J. R. de Weille

Isolation of a Tarantula Toxin Specific for a Class of Proton-gated Na+ Channels

Acid sensing is associated with nociception, taste transduction, and perception of extracellular pH fluctuations in the brain. Acid sensing is carried out by the simplest class of ligand-gated channels, the family of H+-gated Na+ channels. These channels have recently been cloned and belong to the acid-sensitive ion channel (ASIC) family. Toxins from animal venoms have been essential for studies of voltage-sensitive and ligand-gated ion channels. This paper describes a novel 40-amino acid toxin from tarantula venom, which potently blocks (IC50 = 0.9 nm) a particular subclass of ASIC channels that are highly expressed in both central nervous system neurons and sensory neurons from dorsal root ganglia. This channel type has properties identical to those described for the homomultimeric assembly of ASIC1a. Homomultimeric assemblies of other members of the ASIC family and heteromultimeric assemblies of ASIC1a with other ASIC subunits are insensitive to the toxin. The new toxin is the first high affinity and highly selective pharmacological agent for this novel class of ionic channels. It will be important for future studies of their physiological and physio-pathological roles.

The acid-sensitive ionic channel subunit ASIC and the mammalian degenerin MDEG form a heteromultimeric H+-gated Na+ channel with novel properties.

Proton-gated cation channels are acid sensors that are present in both sensory neurons and in neurons of the central nervous system. One of these acid-sensing ion channels (ASIC) has been recently cloned. This paper shows that ASIC and the mammalian degenerin MDEG, which are colocalized in the same brain regions, can directly associate with each other. Immunoprecipitation of MDEG causes coprecipitation of ASIC. Moreover, coexpression of ASIC and MDEG subunits in Xenopus oocytes generates an amiloride-sensitive H+-gated Na+ channel with novel properties (different kinetics, ionic selectivity, and pH sensitivity). In addition, coexpression of MDEG with mutants of the ASIC subunit can create constitutively active channels that become completely nonselective for Na+ versusK+ and H+-gated channels that have a drastically altered pH sensitivity compared with MDEG. These data clearly show that ASIC and MDEG can form heteromultimeric assemblies with novel properties. Heteromultimeric assembly is probably used for creating a diversity of H+-gated cation channels acting as neuronal acid sensors in different pH ranges.

The pre-transmembrane 1 domain of acid-sensing ion channels participates in the ion pore.

The acid-sensing ion channel (ASIC) subunits ASIC1, ASIC2, and ASIC3 are members of the amiloride-sensitive Na+ channel/degenerin family of ion channels. They form proton-gated channels that are expressed in the central nervous system and in sensory neurons, where they are thought to play an important role in pain accompanying tissue acidosis. A splice variant of ASIC2, ASIC2b, is not active on its own but modifies the properties of ASIC3. In particular, whereas most members of the amiloride-sensitive Na+ channel/degenerin family are highly selective for Na+ over K+, ASIC3/ASIC2b heteromultimers show a nonselective component. Chimeras of the two splice variants allowed identification of a 9-amino acid region preceding the first transmembrane (TM) domain (pre-TM1) of ASIC2 that is involved in ion permeation and is critical for Na+ selectivity. Three amino acids in this region (Ile-19, Phe-20, and Thr-25) appear to be particularly important, because channels mutated at these residues discriminate poorly between Na+ and K+. In addition, the pH dependences of the activity of the F20S and T25K mutants are changed as compared with that of wild-type ASIC2. A corresponding ASIC3 mutant (T26K) also has modified Na+selectivity. Our results suggest that the pre-TM1 region of ASICs participates in the ion pore.

Pharmacology and regulation of ATP-sensitive K+ channels

Introduction A new class of K + channels that link membrane potential to the bioenergetic situation of the cell has been recently discovered (Noma 1983). These K + channels (KATP) are normally closed at physiological intracellular ATP concentrations and open upon a diminution of [ATP]in. These channels have been shown to be present in pancreatic B-cells (Ashcroft et al. 1984; Cook and Hales 1984; Rorsman and Trube 1985a; Rorsman and Trube 1985b), cardiac ventricular cells (Noma 1983; Trube and Hescheler 1984) and skeletal muscle (Spruce et al. 1985; Spruce et al. 1986; Spruce et al. 1987). They might also be present in the central nervous system (Ashford et al. 1988; Bernardi et al. 1988) and in smooth muscle (Quast 1988; Quast and Cook 1988).

K+ Channels: Structure, Function, Regulation, Molecular Pharmacology and Role in Diseased States

This chapter will describe the molecular pharmacology and biochemistry of three types of K+ channels: the calcium-activated potassium channels. ATP-regulated potassium channels and voltage-sensitive potassium channels.

ATP-Sensitive K+ Channels : Molecular Pharmacology, Regulation and Role in Diseased States

ATP-dependent K+ (KATP) channels have now been identified in many tissues including β-cells, cardiac cells, skeletal muscle cells and neurons (reviewed in.l). They are the targets of 2 important classes of drugs, the antidiabetic sulfonylureas (2,3), which block the channel, and a series of compounds called K+ channel openers (24,25) and which include cromakalim, pinacidil, nicorandil, minoxidil sulfate, and RP 49356, which tend to maintain the channel in an open conformation. The activity of KATP channels is regulated by the ATP/ADP ratio. ATP or ADP alone inhibit the KATP channel. However, in the presence of ATP, ADP-Mg2+ activates the channel (see for example 6). The KATP channel is an excellent reporter of intracellular variations of the ATP/ADP ratio and has been used to demonstrate the presence of Cl− channels essential for oxydative phosphorylation in mitochondria (7)