1-amino-(IS,3R)-cyclopentanedicarboxylic acid (trans-ACPD), 150 microM] completely and reversibly suppressed stimulus-evoked seizure activity as detected electrophysiologically and optically. The magnitudes of field potentials recorded in the isolated brain also were reduced by trans-ACPD an average of 75.4 +/- 5.39% of control values. 6. These results demonstrate that GABAA-mediated transmission is functionally present and may play an important role in epileptic tissue in limiting the spread of seizure activity from the entorhinal cortex to the posteromedial cortical amygdaloid nucleus and in creating functional pathways or preferential routes of seizure spread. GABAB-mediated postsynaptic inhibition played no significant role in the induction or spread of seizure activity in this study. K/A receptors but not NMDA receptors are necessary for the induction and subsequent spread of seizure activity originating in the entorhinal cortex/hippocampus.     

Inhibition in the human urinary bladder by gamma-amino-butyric acid

OBJECTIVE: To investigate the effects of gamma-amino-butyric acid (GABA) on detrusor activity in man to determine whether it has any inhibitory effect on detrusor contraction. The inhibitory neurotransmitter GABA has been found in mammalian urinary bladders and the effects of GABA on detrusor activity in the rabbit bladder has previously been described [1]. MATERIALS AND METHODS: Human detrusor muscle strips, obtained at cystectomy, were made to contract by electrical stimulation of their autonomic nerves or by the addition of carbachol in a superfusion apparatus. GABA and its analogues were added to the superfusion chamber and any changes in the responses were measured. RESULTS: The electrically evoked nerve-mediated contractions in human bladder muscle were exclusively cholinergic. GABA inhibited nerve-mediated contractions in human detrusor muscle-strips by the activation of the GABAB receptor, since baclofen (a GABAB receptor agonist) produced similar inhibition and muscimol (a GABAA receptor agonist) did not. There was no inhibition of carbachol-mediated contractions by GABA. CONCLUSION: This in vitro study shows that GABA has a peripherally mediated inhibitory effect on excitatory neurotransmission in human detrusor muscle. The site of action is on the post-ganglionic nerves and appears to be mediated via the GABAB receptor.