Double-gating mechanism and diversity of an adenosine triphosphate (ATP)-sensitive K~ channel in neurons acutely dissociated from rat neocortex

Classically, ion channels are classified into 2 groups: chemical-sensitive (ligand-gated) and voltage-sensitive channels. Single ATP-sensitive K (K-ATP) channel currents were recorded in acutely dissociated rat neo-cortical neurons using patch clamp technique. A type of K-ATP channel has been found to be gated not only by intra-cellular ATP, but also by membrane potential ( Vm) , and proved to be a novel mechanism underlying the gating of ion channels, namely bi-gating mechanism. The results also show that the K-ATP channels possess heterogeneity and di-versity. These types of K-ATP channels have been identified in 40.12% of all patches, which are different in activa-tion-threshold and voltage-sensitivity. The present experiment studied the type-3 K-ATP channel with a unitary con-ductance of about 80 pS in detail ( n = 15). Taking account of all the available data, a variety of K-ATP channels are suggested to exist in body, and one type of them is bi-gated by both chemical substances and membrane poten     

Solution structure of the potassium channel inhibitor agitoxin 2: caliper for probing channel geometry.

The structure of the potassium channel blocker agitoxin 2 was solved by solution NMR methods. The structure consists of a triple-stranded antiparallel beta-sheet and a single helix covering one face of the beta-sheet. The cysteine side chains connecting the beta-sheet and the helix form the core of the molecule. One edge of the beta-sheet and the adjacent face of the helix form the interface with the Shaker K+ channel. The fold of agitoxin is homologous to the previously determined folds of scorpion venom toxins. However, agitoxin 2 differs significantly from the other channel blockers in the specificity of its interactions. This study was thus focused on a precise characterization of the surface residues at the face of the protein interacting with the Shaker K+ channel. The rigid toxin molecule can be used to estimate dimensions of the potassium channel. Surface-exposed residues, Arg24, Lys27, and Arg31 of the beta-sheet, have been identified from mutagenesis studies as functionally important for blocking the Shaker K+ channel. The sequential and spatial locations of Arg24 and Arg31 are not conserved among the homologous toxins. Knowledge on the details of the channel-binding sites of agitoxin 2 formed a basis for site-directed mutagenesis studies of the toxin and the K+ channel sequences. Observed interactions between mutated toxin and channel are being used to elucidate the channel structure and mechanisms of channel-toxin interactions.     

Evidence that the crystalline arrays in the outer membrane of Neurospora mitochondria are composed of the voltage-dependent channel protein

Antibodies were raised in rabbits against the outer membrane of Neurospora mitochondria. Antibodies were obtained that were specific for this membrane's major polypeptide ($\text{M, 31 000}$) and its slower-migrating derivatives on SDS-polyacrylamide gels. These antibodies inhibited the insertion into phospholipid bilayers of voltage-dependent ion channels from detergent extracts of the mitochondrial outer membranes. The same antibodies bound preferentially to membranes containing crystalline surface arrays in outer mitochondrial membrane fractions. These results indicate that the 31 kDa polypeptide is a component both of the ion channels and of the membrane arrays, suggesting identity between the functional and structural entities.     

Potassium-induced insulin release and voltage noise measurements in single mouse islets of Langerhans

Summary Insulin release and membrane potential fluctuations in response to increased extracellular potassium [K⁺] _o have been measured in single perifused islets of Langerhans from normal mice. An increase in [K⁺] _o from 5mm to 50mm induced a transient insulin release with a peak at about 1 min. The peak value was [K⁺] _o -dependent but the half-timet _1/2 for the decline was constant at nearly 1 min. 2.5mm cobalt completely inhibited the potassium-induced stimulation of insulin release. The insulin release elicited by 28 and 50mm [K⁺] _o was similar in terms of peak, total release and half-time from maximum release. Stepwise increase in [K⁺] _o from 10 to 28 to 50mm resulted in a normal response to 28mm but no peak of release after the 28 to 50mm increase. The results indicate good correlation between excess voltage noise, thought to reflect calcium channel activity, and insulin release evoked by changing extracellular potassium.     

Membrane damage by a toxin from the sea anemone Stoichactis helianthus. I. Formation of transmembrane channels in lipid bilayers

The addition of nanomolar amounts of a toxin preparation derived from the sea anemone Stoichactis helianthus to black lipid membranes increases their electrical conductance by one million-fold. In addition, the membranes become permeable predominantly to monovalent cations. The elevated bilayer conductance is voltage-dependent, and the current-voltage curves of these bilayers display rectification as well as a region of negative resistance. The membrane activity of the toxin is proportional to the third power of its concentration, and at very low concentrations the membrane conductance increases in discrete uniform steps. These observations indicate that the mechanism of toxin action involves the formation of transmembrane channels constructed by the aggregation of protein molecules which are inserted in the bilayer. The voltage-dependent membrane conductance arises from two distinct channel characteristics: (1) the unit conductance of individual channels is dependent on the polarity of applied voltage; (2) the number of ion-conducting channels is influenced by the polarity as well as the magnitude of applied potential. It is believed that these effects are due to the influence of an electric field on the insertion of toxin molecules into the bilayer or on their subsequent association with each other to produce channels. Partial chemical characterization of the toxin material has shown that the membrane active factor is a basic protein with a molecular weight of 17 500.     

Five New Species of Henneguya (Protozoa: Myxosporida) from Ictalurid Fishes*

SYNOPSIS. Four new species of Henneguya (myxosporidan) are described from Ictalurus punctatus Rafinesque (channel catfish). They are as follows: Henneguya postexilis sp. n. and Henneguya longicauda sp. n. from the gills; Henneguya adiposa sp. n. from the adipose fin; and Henneguya diversis sp. n. from the liver, kidney, connective tissue of muscles and fins, and tumor-like external growths. Henneguya pellis sp. n. is described from the dermis of Ictalurus furcatus (LeSueur) (blue catfish). The development stages of all described species are discussed. Henneguya exilis Kudo was found on the gills of one I. punctatus; notes on its spore characteristics are presented.     

Whole-cell K+ current activation in response to voltages and carbachol in gastric parietal cells isolated from guinea pig

Summary Patch-clamp studies of whole-cell ionic currents were carried out in parietal cells obtained by collagenase digestion of the gastric fundus of the guinea pig stomach. Applications of positive command pulses induced outward currents. The conductance became progressively augmented with increasing command voltages, exhibiting an outwardly rectifying current-voltage relation. The current displayed a slow time course for activation. In contrast, inward currents were activated upon hyperpolarizing voltage applications at more negative potentials than the equilibrium potential to K⁺ (E _K). The inward currents showed time-dependent inactivation and an inwardly rectifying current-voltage relation. Tail currents elicited by voltage steps which had activated either outward or inward currents reversed at nearE _K, indicating that both time-dependent and voltagegated currents were due to K⁺ conductances. Both outward and inward K⁺ currents were suppressed by extracellular application of Ba²⁺, but little affected by quinine. Tetraethylammonium inhibited the outward current without impairing the inward current, whereas Cs⁺ blocked the inward current but not the outward current. The conductance of inward K⁺ currents, but not outward K⁺ currents, became larger with increasing extracellular K⁺ concentration. A Ca²⁺-mobilizing acid secretagogue, carbachol, and a Ca²⁺ ionophore, ionomycin, brought about activation of another type of outward K⁺ currents and voltage-independent cation currents. Both currents were abolished by cytosolic Ca²⁺ chelation. Quinine preferentially inhibited this K⁺ current. It is concluded that resting parietal cells of the guinea pig have two distinct types of voltage-dependent K⁺ channels, inward rectifier and outward rectifier, and that the cells have Ca²⁺-activated K⁺ channels which might be involved in acid secretion under stimulation by Ca²⁺-mobilizing secretagogues.     

Comparisons of benthic invertebrates between riffles and pools

Benthic species assemblages in upstream and downstream ends of riffles and in pools were investigated seasonally in the first five orders of an alluvial gravel stream with distinct pool and riffle channel form. Riffles comprised < 10% of stream area and were separated by pools with extensive bedrock substrate (ca. 15–85% of total surface area) which was scoured during periodic high flow. Virtually all taxa were more abundant in riffles than in pools, except chironomids which were more equally distributed. Inconsistent results were obtained for upstream-downstream comparisons within riffles. Intermittent headwater reaches (orders 1 & 2) which supported half as many taxa retained this pattern during periods of flow, although riffles at these sites were dry from mid-June to mid-November. Pools which contained more gravel, indicating less disturbance during high flow, had a richer assemblage of benthic species than other pools. Many invertebrates in pools may have been there as a result of drift from their preferred riffle habitats, but the presence of gravel in the pools indicates less intense flow disturbance during floods, provides protection from the mild scouring that does occur during floods, and provides refugia from predators.     

Incorporation of the gene for a cell-cell channel protein into transformed cells leads to normalization of growth

Summary Incorporation of the gene for connexin 43, a cell-cell channel protein of gap junction, into the genome of communication-deficient transformed mouse 10T1/2 cells restored junctional communication and inhibited growth. Growth was slowed, saturation density reduced and focus formation suppressed, and these effects were contingent on overexpression of the exogenous gene and the consequent enhancement of communication. In coculture with normal cells the growth of the connexin overexpressors was completely arrested, as these cells established strong communication with the normal ones. Thus, in culture by themselves or in coculture, the connexin overexpressor cells grew like normal cells. These results demonstrate that the cell-cell channel is instrumental in growth control; they are the expected behavior if the channel transmits cytoplasmic growth-regulatory signals.