The specific mechanisms underlying general anesthesia are primarily unknown. The intravenous general anesthetic etomidate acts by potentiating GABA(A) receptors, with selectivity for beta2 and beta3 subunit-containing receptors determined by a single asparagine residue. We generated a genetically modified mouse containing an etomidate-insensitive beta2 subunit (beta2 N265S) to determine the role of beta2 and beta3 subunits in etomidate-induced anesthesia. Loss of pedal withdrawal reflex and burst suppression in the electroencephalogram were still observed in the mutant mouse, indicating that loss of consciousness can be mediated purely through beta3-containing receptors. The sedation produced by subanesthetic doses of etomidate and during recovery from anesthesia was present only in wild-type mice, indicating that the beta2 subunit mediates the sedative properties of anesthetics. These findings show that anesthesia and sedation are mediated by distinct GABA(A) receptor subtypes.

does not interact with N-type calcium channels directly but inhibits them via a voltage-independent mechanism involving a PTX-sensitive, G-protein-coupled receptor. Preincubation with a variety of selective receptor antagonists demonstrated that only the GABA B receptor antagonists, [S-(R*,R*)][-3-[[1-(3,4-dichlorophenyl)ethyl]amino]-2-hydroxy propyl]([3,4]-cyclohexylmethyl) phosphinic acid hydrochloride (2S)-3[[(1S)- 1-(3,4-dichlorophenyl

Neuronal (N)-type Ca2ϩ channel-selective -conotoxins have emerged as potential new drugs for the treatment of chronic pain. In this study, two new -conotoxins, CVIE and CVIF, were discovered from a Conus catus cDNA library. Both conopeptides potently displaced 125 I-GVIA binding to rat brain membranes. In Xenopus laevis oocytes, CVIE and CVIF potently and selectively inhibited depolarizationactivated Ba 2ϩ currents through recombinant N-type (␣1 B-b / ␣ 2 ␦1/␤ 3 ) Ca 2ϩ channels. Recovery from block increased with membrane hyperpolarization, indicating that CVIE and CVIF have a higher affinity for channels in the inactivated state. The link between inactivation and the reversibility of -conotoxin action was investigated by creating molecular diversity in ␤ subunits: N-type channels with ␤ 2a subunits almost completely recovered from CVIE or CVIF block, whereas those with ␤ 3 subunits exhibited weak recovery, suggesting that reversibility of the -conotoxin block may depend on the type of ␤-subunit isoform. In rat dorsal root ganglion sensory neurons, neither peptide had an effect on low-voltageactivated T-type channels but potently and selectively inhibited high voltage-activated N-type Ca 2ϩ channels in a voltage-dependent manner. In rat spinal cord slices, both peptides reversibly inhibited excitatory monosynaptic transmission between primary afferents and dorsal horn superficial lamina neurons. Homology models of CVIE and CVIF suggest that -conotoxin/voltage-gated Ca 2ϩ channel interaction is dominated by ionic/electrostatic interactions. In the rat partial sciatic nerve ligation model of neuropathic pain, CVIE and CVIF (1 nM) significantly reduced allodynic behavior. These N-type Ca 2ϩ channel-selective -conotoxins are therefore useful as neurophysiological tools and as potential therapeutic agents to inhibit nociceptive pain pathways.

. By contrast, mIPSCs were not influenced by deletion of these genes. Enhancement of the tonic current by the β 2/3 subunit-selective agent etomidate was significantly reduced for DGGCs derived from β 2N265S mice, whereas this manipulation had no effect on the prolongation of mIPSCs produced by this anaesthetic. Collectively, these observations, together with previous studies on α 4 −/− mice, identify a population of extrasynaptic α 4 β 2 δ receptors, whereas synaptic GABA A receptors appear to primarily incorporate the β 3 subunit. A component of the tonic current is diazepam sensitive and is mediated by extrasynaptic receptors incorporating α 5 and γ 2 subunits. Deletion of the β 2 subunit had no effect on the diazepam-induced current and therefore these extrasynaptic receptors do not contain this subunit. The unambiguous identification of these distinct pools of synaptic and extrasynaptic GABA A receptors should aid our understanding of how they act in harmony, to regulate hippocampal signalling in health and disease.

This study examined the effect of chronic exposure to ethanol and brain-derived neurotrophic factor (BDNF) on the responsiveness of cerebellar granule cells to gamma-aminobutyric acid (GABA). Cerebellar granule cell cultures were chronically exposed to ethanol (100 mM), BDNF (20 ng/ml), or the combination of ethanol and BDNF. Whole-cell current responses of granule cells to exogenously applied GABA were monitored following at least 5 days of chronic exposure. In the ethanol-treated cultures, granule cell responsiveness to GABA was attenuated. Concomitant exposure of cultures to ethanol and BDNF mitigated the ethanol-induced attenuation of GABA response, although BDNF, by itself, did not affect responsiveness to GABA. BDNF increased the expression of the GABA(A) receptor alpha6 subunit, whereas ethanol had no effect, in chronically treated granule cell cultures. In addition, concomitant treatment with BDNF and ethanol did not increase the expression of the GABA(A) receptor alpha6 subunit, so the subunit expression alone could not account for the mitigating effect of BDNF. We propose that different mechanisms regulating responsiveness to GABA underlie the effects induced by ethanol and BDNF, with the former influencing the expression of functional GABA(A) receptors and the latter involving the activation of the TrkB receptor and its downstream signaling pathways.

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