Robert Harvey

There are concerns regarding the potential development of drug resistance in scabies. Recently, there has been a substantial increase in the global incidence of scabies, together with reports of treatment failure in parts of Europe. While it can be difficult to demonstrate cases of genuine resistance, there is now clinical evidence that permethrin resistance may indeed explain a proportion of these treatment failures. Permethrin resistance has been confirmed in a laboratory model of Sarcoptes scabiei var. canis and is associated with sodium channel mutations and increased detoxification activity. Ivermectin resistance has also been noted in cases of crusted scabies, with several other cases of ivermectin treatment failure likely linked to resistance. Resistance to ivermectin and other macrocyclic lactones is likely a polyfactorial trait. Increased transcription of a P-glycoprotein efflux pump and detoxifying Glutathione S- transferases was found in mites collected from patients after ivermectin treatment. Mutations in ligand-gated chloride channels have been associated with ivermectin resistance in other mites and lice and would be worthy of investigation in S. scabiei. Research efforts should focus on the collection of mites in cases of suspected resistance for molecular analysis. Further characterisation of the scabies mite ion channels would better develop understanding of the physiological targets of acaricides and help guide treatment strategies to avoid further development of resistance. This may include using synergised pyrethroids and/or combination therapies with different target effects in S. scabiei.

This book reviews the recent advances in our understanding of the molecular and cellular mechanisms that control the assembly, transport, targeting and anchoring of the protein complexes making up the most important ion channels and receptor families, fundamental to synaptic function. Improved understanding of these processes is expected to reveal novel therapeutic targets relevant to a range of disease states. Receptor and Ion-Channel Trafficking provides an integrated overview of the significant recent advances in the field of molecular neurobiology and will be essential reading for all researchers in neuroscience, neurobiology, biochemistry, cellular and molecular biology. [Book Synopsis]

The amino acid glycine is highly concentrated in the ventral and dorsal horns of the spinal cord, in many brain stem nuclei, and in sensory relay stations such as the cochlear nucleus and the retina. Traditional physiological studies have shown that glycine is a major inhibitory neurotransmitter that performs a vital role in the control of both motor and sensory pathways (Aprison 1990). In presynaptic nerve terminals of glycinergic interneurons in spinal cord and brain stem, cytosolic glycine is concentrated in small clear synaptic vesicles by an H+-dependent vesicular glycine transporter. Excitation of these interneurons leads to the calcium-triggered fusion of these synaptic vesicles with the presynaptic plasma membrane, thus liberating glycine into the synaptic cleft. Glycine then binds to postsynaptic glycine receptors (GlyRs), causing gating of an integral anion channel that increases the chloride ion conductance of the plasma membrane. This postsynaptic action of glycine is selectively antagonized by the plant alkaloid strychnine. In mature neurons, where the chloride equilibrium potential approximates the resting potential, GlyR activation results in chloride ion influx. This neutralizes depolarization by sodium ion influx, thereby inhibiting the propagation of action potentials. However, a different response is found in the developing nervous system, where immature neurons contain very high intracellular chloride concentrations (Wang et al. 1994). Here, glycine-induced increases in chloride conductance cause CI-efflux, resulting in depolarization of the postsynaptic cell (see Reichling et al. 1994; Boehm et al. 1997).

Glycine serves as a major inhibitory neurotransmitter in the adult vertebrate central nervous system and is implicated in the control of many motor and sensory pathways. In addition, this amino acid functions as an excitatory transmitter during embryonic development and is an essential coagonist at glutamatergic synapses containing the N-methyl-d-aspartate (NMDA) subtype of glutamate receptors

Glycine, the simplest of all amino acids, is highly enriched in spinal cord and brain stem compared with other regions of the central nervous system. Classical physiological analysis has revealed that glycine serves as a major inhibitory neurotransmitter in the control of motor and sensory pathways (APRISON 1990). In the nerve terminals of glycinergic interneurons in spinal cord and brain stem, cytosolic glycine is concentrated in small clear synaptic vesicles by an H+-dependent vesicular transporter. Excitation of the interneurons causes Ca2+-triggered fusion of these synaptic vesicles with the presynaptic plasma membrane, thus initiating glycine release into the synaptic cleft. This results in the activation of postsynaptic glycine receptors (GlyRs) which mediate an increase in chloride conductance by opening an integral anion channel in response to agonist binding. As the chloride equilibrium of mature neurons is close to their resting potential, glycine-mediated Cl-influx normally antagonizes depolarization by Na+ influx and thus inhibits the propagation of action potentials. However, glycine can also serve as an excitatory neurotransmitter. Immature neurons in the developing central nervous system often contain very high intracellular chloride concentrations (WANG et al. 1994). In these neurons, glycine-induced increases in chloride conductance cause Cl-efflux, resulting in membrane depolarization and neurotransmitter release (REICHLING et al.1994; BOEHM et al.1997). Excitatory GlyRs may be especially relevant to synaptogenesis, since glycine-triggered rises in intracellular Ca2+ have recently been shown to be crucial for the correct formation of postsynaptic glycinergic membrane specializations (KIRSCH and BETZ 1998). Thus, the developmental regulation of intracellular Cl-oncentration critically controls the nature of the postsynaptic response to glycine.

Rapid chemical communication between cells in the vertebrate central nervous system is mediated by ligand-gated ion-channel receptors (also called ionotropic receptors), which are multisubunit complexes, that each contain an ion-selective channel. In response to an appropriate signal, neurotransmitter is released from the storage vesicles in the presynaptic terminal of a neuron into the synaptic cleft; the signaling molecules then diffuse across this intercellular compartment and bind to receptors located in the membrane of the postsynaptic cell. The binding of neurotransmitter to an ionotropic receptor results, via an unknown mechanism that is assumed to involve a conformational change in the protein, in the opening of the ion channel and the flux of either cations or anions. This rapid neurotransmission, which occurs on the millisecond time scale, is terminated by the closure of the channel as a result of either agonist dissociation, or receptor desensitization, and either transmitter reuptake or hydrolysis.

In this chapter we introduce the reader to the structures of the different types of ligand-gated ion-channel receptor, and the numerous receptor subtypes that have recently been revealed to exist, in both invertebrate and vertebrate species, by the application of molecular biological methods. We then review some of the data in support of the existence, in molluscs, of such receptor/channel complexes for γ-aminobutyric acid, glutamate and acetylcholine. Finally, recent results from our laboratory on the cloning and expression of complementary DNAs, from the pond-snail Lymnaea stagnalis, that encode GABAA and glutamate receptor subunits will be described.

A cDNA encoding part of a polypeptide (Loc1) that exhibits similarity to the corresponding portion of the rat GluRl subunit has been identified by screening an amplified locust cDNA library. This polypeptide is deduced to be missing about 200 amino acids of the amino-terminus and about 100 amino acids of the carboxy-terminus. cDNAs encoding two other glutamate receptor-like polypeptides (Loc2 and Loc3), which both exhibit good sequence homology with Loci, have also been identified. So far, there is no evidence for 'flip' and 'flop' variants of Loci, 2 and 3. A 27-mer peptide including the Ml sequence of Loci and a 25-mer peptide including the M2 sequence of this putative glutamate receptor subunit have been synthesised and incorporated into artificial bilayers. Channel openings, of minimum conductance 20 pS, were seen more frequently with the M2 peptide. These studies are designed to lead to the isolation of full-length cDNAs for Loci, 2 and 3 and to the electrophysiological characterisation of their ion transport properties.