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Biomedical subjects

W F Gilly

Publications and source records attributed to W F Gilly.

At least 37 records · Page 2Linked to original sources
PubMed1995

Tissue distribution and subcellular localization of Na+ channel mRNA in the nervous system of the squid, Loligo opalescens.

Recent cloning of a putative Na+ channel alpha subunit cDNA, GFLN1, from the squid stellate ganglion has allowed us to study the expression of this ion channel at a cellular level. In situ hybridizations with a probe derived from and specific to 3' untranslated and coding sequence of GFLN1 were used to determine its tissue distribution as well as its subcellular localization. In sections of the stellate ganglion, the probe labeled all of the cells in the giant fiber lobe (GFL) and most cells in the cellular layer of the main ganglion. In these non-GFL portions of the stellate ganglion, labeling was particularly intense in the ventral large cells and weak or absent in the dorsal small cells. In the optic lobe, only a select group of cells, the second-order visual giant neurons, were intensely labeled. These results are consistent with electrophysiological data that show GFL-like Na+ currents in rare large cells dissociated from the optic lobe and in most but not all cells from the non-GFL part of the stellate ganglion. In sections of the subesophageal mass of the central nervous system, strong labeling for GFLN1 mRNA occurred in the fin lobe, posterior chromatophore lobe, central and latero-ventral palliovisceral lobes, and posterior pedal lobe. In all cases, labeling was detected only in the cellular layer of these tissues and never in nerves or neuropil. In situ hybridization with dissociated GFL neurons maintained in primary culture verified that Na+ channel mRNA is confined to the cell body. These results indicate that GFLN1 is expressed predominately in large cells with large or long axons, and that this mRNA is restricted to the cell bodies of these neurons.

Animals↗
PubMed1993

Amino acid sequence of a putative sodium channel expressed in the giant axon of the squid Loligo opalescens.

A full-length cDNA encoding a putative Na+ channel (GFLN1) has been cloned from a library prepared from the stellate ganglion of Loligo opalescens. The cDNA encodes a predicted protein of 1784 amino acids. Regions of the GFLN1 protein with defined functional importance (membrane span S4, the SS1 and SS2 segments, and interdomain III-IV) are highly conserved among all vertebrate Na+ channel alpha-subunit structures. Northern blot hybridization and RNase protection assays verify that mRNA corresponding to GFLN1 is expressed in neurons of the giant fiber lobe that form the giant axon. We propose that GFLN1 encodes the Na+ channel that has been extensively studied in the squid axon.

Amino Acid Sequence↗
PubMed1993

Voltage-dependent calcium and potassium conductances in striated muscle fibers from the scorpion, Centruroides sculpturatus.

Ionic currents responsible for the action potential in scorpion muscle fibers were characterized using a three-intracellular microelectrode voltage clamp applied at the fiber ends (8-12 degrees C). Large calcium currents (ICa) trigger contractile activation in physiological saline (5 mM Ca) but can be studied in the absence of contractile activation in a low Ca saline (< or = 2.5 mM). Barium (Ba) ions (1.5-3 mM) support inward current but not contractile activation. Ca conductance kinetics are fast (time constant of 3 msec at 0 mV) and very voltage dependent, with steady-state conductance increasing e-fold in approximately 4 mV. Half-activation occurs at -25 mV. Neither ICa nor IBa show rapid inactivation, but a slow, voltage-dependent inactivation eliminates ICa at voltages positive to -40 mV. Kinetically, scorpion channels are more similar to L-type Ca channels in vertebrate cardiac muscle than to those in skeletal muscle. Outward K currents turn on more slowly and with a longer delay than do Ca currents, and K conductance rises less steeply with voltage (e-fold change in 10 mV; half-maximal level at 0 mV). K channels are blocked by externally applied tetraethylammonium and 3,4 diaminopyridine.

4-Aminopyridine↗
PubMed1990

Control of the spatial distribution of sodium channels in giant fiber lobe neurons of the squid.

Na+ channels are present at high density in squid giant axon but are absent from its somata in the giant fiber lobe (GFL) of the stellate ganglion. GFL cells dispersed in vitro maintain growing axons and develop a Na+ channel distribution similar to that in vivo. Tunicamycin, a glycosylation inhibitor, selectively disrupts the spatially appropriate, high level expression of Na+ channels in axonal membrane but has no effect on expression in cell bodies, which show low level, inappropriate expression in vitro. This effect does not appear to involve alteration in Na+ channel turnover or axon viability. K+ channel distribution is unaffected. Thus, glycosylation appears to be involved in controlling Na+ channel localization in squid neurons.

Animals↗
PubMed1990

Jet-propelled escape in the squid Loligo opalescens: concerted control by giant and non-giant motor axon pathways.

Recordings of stellar nerve activity were made during escape responses in living squid. Short-latency activation of the giant axons is triggered by light-flash stimulation that elicits a stereotyped startle-escape response and powerful jet. Many other types of stimuli produce a highly variable, delayed-escape response with strong jetting primarily controlled by a small axon motor pathway. In such cases, activation of the giant axons is not necessary for a vigorous escape jet. When they are utilized, the giant axons are not activated until well after the non-giant system initiates the escape response, and excitation is critically timed to boost the rise in intramantle pressure. Squid thus show at least two escape modes in which the giant axons can contribute in different ways to the control of a highly flexible behavior.

Animals↗
PubMed1989

Properties of appropriately and inappropriately expressed sodium channels in squid giant axon and its somata.

Neurons that form the giant axons in squid by axonal fusion in the stellate ganglion are inexcitable and do not express functional voltage-controlled sodium (Na) channels in their somata in vivo. These cells do express Na channels in the soma membrane in vitro, however, provided they have been axotomized. We describe here voltage-clamp experiments on the isolated cell bodies maintained in primary culture and on acutely isolated giant axons designed to compare the functional properties of the Na channels expressed inappropriately in the soma with those of channels expressed normally in the axon. Approximately 85% of Na channels in the soma are essentially indistinguishable from those in the giant axon with regard to gating properties and sensitivity to tetrodotoxin or saxitoxin. Thus, the isolated soma is capable of processing Na channels to a state of apparent functional perfection. In addition to these normal Na channels, another type is regularly expressed in the cultured somata. This second type lacks inactivation and is preferentially sensitive to block by cadmium ions, but is otherwise indistinguishable from the more prevalent normal type of channels.

Animals↗
PubMed1987

Physiological properties of three muscle fibre types controlling dorsal fin movements in a flatfish, Citharichthys sordidus.

Pacific sand dabs utilize their dorsal and anal fins in different behaviours which are characterized by extremely rapid fin movements, on one hand, and essentially isometric force generation on the other. Muscle fibres controlling fin movements were examined physiologically. Direct electrical shocks to localized regions of fin muscles reveal three fibre types. The longest fibres in the muscle are very fast and functionally analogous to frog twitch fibres. The shortest fibres are extremely, slow and show properties much like frog tonic fibres. The mid-length fibres produce contractile responses which are intermediate in time course. Even the fastest muscle fibres do not generate action potentials, but instead rely on summating junction potentials to drive membrane voltage to a stable level just beyond contraction threshold (-35 to -40 mV). Twitch amplitude can be finely graded by the time that membrane depolarization exceeds this threshold level.

Action Potentials↗
PubMed1987

Golgi stain identifies three types of fibres in fish muscle.

Using Golgi infiltration we have studied the structure and disposition of transverse tubules in muscle fibres from the sand dab fin musculature. Three types of fibres differ significantly from each other in the extent and disposition of junctions between transverse tubules and the sarcoplasmic reticulum. These correlate with the three groups of fibres having different relaxation times shown in the accompanying paper (Gilly & Aladjem, 1987). Fibres with very slow relaxation (tonic fibres) correspond to those which have an unusual disposition of T tubules and very rare T-SR junctions. In the fast twitch fibres the peripheral T tubules segments converge into tangentially arranged tubules before joining the plasmalemma.

Animals↗
PubMed1987

Synthesis of sodium channels in the cell bodies of squid giant axons.

Giant axons in squid are formed by fusion of axons from many small cell bodies in the giant fiber lobe (GFL) of the stellate ganglion. Somata of GFL cells in vivo are inexcitable and do not have measurable sodium current (INa) when studied with microelectrode or patch-electrode voltage-clamp techniques. If GFL cells are separated from the giant axons and maintained in primary culture, axon-like INa can be recorded from the somata after several days. Incorporation of Na channels into GFL cell bodies requires protein synthesis, intracellular microtubule-based transport, and the lack of a morphologically defined axon to serve as a sink for channels synthesized in culture.

Animals↗
PubMed1986

Morphological and physiological properties of non-striated muscle from the tunicate, Ciona intestinalis: parallels with vertebrate skeletal muscle.

Non-striated muscle from the longitudinal body wall muscle in an adult ascidian (tunicate) is characterized morphologically and physiologically. These muscles are unique among chordate non-striated (i.e. 'smooth') muscles in that they are composed of discrete bundles of several small diameter (4-10 microns) muscle cells (fibers) arranged in parallel functional units. Each bundle is wrapped by a basal lamina, and at least some appear to be directly innervated at a neuromuscular junction similar to an end plate. In these regards, a bundle of Ciona smooth muscle cells is analogous to a skeletal muscle fiber of a vertebrate. This analogy also extends to physiological properties. Ciona muscle generates a rapid all-or-none Ca action potential which gives rise to a brisk twitch with brief latency. These anatomical and physiological adaptations are discussed in terms of the evolution of vertebrate skeletal muscle.

Action Potentials↗
PubMed1986

Charge movement and depolarization-contraction coupling in arthropod vs. vertebrate skeletal muscle.

Voltage-dependent charge movement has been characterized in arthropod skeletal muscle. Charge movement in scorpion (Centuroides sculpturatus) muscle is distinguishable from that in vertebrate skeletal muscle by criteria of kinetics, voltage dependence, and pharmacology. The function of scorpion charge movement is gating of calcium channels in the sarcolemma, and depolarization-contraction coupling relies on calcium influx through these channels.

Animals↗
PubMed1984

Contractile activation in scorpion striated muscle fibers. Dependence on voltage and external calcium.

Excitation-contraction coupling was characterized in scorpion striated muscle fibers using standard microelectrode techniques as employed in studies on vertebrate skeletal muscle. The action potential of scorpion muscle consists of two phases of regenerative activity. A relatively fast, overshooting initial spike is followed by a prolonged after-discharge of smaller, repetitive spikes. This after-discharge is accompanied by a twitch that relaxes promptly upon repolarization. Twitches fail in Na-free, tetrodotoxin (TTX)-containing, or Ca-free media. However, caffeine causes contractures in muscles paralyzed by Na- and Ca-free solutions. Experiments on muscle fibers voltage-clamped at a point with two microelectrodes in Na-free or TTX-containing media indicate that: (a) the strength-duration relation for threshold contractions has a shape similar to that in frog muscle, but mean values are displaced approximately 20 mV in the positive direction; (b) tetracaine exerts a parallel effect on strength-duration curves from scorpion and frog; (c) contractile activation in scorpion is abolished in Ca-free media; and (d) the contractile threshold is highly correlated with the occurrence of inward Ca current for pulses of all durations. Thus, the voltage dependence of contractile activation in scorpion and frog muscle is similar. However, the preparations differ in their dependence on extracellular Ca for contraction. These results are discussed in relation to possible mechanisms coupling tubular depolarization to Ca release from the sarcoplasmic reticulum in vertebrate and invertebrate skeletal muscle.

Animals↗
PubMed1982

Slowing of sodium channel opening kinetics in squid axon by extracellular zinc.

The interaction of Zn ion on Na channels was studied in squid giant axons. At a concentration of 30 mM Zn2+ slows opening kinetics of Na channels with almost no alteration of closing kinetics. The effects of Zn2+ can be expressed as a "shift" of the gating parameters along the voltage axis, i.e., the amount of additional depolarization required to overcome the Zn2+ effect. In these terms the mean shifts caused by 30 mM Zn2+ were +29.5 mV for Na channel opening (on) kinetics (t1/2 on), +2 mV for closing (off) kinetics (tau off), and +8.4 mV for the gNa-V curve. Zn2+ does not change the shape of the instantaneous I-V curve for inward current, but reduces it in amplitude by a factor of or approximately 0.67. Outward current is unaffected. Effects of Zn2+ on gating current (measured in the absence of TTX) closely parallel its actions on gNa. On gating current kinetics are shifted by +27.5 mV, off kinetics by +6 mV, and the Q-V distribution by +6.5 mV. Kinetic modeling shows that Zn2+ slows the forward rate constants in activation without affecting backward rate constants. More than one of the several steps in activation must be affected. The results are not compatible with the usual simple theory of uniform fixed surface charge. They suggest instead that Zn2+ is attracted by a negatively charged element of the gating apparatus that is present at the outer membrane surface at rest, and migrates inward on activation.

Animals↗
PubMed1982

Divalent cations and the activation kinetics of potassium channels in squid giant axons.

The effects of external Zn+2 and other divalent cations on K channels in squid giant axons were studied. At low concentration (2 mM) Zn+2 slows opening kinetics without affecting closing kinetics. Higher concentrations (5-40 mM) progressively slow opening and speed channel closing to a lesser degree. In terms of "shifts," opening kinetics are strongly shifted to the right on the voltage axis, and off kinetics much less so. The shift of the conductance-voltage relation along the axis is intermediate. Zinc's kinetic effects show little sign of saturation at the highest concentration attainable. Zn does not alter the shape of the instantaneous current-voltage relation of open channels. Some other divalent cations have effects similar to Zn+2, Hg2+ being the most potent and Ca+2 the least. After treatment with Hg+2, which is irreversible, Zn+2 still slows opening kinetics, which suggests that each channel has at least two sites for divalent cation action. The results are not compatible with a simple theory of fixed, uniform surface charges. They suggest that external cations interact directly with a negatively charged element of the gating apparatus that moves inward from the membrane's outer surface during activation. Examination of normal kinetics shows that there is a slow step somewhere in the chain leading to channel opening. But the slowest step must not be the last one.

Animals↗
PubMed1980

Gating current and potassium channels in the giant axon of the squid.

Gating current (Ig) underlying Na-channel activation is large enough to enable resolution of components both preceding and paralleling Na conductance (gNa) turn-on. For large depolarizations (beyond +20 mV), an additional "slow phase" of Ig is observed during a time when Na activation is already complete, but when K-channel opening is just becoming detectable. If Na- and K-channel gating are similar, the slow kinetics and long delay for K activation predict that K channel Ig must be relatively small and slow. Externally applied dibucaine almost totally blocks gNa and greatly reduces the fast (Na channel) Ig without altering gK or the Ig slow phase. The slow phase of Ig depends in part of the presence of functional K channels. Selective diminution in amplitude of the slow phase is consistently observed after a 30-min perfusion with both external and internal K-free media, a procedure which destroys nearly all K channels. This decrease of Ig amounts to approximately 10% of the total charge movements at +40 to +80 mV, with gating charge and K channels disappearing in a ratio of less than 1 e- per picosiemens of gK. These findings are consistent with the idea that part of the Ig slow phase represents gating current generated by the early steps in K-channel activation.

Animals↗
PubMed1980

Mechanical activation in slow and twitch skeletal muscle fibres of the frog.

1. Slow and twitch muscle fibres of the frog were studied with a two-micro-electrode point voltage-clamp method. Slow fibres were identified in pyriformis and cruralis muscles by their appearance in the light microscope, electrical characteristics, and rate of sarcomere shortening or of tension development. 2. The relation between the amplitude and duration of threshold depolarizing pulses was determined in sartorius twitch and pyriformis slow fibres. Strength-duration relations for contractile activation are very similar in the two fibre types. 3. The effect of a brief subthreshold pulse on the threshold voltage level decays with a half-time of 1-2 msec at 9 degrees C in both slow and twitch fibres. This fast decay, thought to reflect voltage-dependent deactivation of Ca2+ release following repolarization, is followed by a slower decay of greatly different rates in the two fibre types. the slower components of decay might reflect the rate of background Ca2+ removal by the sarcoplasmic reticulum. 4. Reducing external Ca2+ levels to about about 0.1 microM with 2.5 mM-EGTA has no effect on the shapes of strength-duration curves for both slow and twitch fibres, suggesting that activator Ca2+ in both fibre types originates entirely from intracellular stores. 5. "Tonic' contractions were studied using voltage-clamped short cruralis slow fibres at 20 degrees C. Reducing external Ca2+ to about 0.1 microM had no effect on the steepness of the steady-state tension-voltage relation or on the ability of slow fibres to maintain maximal tension during long (200 sec) depolarizations to membrane potentials of up to +50 mV. 6. Functional similarities in activation kinetics of slow and twitch fibres are discussed in relation to the sensing of tubular membrane potential by the sarcoplasmic reticulum, to Ca2+ release from it, and to possible mechanisms involved in these processes. Processes leading to the rapid turning on and off of Ca2+ release in response to changes in tubular membrane potential are probably similar in slow and twitch fibres. However, the apparent lack of voltage-and time-dependent inactivation of Ca2+ release in slow fibre points to a major difference in the two types of muscle.

Animals↗
PubMed1980

Membrane electrical properties of frog slow muscle fibres.

1. Pyriformis slow (and sartorius twitch) fibres from Rana temporaria were studied with a three-micro-electrode voltage-clamp technique to obtain an approximate measurement of membrane current density at a fibre end. In most experiments, a modified Ringer solution containing 2H20 and 230 mM-sucrose was used to reduce movement. 2. Linear membrane properties of slow fibres obtained with this method are consistent with results from previous studies. Measured Cm (microF/cm2) increases with fibre diameter in a manner consistent with a tubular location of part of the fibre capacitance. 3. Voltage steps to -50mV and more positive potentials result in outward membrane currents in both slow and twitch fibres. These currents develop along similar sigmoid time courses and are blocked by tetraethylammonium (TEA+) ions. The reversal potential for delayed current channels in slow fibres varies with external K+ concentration, suggesting that the delayed current in slow fibres, as in twitch, is carried by K+ ions. 4. Maximum GK,GK, in slow fibres is an order of magnitude smaller than twitch fibres. The steady-state GK-V curve of slow fibres is very broad (e-fold for approximately 15 mV), saturating at very positive voltages, whereas the GK of twitch fibres varies more steeply with voltage. 5. No evidence of inward currents was seen in slow fibres during pulses of duration up to 96 msec. 6. Slow outward currents, which do not inactivate appreciably, are seen in slow fibres during long (10 sec) pulses. Tail currents following such long pulses are very slow. The reversal potential shifts to more positive values with increasing pulse duration.

Animals↗