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The effective management of marine species relies on a comprehensive understanding of how the environment can influence biodiversity and distributions. This study quantified the biodiversity of invertebrates across the phyla Arthropoda, Echinodermata, Mollusca and Porifera from 132 sampling stations along the coast of the Ningaloo Marine Park (NMP) and across depths ranging from 30 to 123 m, providing a broad assessment of the biodiversity of marine invertebrates in mesophotic habitats of the NMP. Benthic sleds were used to sample biodiversity and specimens identified to the highest taxonomic resolution. Single-beam and multibeam surveys of the seabed were performed to derive 58 spatial and bathymetric variables used to assess their relevance to the distributions of benthic diversity. A total of 787 taxa were identified across the four phyla, of which 51% were rare and sampled from a single station. Latitude and depth were significant in structuring the invertebrate diversity in the mesophotic, with Point Cloates notably identified as a significant geographical feature in partitioning diversity to the north and south of the NMP. Depth was more influential at structuring diversity in the northern section of the NMP due to the narrower Ningaloo Shelf, and southern localities including Turtles and Red Bluff were identified as biodiversity hotspots for benthic invertebrates. Importantly, comparisons of diversity within ‘no-take’ and ‘general use’ zones along the NMP indicate that sanctuary zones represent the diversity contained in the marine park, thus highlighting their importance in the protection and conservation of invertebrate diversity for the region.

A new species of sponge-inhabiting barnacle, Membranobalanus porphyrophilussp. nov., is described herein. This species can be distinguished from all other congeners by a combination of characters, in particular by the shapes of the tergum and scutum and the armament of the cirri. COI sequence data from the type specimens have been lodged with GenBank and a morphological key to the species of Membranobalanus is provided to aid future research. The host of the new species is the southern Australian endemic demosponge Spheciospongia purpurea. The new species of barnacle is thought to be host species specific.

Sponges represent one of the most species-rich hosts for commensal barnacles yet host utilisation and diversity have not been thoroughly examined. This study investigated the diversity and phylogenetic relationships of sponge-inhabiting barnacles within a single, targeted host group, primarily from Western Australian waters. Specimens of the sponge order Dictyoceratida were surveyed and a total of 64 host morphospecies, representing four families, were identified as barnacle hosts during the study. Utilising molecular (COI, 12S) and morphological methods 42 molecular operational taxonomic units (MOTUs) of barnacles, representing Acasta, Archiacasta, Euacasta and Neoacasta were identified. Comparing inter- and intra-MOTU genetic distances showed a barcode gap between 2.5% and 5% for COI, but between 1% and 1.5% in the 12S dataset, thus demonstrating COI as a more reliable barcoding region. These sponge-inhabiting barnacles were demonstrated to show high levels of host specificity with the majority being found in a single sponge species (74%), a single genus (83%) or a single host family (93%). Phylogenetic relationships among the barnacles were reconstructed using mitochondrial (12S, COI) and nuclear (H3, 28S) markers. None of the barnacle genera were recovered as monophyletic. Euacasta was paraphyletic in relation to the remaining Acastinae genera, which were polyphyletic. Six well-supported clades of molecular operational taxonomic units, herein considered to represent species complexes, were recovered, but relationships between them were not well supported. These complexes showed differing patterns of host usage, though most were phylogenetically conserved with sister lineages typically occupying related hosts within the same genus or family of sponge. The results show that host specialists are predominant, and the dynamics of host usage have played a significant role in the evolutionary history of the Acastinae.

A new species, Paranaxia keesingi sp. nov., is described based on specimens collected in northern Australia. The new species differs from its only congener, P. serpulifera (Guérin, 1832, in Guérin-Méneville 1829-1837), by several characters including carapace setation, sternal cavities, absence of a subhepatic spine, presence of a sharp spine on the posterodistal angle of the cheliped merus, relatively shorter chelipeds, and longer and more slender ambulatory legs. Morphological separation of the two species is supported by 12s rDNA sequence divergences of 7.4-8.2%. Like P. serpulifera, the newly described species exhibits direct development with females carrying juvenile individuals under the pleon. Both species are sympatric, but Paranaxia keesingi sp. nov. is found in deeper waters than P. serpulifera.

Bioavailable contaminant concentrations are an important component in assessing environmental effects as they directly affect ecosystem health. Shellfish contaminant monitoring programmes have traditionally filled this requirement but are being phased out in some jurisdictions. Passive sampling devices (PSDs) have the potential to replace shellfish monitoring; however, there are still knowledge gaps to address before this can occur. This study assessed the suitability of three different PSDs in providing the required information to replace shellfish monitoring. PSDs were deployed at three historic mussel monitoring sites with different levels of urban influence in the Waitemata Harbour, Auckland, New Zealand. Contaminants of interest were urban heavy metals, plus current and emerging organic contaminants. PSDs provided extremely low detection limits and, for some contaminants, very strong correlations to shellfish. PSDs can currently complement shellfish in monitoring, but it is premature to make conclusions as to the suitability of PSDs in replacing shellfish monitoring until more information is available.

Controlling neuronal excitability is vitally important for maintaining a healthy central nervous system (CNS) and this relies on the activity of type A gamma-aminobutyric acid (GABA(A)) neurotransmitter receptors. Given this role, it is therefore important to understand how these receptors are regulated by endogenous modulators in the brain and determine where they bind to the receptor. One of the most potent groups of modulators is the neurosteroids which regulate the activity of synaptic and extrasynaptic GABAA receptors. This level of regulation is thought to be physiologically important and its dysfunction may be relevant to numerous neurological conditions. The aim of this review is to summarise those studies that over the last 20 years have focussed upon finding the binding sites for neurosteroids on GABA(A) receptors. We consider the nature of steroid binding sites in other proteins where this has been determined at atomic resolution and how their generic features were mapped onto GABAA receptors to help locate 2 putative steroid binding sites. Altogether, the findings strongly suggest that neurosteroids do bind to discrete sites on the GABAA receptor and that these are located within the transmembrane domains of of and receptor subunits. The implications for neurosteroid binding to other inhibitory receptors such as glycine and GABA(C) receptors are also considered. Identifying neurosteroid binding sites may enable the precise pathophysiological role(s) of neurosteroids in the CNS to be established for the first time, as well as providing opportunities for the design of novel drug entities. (c) 2007 Elsevier Inc. All rights reserved.

Phosphorylation can affect both the function and trafficking of GABA(A) receptors with significant consequences for neuronal excitability. Serine/threonine kinases can phosphorylate the intracellular loops between M3-4 of GABA(A) receptor beta and gamma subunits thereby modulating receptor function in heterologous expression systems and in neurons (1, 2). Specifically, CaMK-II has been demonstrated to phosphorylate the M3-4 loop of GABA(A) receptor subunits expressed as GST fusion proteins (3, 4). It also increases the amplitude of GABA(A) receptor-mediated currents in a number of neuronal cell types (5-7). To identify which substrate sites CaMK-II might phosphorylate and the consequent functional effects, we expressed recombinant GABA(A) receptors in NG108-15 cells, which have previously been shown to support CaMK-II modulation of GABA(A) receptors containing the beta3 subunit (8). We now demonstrate that CaMK-II mediates its effects on alpha1beta3 receptors via phosphorylation of Ser(383) within the M3-4 domain of the beta subunit. Ablation of beta3 subunit phosphorylation sites for CaMK-II revealed that for alphabetagamma receptors, CaMK-II has a residual effect on GABA currents that is not mediated by previously identified sites of CaMK-II phosphorylation. This residual effect is abolished by mutation of tyrosine phosphorylation sites, Tyr(365) and Tyr(367), on the gamma2S subunit, and by the tyrosine kinase inhibitor genistein. These results suggested that CaMK-II is capable of directly phosphorylating GABA(A) receptors and activating endogenous tyrosine kinases to phosphorylate the gamma2 subunit in NG108-15 cells. These findings were confirmed in a neuronal environment by expressing recombinant GABA(A) receptors in cerebellar granule neurons.

Inhibitory neurotransmission mediated by GABA(A) receptors can be modulated by the endogenous neurosteroids, allopregnanolone and tetrahydro-deoxycorticosterone(1). Neurosteroids are synthesized de novo in the brain during stress(2), pregnancy(3) and after ethanol consumption(4), and disrupted steroid regulation of GABAergic transmission is strongly implicated in several debilitating conditions such as panic disorder, major depression, schizophrenia, alcohol dependence and catamenial epilepsy(3,5-8). Determining how neurosteroids interact with the GABA(A) receptor is a prerequisite for understanding their physiological and pathophysiological roles in the brain. Here we identify two discrete binding sites in the receptor's transmembrane domains that mediate the potentiating and direct activation effects of neurosteroids. They potentiate GABA responses from a cavity formed by the alpha-subunit transmembrane domains, whereas direct receptor activation is initiated by interfacial residues between alpha and beta subunits and is enhanced by steroid binding to the potentiation site. Thus, significant receptor activation by neurosteroids relies on occupancy of both the activation and potentiation sites. These sites are highly conserved throughout the GABA(A) receptor family, and their identification provides a unique opportunity for the development of new therapeutic, neurosteroid-based ligands and transgenic disease models of neurosteroid dysfunction.

Regulation of GABAA receptors by extracellular pH exhibits a dependence on the receptor subunit composition. To date, the molecular mechanism responsible for the modulation of GABAA receptors at alkaline pH has remained elusive. We report here that the GABA-activated current can be potentiated at pH 8.4 for both αβ and αβγ subunit-containing receptors, but only at GABA concentrations below the EC40. Site-specific mutagenesis revealed that a single lysine residue, K279 in the β subunit TM2–TM3 linker, was critically important for alkaline pH to modulate the function of both α1β2 and α1β2γ2 receptors. The ability of low concentrations of GABA to reveal different pH titration profiles for GABAA receptors was also examined at acidic pH. At pH 6.4, GABA activation of αβγ receptors was enhanced at low GABA concentrations. This effect was ablated by the mutation H267A in the β subunit. Decreasing the pH further to 5.4 inhibited GABA responses via αβγ receptors, whereas those responses recorded from αβ receptors were potentiated. Inserting homologous β subunit residues into the γ2 subunit to recreate, in αβγ receptors, the proton modulatory profile of αβ receptors, established that in the presence of β2H267, the mutation γ2T294K was necessary to potentiate the GABA response at pH 5.4. This residue, T294, is homologous to K279 in the β subunit and suggests that a lysine at this position is an important residue for mediating the allosteric effects of both acidic and alkaline pH changes, rather than forming a direct site for protonation within the GABAA receptor.

Zinc ions are concentrated in the central nervous system and regulate GABA(A) receptors, which are pivotal mediators of inhibitory synaptic neurotransmission. Zinc ions inhibit GABA(A) receptor function by an allosteric mechanism that is critically dependent on the receptor subunit composition: alphabeta subunit combinations show the highest sensitivity, and alphabetagamma isoforms are the least sensitive. Here we propose a mechanistic and structural basis for this inhibition and its dependence on the receptor subunit composition. We used molecular modeling to identify three discrete sites that mediate Zn2+ inhibition. One is located within the ion channel, and the other two are on the external amino (N)-terminal face of the receptor at the interfaces between alpha and beta subunits. We found that the characteristically low Zn2+ sensitivity of GABA(A) receptors containing the gamma2 subunit results from disruption to two of the three sites after receptor subunit co-assembly.

PMBA is a novel antagonist of strychnine-sensitive glycine receptors in the rat spinal cord, however, its mode of action is unknown. The actions of PMBA on rat glycine receptor α1 and α2 homomers in Xenopus oocytes were studied under two-electrode voltage-clamp. Co-application of PMBA and glycine to both α1 and α2 homomers yielded inward currents which decayed to a steady-state. Responses rose slowly to the same steady-state amplitude following a 2 min pre-incubation in PMBA. Strychnine, but not picrotoxinin, showed similar antagonism to PMBA. The potency of PMBA was independent of membrane potential between −100 and 0 mV. When tested against EC50 concentrations of glycine, PMBA was almost equally potent on α1 (IC50, 406±41 nM: Hill coefficient, 1.5±0.2) and α2 (IC50, 539±56 nM; Hill coefficient, 1.4±0.2) homomers. PMBA (1–10 μM) and strychnine (200 nM) reduced the potency of glycine and the amplitude of the maximal agonist response of α1 and α2 homomers. In 10 μM PMBA, two distinct classes of glycine response were observed on α2, only a single class of responses were observed on α1. There are similarities in PMBA and strychnine antagonism, although these compounds are structurally distinct. The possibility that PMBA interacts at two binding sites which differ in α1 and α2 subunits is discussed. PMBA may provide a lead structure for novel antagonists with which to investigate structural differences in glycine receptor at α1 and α2 subunits. British Journal of Pharmacology (1999) 126, 1230–1236; doi:10.1038/sj.bjp.0702402

We describe a new species from the genus Trevathana Anderson, 1992, collected from the Cocos/Keeling Islands in the Indian Ocean. Trevathana noae sp. nov. is similar to other species of Trevathana by its external shell and opercular valve morphology. It is distinct from congeners in that the tergum has a distinct spur which lacks an internal tooth in adult specimens. A key to the known species of Trevathana is given.

Alcohol dependence is a common, complex and debilitating disorder with genetic and environmental influences. Here we show that alcohol consumption increases following mutations to the γ-aminobutyric acidA receptor (GABAAR) β1 subunit gene (Gabrb1). Using N-ethyl-N-nitrosourea mutagenesis on an alcohol-averse background (F1 BALB/cAnN x C3H/HeH), we develop a mouse model exhibiting strong heritable preference for ethanol resulting from a dominant mutation (L285R) in Gabrb1. The mutation causes spontaneous GABA ion channel opening and increases GABA sensitivity of recombinant GABAARs, coupled to increased tonic currents in the nucleus accumbens, a region long-associated with alcohol reward. Mutant mice work harder to obtain ethanol, and are more sensitive to alcohol intoxication. Another spontaneous mutation (P228H) in Gabrb1 also causes high ethanol consumption accompanied by spontaneous GABA ion channel opening and increased accumbal tonic current. Our results provide a new and important link between GABAAR function and increased alcohol consumption that could underlie some forms of alcohol abuse.

The actions of picrotoxinin and four of its analogues were tested on a Drosophila melanogaster homo-oligomeric GABA (gamma-aminobutyric acid) receptor formed when RDL (resistance to dieldrin) subunits were expressed in Xenopus oocytes. In agreement with previously reported studies on native insect GABA receptors and native expressed vertebrate GABA receptors, acetylation of the bridgehead hydroxyl group (picrotoxinin acetate) greatly reduced the activity of the molecule, but surprisingly, substitution with flourine at the same position also reduced the activity. Conversion of the terminal isopropenyl group to an acetyl (alpha-picrotoxinone) or hydration of the double bond (picrotin) also reduced activity, in agreement with findings for native insect and mammalian receptors. The present results suggest that interactions of convulsants with homo-oligomeric and multimeric GABA receptors are qualitatively similar. Thus, the RDL homo-oligomer exhibits a pharmacological profile for picrotoxinin analogues resembling that of native GABA receptors.

High-affinity extrasynaptic GABAAreceptors are persistently activated by the low ambient GABA levels that are known to be present in extracellular space. The resulting tonic conductance generates a form of shunting inhibition that is capable of altering cellular and network behavior. It has been suggested that this tonic inhibition will be enhanced by neurosteroids, antiepileptics, and sedative/hypnotic drugs. However, we show that the ability of sedative/hypnotic drugs to enhance tonic inhibition in the mouse cerebellum will critically depend on ambient GABA levels. For example, we show that the intravenous anesthetic propofol enhances tonic inhibition only when ambient GABA levels are m. More surprisingly, the actions of the sleep-promoting drug 4,5,6,7-tetrahydroisothiazolo-[5,4-c]pyridin-3-ol (THIP) are attenuated at ambient GABA levels of just 20 nm. In contrast, our data suggest that neurosteroid enhancement of tonic inhibition will be greater at high ambient GABA concentrations. We present a model that takes into account realistic estimates of ambient GABA levels and predicted extrasynaptic GABAAreceptor numbers when considering the ability of sedative/hypnotic drugs to enhance tonic inhibition. These issues will be important when considering drug strategies designed to target extrasynaptic GABAAreceptors in the treatment of sleep disorders and other neurological conditions.

High-affinity extrasynaptic GABAAreceptors (GABAARs) are a prominent feature of cerebellar granule neurons and thalamic relay neurons. In both cell types, the presence of synaptic glomeruli would be expected to promote activation of these GABAARs, contributing to phasic spillover-mediated currents and tonic inhibition. However, the precise role of different receptor subtypes in these two phenomena is unclear. To address this question, we made recordings from neurons in acute brain slices from mice, and from tsA201 cells expressing recombinant GABAARs. We found that δ subunit-containing GABAARs of both cerebellar granule neurons and thalamic relay neurons of the lateral geniculate nucleus contributed to tonic conductance caused by ambient GABA but not to spillover-mediated currents. In the presence of a low “ambient” GABA concentration, recombinant “extrasynaptic” δ subunit-containing GABAARs exhibited profound desensitization, rendering them insensitive to brief synaptic- or spillover-like GABA transients. Together, our results demonstrate that phasic spillover and tonic inhibition reflect the activation of distinct receptor populations.

Modulation of GABAAreceptor function and inhibitory synaptic transmission by phosphorylation has profound consequences for the control of synaptic plasticity and network excitability. We have established that activating α-calcium/calmodulin-dependent protein kinase II (α-CaMK-II) in cerebellar granule neurons differentially affects populations of IPSCs that correspond to GABAAreceptors containing different subtypes of β subunit. By using transgenic mice, we ascertained that α-CaMK-II increased IPSC amplitude but not the decay time by acting via β2 subunit-containing GABAAreceptors. In contrast, IPSC populations whose decay times were increased by α-CaMK-II were most likely mediated by β3 subunit-containing receptors. Expressing α-CaMK-II with mutations that affected kinase function revealed that Ca2+and calmodulin binding is crucial for α-CaMK-II modulation of GABAAreceptors, whereas kinase autophosphorylation is not. These findings have significant consequences for understanding the role of synaptic GABAAreceptor heterogeneity within neurons and the precise regulation of inhibitory transmission by CaMK-II phosphorylation.

This study addresses whether the potentiation site for neurosteroids on GABA(A) receptors is conserved amongst different GABA(A) receptor isoforms. The neurosteroid potentiation site was previously identified in the alpha1beta2gamma2S receptor by mutation of Q241 to methionine or leucine, which reduced the potentiation of GABA currents by the naturally occurring neurosteroids, allopregnanolone or tetrahydrodeoxycorticosterone (THDOC). By using heterologous expression of GABA(A) receptors in HEK cells, in combination with whole-cell patch clamp recording methods, a relatively consistent potentiation by allopregnanolone of GABA-activated currents was evident for receptors composed of one alpha subunit isoform (alpha2-5) assembled with beta3 and gamma2S subunits. Using mutant alphabetagamma receptors, the neurosteroid potentiation was universally dependent on the conserved glutamine residue in M1 of the respective alpha subunit. Studying wild-type and mutant receptors composed of alpha4beta3delta subunits revealed that the delta subunit is unlikely to contribute to the neurosteroid potentiation binding site and probably affects the efficacy of potentiation. Thus, in keeping with the ability of neurosteroids to potentiate GABA currents via a broad variety of GABA(A) receptor isoforms in neurons, the potentiation site is structurally highly conserved on this important neurotransmitter receptor family.

The actions of penicillin-G (PCG) on wild-type and mutant Drosophila GABA receptor (RDL) subunits expressed in Xenopus oocytes were studied under two-electrode voltage-clamp. PCG was found to be a non-competitive antagonist of homomeric Drosophila RDL receptors with an IC(50) of 20.41 +/- 1.66 mM at EC(50) GABA. Substitution of a single amino acid (N318R) at the extracellular end of the channel lining region of the RDL subunit increased the potency of GABA approximately four fold, and increased the IC(50) of PCG to 5.09 +/- 0.38 mM. Although the antagonism by PCG on wild-type RDL receptors was independent of membrane potential, PCG action on the N318R mutant showed pronounced voltage-dependency, being much more effective at positive membrane potentials. Thus, in RDL homomers, the replacement of N318 by R318, a residue present at the equivalent position in vertebrate GABA(A) receptors, confers a vertebrate-like PCG pharmacology to the N318R mutant receptor. The A301S mutation that confers resistance to dieldrin did not significantly affect the antagonism by PCG.

Regulation of GABA(A) receptors by extracellular pH exhibits a dependence on the receptor subunit composition. To date, the molecular mechanism responsible for the modulation of GABA(A) receptors at alkaline pH has remained elusive. We report here that the GABA-activated current can be potentiated at pH 8.4 for both alphabeta and alphabeta gamma subunit-containing receptors, but only at GABA concentrations below the EC40. Site-specific mutagenesis revealed that a single lysine residue, K279 in the beta subunit TM2-TM3 linker, was critically important for alkaline pH to modulate the function of both alpha1beta2 and alpha1beta2 gamma2 receptors. The ability of low concentrations of GABA to reveal different pH titration profiles for GABA(A) receptors was also examined at acidic pH. At pH 6.4, GABA activation of alphabeta gamma receptors was enhanced at low GABA concentrations. This effect was ablated by the mutation H267A in the beta subunit. Decreasing the pH further to 5.4 inhibited GABA responses via alphabeta gamma receptors, whereas those responses recorded from alphabeta receptors were potentiated. Inserting homologous beta subunit residues into the gamma2 subunit to recreate, in alphabeta gamma receptors, the proton modulatory profile of alphabeta receptors, established that in the presence of beta2(H267), the mutation gamma2(T294K) was necessary to potentiate the GABA response at pH 5.4. This residue, T294, is homologous to K279 in the beta subunit and suggests that a lysine at this position is an important residue for mediating the allosteric effects of both acidic and alkaline pH changes, rather than forming a direct site for protonation within the GABA(A) receptor.

Importing functional GABAA receptors into synapses is fundamental for establishing and maintaining inhibitory transmission and for controlling neuronal excitability. By introducing a binding site for an irreversible inhibitor into the GABAA receptor alpha1 subunit channel lining region that can be accessed only when the receptor is activated, we have determined the dynamics of receptor mobility between synaptic and extrasynaptic locations in hippocampal pyramidal neurons. We demonstrate that the cell surface GABAA receptor population shows no fast recovery after irreversible inhibition. In contrast, after selective inhibition, the synaptic receptor population rapidly recovers by the import of new functional entities within minutes. The trafficking pathways that promote rapid importation of synaptic receptors do not involve insertion from intracellular pools, but reflect receptor diffusion within the plane of the membrane. This process offers the synapse a rapid mechanism to replenish functional GABAA receptors at inhibitory synapses and a means to control synaptic efficacy.

The role of Zn2+ in the CNS has remained enigmatic for several decades. This divalent cation is accumulated by specific neurons into synaptic vesicles and can be released by stimulation in a Ca2+-dependent manner. Using Zn2+ fluorophores, radiolabeled Zn2+, and selective chelators, the location of this ion and its release pattern have been established across the brain. Given the distribution and possible release under physiological conditions, Zn2+ has the potential to act as a modulator of both excitatory and inhibitory neurotransmission. Excitatory N-methyl-D-aspartate (NMDA) receptors are directly inhibited by Zn2+, whereas non-NMDA receptors appear relatively unaffected. In contrast, inhibitory transmission mediated via GABAAreceptors can be potentiated via a presynaptic mechanism, influencing transmitter release; however, although some tonic GABAergic inhibition may be suppressed by Zn2+, most synaptic GABA receptors are unlikely to be modulated directly by this cation. In the spinal cord, glycinergic transmission may also be affected by Zn2+ causing potentiation. Recently, the penetration of synaptically released Zn2+ into neurons suggests that this ion has the potential to act as a direct transmitter, by affecting postsynaptic signaling pathways. Taken overall, present studies are broadly supportive of a neuromodulatory role for Zn2+ at specific excitatory and inhibitory synapses.

GABA type A (GABA(A)) receptors are functionally regulated by external protons in a manner dependent on the receptor subunit composition. Although H(+) can regulate the open probability of single GABA ion channels, exactly what residues and receptor subunits are responsible for proton-induced modulation remain unknown. This study resolves this issue by using recombinant alpha1betai subunit GABA(A) receptors expressed in human embryonic kidney cells. The potentiating effect of low external pH on GABA responses exhibited p(Ka) in accord with the involvement of histidine and/or cysteine residues. The exposure of GABA(A) receptors to the histidine-modifying reagent DEPC ablated regulation by H(+), implicating the involvement of histidine residues rather than cysteines in proton regulation. Site-specific substitution of all conserved external histidines to alanine on the beta subunits revealed that H267 alone, in the TM2 domain, is important for H(+) regulation. These results are interpreted as a direct protonation of H267 on alpha1betai receptors rather than an involvement in signal transduction. The opposing functional effects induced by Zn(2+) and H(+) at this single histidine residue most likely reflect differences in charge delocalization on the imidazole rings in the mouth of the GABA(A) receptor ion channel. Additional substitutions of H267 in beta subunits with other residues possessing charged side chains (glutamate and lysine) reveal that this area of the ion channel can profoundly influence the functional properties of GABA(A) receptors.

The polycyclic dinitriles are a potent class of insecticides which are non-competitive GABA (gamma-aminobutyric acid) antagonists acting at the convulsant site. Comparison with other classes of GABA convulsant site ligands using molecular modelling has shown significant structural similarities. We have developed a pharmacophore model which unifies this class and some previous classes of GABA convulsants. Key pharmacophore elements are a polarizable functionality separated by a fixed distance from two H-bond accepting elements. This model is based on information from X-ray crystal structures and Sybyl using the Tripos force field. Using this pharmacophore model, numerous structural modifications were explored to enhance understanding of structure-activity relationships at the GABA receptor convulsant site of insects and mammals. A radiolabelled bicyclic dinitrile, [3H]BIDN [3H]3,3-bis-trifluoromethyl-bicyclo[2,2,1]heptane-2,2-dicarbonitrile+ ++), was prepared from this area of chemistry and was used as a probe for the interaction of polycyclic dinitriles at the target site.

Ionotropic gamma-aminobutyric acid (GABA) receptors are distributed throughout the nervous systems of many insect species. As with their vertebrate counterparts, GABAA receptors and GABAC receptors, the binding of GABA to ionotropic insect receptors elicits a rapid, transient opening of anion-selective ion channels which is generally inhibitory. Although insect and vertebrate GABA receptors share a number of structural and functional similarities, their pharmacology differs in several aspects. Recent studies of cloned Drosophila melanogaster GABA receptors have clarified the contribution of particular subunits to these differences. Insect ionotropic GABA receptors are also the target of numerous insecticides and an insecticide-resistant form of a Drosophila GABA-receptor subunit has enhanced our understanding of the structure-function relationship of one aspect of pharmacology common to both insect and vertebrate GABA receptors, namely antagonism by the plant-derived toxin picrotoxinin.

1. Blocking actions of the novel insecticide, fipronil, were examined on GABA responses recorded from Xenopus oocytes expressing either wild type (dieldrin-sensitive) or mutant (dieldrin-resistant) forms of the Drosophila melanogaster GABA-gated chloride channel homo-oligomer, RDL (the product of the resistance to dieldrin locus: Rdl). 2. In the case of the wild type receptor, fipronil blocked GABA-induced currents inducing both a shift to the right in the GABA dose-response curve and depressing the maximum amplitude of responses to GABA. The potency of fipronil was dependent on the GABA concentration but was unaffected by membrane potential. 3. Mutant RDL GABA-receptors, which have a naturally occurring amino acid substitution (A302-->S) in the putative ion-channel lining region, conferring resistance to dieldrin and picrotoxinin, were markedly less sensitive to fipronil than the wild-type receptors. 4. Fipronil antagonism is qualitatively similar to that produced by the structurally distinct compound, picrotoxinin. As the mutation A302-->S reduces the potency of both fipronil and picrotoxinin, homooligomeric RDL receptors should facilitate detailed studies of the molecular basis of convulsant/insecticide antagonist actions on GABA receptors.

Four new species of the families Hemioniscidae and Crinoniscidae are described from New Zealand waters: Crinoniscus politosummus sp. nov., C. cephalatus sp. nov., Scalpelloniscus vomicus sp. nov. and S. nieli sp. nov. Mature males and females are described for all species, and the epicaridium larva is described for S. vomicus sp. nov. Males of the two species of Scalpelloniscus can be distinguished from S. penicillatus and S. binoculis by coxal plate dentition, relative sizes of propodus and dactylus. Crinoniscus politosummus sp. nov. and C. cephalatus sp. nov. can be separated from the only other congener Crinoniscus equitans by the body shape of the mature females, and shape of antennule article 1 of the males. A single specimen tentatively identified as S. cf. penicillatus is also recorded from New Zealand waters. These species are the first records of cryptoniscoid isopods infecting thoracic barnacles from the South West Pacific.

The paper reviews very briefly the development of the Pescara free-piston engine and outlines the general constructional features of the GS 34 design of free-piston gas generator. The arrangements for the application of lubricants are described, illustrating the total loss and circulating oil systems. The requirements of lubricants are discussed together with some operating troubles associated with lubrication. The paper concludes with the contention that the individual efforts of the engine and petroleum industries will find their best return in co-operative action at all stages of further development.