Propofol anaesthesia in mice is potentiated by muscimol and reversed by bicuculline.
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We have examined the role of gamma-aminobutyric acid (GABA) neurones in propofol anaesthesia in mice using the righting reflex. Propofol i.p. increased the percentage of loss of the righting reflex in a dose-dependent manner with an ED50 value of 140 (95% confidence limits 123-160) mg kg-1 (n = 40; eight animals per dose, five doses per dose-response curve). The ED50 for propofol decreased significantly to 66 (58-75) mg kg-1 in the presence of the GABAA receptor agonist muscimol 1 mg kg-1 i.p. (n = 40) (P < 0.05). In contrast, the ED50 increased significantly to 240 (211-274) mg kg-1 in the presence of the antagonist bicuculline 5 mg kg-1 i.p. (n = 40) (P < 0.05). Our results suggest that propofol anaesthesia may be mediated, at least in part by GABA neurons. (+info)
Afferent regulation of inhibitory synaptic transmission in the developing auditory midbrain.
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To determine whether afferent innervation regulates the strength of inhibitory connections in the gerbil auditory midbrain, both cochleas were surgically removed in postnatal day 7 animals, before sound-driven activity is first observed. Inhibitory synaptic currents were measured in a brain slice preparation 1-7 d after the ablations. Whole-cell and gramicidin-perforated patch recordings were obtained from inferior colliculus neurons, and IPSCs were evoked by stimulation of the commissure of the inferior colliculus (CIC) or the ipsilateral lateral lemniscus (LL) in the presence of kynurenic acid. Deafferentation led to a 24 mV depolarizing shift in the IPSC equilibrium potential within 1 d of deafferentation. As a consequence, there was a large reduction of IPSC amplitude at a holding potential of -20 mV in neurons from bilaterally ablated animals. Furthermore, both afferent pathways displayed a 50% reduction of the inhibitory synaptic conductance after deafferentation, indicating that driving force was not solely responsible for the decline in IPSC amplitude. When paired pulses were delivered to the LL or CIC pathway in control neurons, the evoked IPSCs exhibited facilitation. However, paired pulse facilitation was nearly eliminated after deafferentation. Thus, normal innervation affects inhibitory synaptic strength by regulating postsynaptic chloride homeostasis and presynaptic transmitter release properties. (+info)
Three levels of lateral inhibition: A space-time study of the retina of the tiger salamander.
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The space-time patterns of activity generated across arrays of retinal neurons can provide a sensitive measurement of the effects of neural interactions underlying retinal activity. We measured the excitatory and inhibitory components associated with these patterns at each cellular level in the retina and further dissected inhibitory components pharmacologically. Using perforated and loose patch recording, we measured the voltages, currents, or spiking at 91 lateral positions covering approximately 2 mm in response to a flashed 300-microm-wide bar. First, we showed how the effect of well known lateral inhibition at the outer retina, mediated by horizontal cells, evolved in time to compress the spatial representation of the stimulus bar at ON and OFF bipolar cell bodies as well as horizontal cells. Second, we showed, for the first time, how GABA(C) receptor mediated amacrine cell feedback to bipolar terminals compresses the spatial representation of the stimulus bar at ON bipolar terminals over time. Third, we showed that a third spatiotemporal compression exists at the ganglion cell layer that is mediated by feedforward amacrine cells via GABA(A) receptors. These three inhibitory mechanisms, via three different receptor types, appear to compensate for the effects of lateral diffusion of activity attributable to dendritic spread and electrical coupling between retinal neurons. As a consequence, the width of the final representation at the ganglion cell level approximates the dimensions of the original stimulus bar. (+info)
Neuroactive steroid 3alpha-hydroxy-5alpha-pregnan-20-one modulates electrophysiological and behavioral actions of ethanol.
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Neuroactive steroids are synthesized de novo in brain, yet their physiological significance remains elusive. We provide biochemical, electrophysiological, and behavioral evidence that several specific actions of alcohol (ethanol) are mediated by the neurosteroid 3alpha-hydroxy-5alpha-pregnan-20-one (3alpha,5alpha-THP; allopregnanolone). Systemic alcohol administration elevates 3alpha, 5alpha-THP levels in the cerebral cortex to pharmacologically relevant concentrations. The elevation of 3alpha,5alpha-THP is dose- and time-dependent. Furthermore, there is a significant correlation between 3alpha,5alpha-THP levels in cerebral cortex and the hypnotic effect of ethanol. Blockade of de novo biosynthesis of 5alpha-reduced steroids using the 5alpha-reductase inhibitor finasteride prevents several effects of ethanol. Pretreatment with finasteride causes no changes in baseline bicuculline-induced seizure threshold but reverses the anticonvulsant effect of ethanol. Finasteride pretreatment also reverses ethanol inhibition of spontaneous neural activity in medial septal/diagonal band of Broca neurons while having no direct effect on spontaneous firing rates. Thus, elevation of 3alpha,5alpha-THP levels by acute ethanol administration represents a novel mechanism of ethanol action as well as an important modulatory role for neurosteroids in the CNS. (+info)
Long-lasting depolarizations in mitral cells of the rat olfactory bulb.
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We investigated the mechanisms of long-lasting depolarizing potentials (LLDs) generated in mitral cells with whole-cell patch recordings in the rat olfactory bulb slice. LLDs occur spontaneously and are evoked by either orthodromic stimulation of the olfactory nerve or antidromic stimulation of mitral and tufted (M/T) cells. LLDs are followed by a long refractory period, limiting LLD generation to approximately 1 Hz. LLD production does not appear to involve either intrinsic voltage-activated or metabotropic mechanisms. The initiation of LLDs requires activation of non-NMDA but not NMDA receptors. Dual recordings from the apical dendrites and somata of mitral cells show that LLDs are generated in the distal portion of the apical dendrite, most likely in the glomerulus. The rising phase of LLDs shows characteristics of polyneuronal input, including a high variability and sensitivity to charge screening. Paired recordings from adjacent mitral cells suggest that LLDs occur synchronously only in cells whose apical dendrites ramify in the same glomerulus. These findings suggest that LLDs involve recurrent, intraglomerular dendrodendritic interactions among M/T cells. (+info)
Modulation by bicuculline and penicillin of the block by t-butyl-bicyclo-phosphorothionate (TBPS) of GABA(A)-receptor mediated Cl(-)-current responses in rat striatal neurones.
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1. T-butyl-bicyclo-phosphorothionate (TBPS) is a prototypical representative of the cage-convulsants which act through a use-dependent block of the GABA(A)-receptor-ionophore complex. Using current recordings from cultured neurones of rat striatum the manner was investigated in which two antagonists, bicuculline and penicillin, presumably acting at the agonist binding site and in the ionic channel, respectively, modify the rate of block by TBPS. 2. Penicillin (5 or 10 mM) did not slow the rate of block by TBPS, but produced a significant enhancement of block rate, which, however, was inversely related to the degree of antagonism by penicillin of the GABA-induced current. 3. Bicuculline (10 microM) reduced the rate of block by TBPS. However, this effect was 3 fold weaker than its GABA-antagonistic action. The slowing of block rate and the current antagonism exhibited a biphasic, positive-negative relationship. Co-application of bicuculline (100 microM) in a concentration that produced nearly complete antagonism and TBPS (10 microM) resulted in a marked ( approximately 40%) reduction of subsequent GABA response amplitudes compatible with a direct, bicuculline-induced conformational change in the receptor required for the binding of and block by TBPS. 4. The lack of protection afforded by the channel blocker penicillin as well as the lack of correlation between bicuculline antagonism of the Cl(-)-current and its efficiency in protecting against TBPS block is evidence against an open channel blocking mechanism for TBPS. TBPS does, therefore, not appear to gain access to its binding site via the open pore but through alternative routes regulated from the agonist binding site. (+info)
Pharmacological characterization of small-conductance Ca(2+)-activated K(+) channels stably expressed in HEK 293 cells.
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Three genes encode the small-conductance Ca(2+)-activated K(+) channels (SK channels). We have stably expressed hSK1 and rSK2 in HEK 293 cells and addressed the pharmacology of these subtypes using whole-cell patch clamp recordings. The bee venom peptide apamin blocked hSK1 as well as rSK2 with IC(50) values of 3.3 nM and 83 pM, respectively. The pharmacological separation between the subtypes was even more prominent when applying the scorpion peptide blocker scyllatoxin, which blocked hSK1 with an IC(50) value of 80 nM and rSK2 at 287 pM. The potent small molecule blockers showed little differentiation between the channel subtypes. The bis-quinolinium cyclophane UCL 1684 blocked hSK1 with an IC(50) value of 762 pM and rSK2 at 364 pM. The antiseptic compound dequalinium chloride blocked hSK1 and rSK2 with IC(50) values of 444 nM and 162 nM, respectively. The nicotinic acetylcholine receptor antagonist d-tubocurarine was found to block hSK1 and rSK2 with IC(50) values of 27 microM and 17 microM when measured at +80 mV. The inhibition by d-tubocurarine was voltage-dependent with increasing affinities at more hyperpolarized potentials. The GABA(A) receptor antagonist bicuculline methiodide also blocked hSK1 and rSK2 in a voltage-dependent manner with IC(50) values of 15 and 25 microM when measured at +80 mV. In conclusion, the pharmacological separation between SK channel subtypes expressed in mammalian cells is too small to support the notion that the apamin-insensitive afterhyperpolarization of neurones is mediated by hSK1. (+info)
Novel role for the NMDA receptor redox modulatory site in the pathophysiology of seizures.
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Redox-active compounds modulate NMDA receptors (NMDARs) such that reduction of NMDAR redox sites increases, and oxidation decreases, NMDAR-mediated activity. Because NMDARs contribute to the pathophysiology of seizures, redox-active compounds also may modulate seizure activity. We report that the oxidant 5, 5'-dithio-bis(2-nitrobenzoic acid) (DTNB) and the redox cofactor pyrroloquinoline quinone (PQQ) suppressed low Mg(2+)-induced hippocampal epileptiform activity in vitro. Additionally, in slices exposed to 4-7 microM bicuculline, DTNB and PQQ reversed the potentiation of evoked epileptiform responses by the reductants dithiothreitol and Tris(2-carboxyethyl)phosphine (TCEP). NMDA-evoked whole-cell currents in CA1 neurons in slices were increased by TCEP and subsequently decreased by DTNB or PQQ at the same concentrations that modulated epileptiform activity. However, DTNB and PQQ had little effect on baseline NMDA-evoked currents in control medium, and PQQ did not alter NMDAR-dependent long-term potentiation. In contrast, in slices returned to control medium after low Mg(2+)-induced ictal activity, DTNB significantly inhibited NMDAR-mediated currents, indicating endogenous reduction of NMDAR redox sites under this epileptogenic condition. These data suggested that PQQ and DTNB suppressed spontaneous ictal activity by reversing pathological NMDAR redox potentiation without inhibiting physiological NMDAR function. In vivo, PQQ decreased the duration of chemoconvulsant-induced seizures in rat pups with no effect on baseline behavior. Our results reveal endogenous potentiation of NMDAR function via mass reduction of redox sites as a novel mechanism that may enhance epileptogenesis and facilitate the transition to status epilepticus. The results further suggest that redox-active compounds may have therapeutic use by reversing NMDAR-mediated pathophysiology without blocking physiological NMDAR function. (+info)