Search PubMed⌕ Search

Biomedical subjects

E Stefani

Publications and source records attributed to E Stefani.

At least 127 records · Page 7Linked to original sources
PubMed1990

Increased MEPP frequency as an early sign of experimental immune-mediated motoneuron disease.

Intracellular recordings of miniature end-plate potentials were performed in extensor digitorum longus muscles from guinea pigs with experimental immune-mediated motoneuron destruction. In the early stages of the disease, the miniature end-plate potential frequency was elevated compared to that in control and normal animals. The amplitude and time course of the miniature end-plate potentials as well as the resting potential of the muscle fibers were not altered, which implies integrity of the postjunctional membrane. The increase in frequency of miniature end-plate potential reflects an increase of basal acetylcholine release and documents dysfunction of the presynaptic terminal of the neuromuscular junction. The increased frequency was associated with high levels of antimotoneuronal IgG in the blood and the presence of IgG at motor end-plates. These data suggest that the presynaptic terminal of the neuromuscular junction may be involved in the immune attack in animal models of motoneuron degeneration.

Animals↗
PubMed1990

Repetitive stimulation increases the activation rate of skeletal muscle Ca2+ currents.

Voltage clamp experiments were conducted in frog skeletal muscle using repetitive stimulation protocols. The activation rate of Ca2+ currents increased by prepulses to depolarizing potentials or by stimulating the fiber with a frequency of 1.7 Hz at 0 mV. The effect was observable with Ca2+ or Ba2+ ions, and was clearly voltage-dependent. Physiologically, it is relevant that such activation rate increase can take place during a train of action potentials.

Action Potentials↗
PubMed1990

Charge movement and calcium currents in skeletal muscle fibers are enhanced by GTP gamma S.

G-proteins play several regulatory roles in the cell. They can modulate ionic channels directly or in association with second messengers. In skeletal muscle, G-proteins modulate the activity of calcium channels either by acting directly on the channel and/or through a cAMP-dependent phosphorylating mechanism. The activation of G-proteins by GTP gamma S can also induce force generation in skinned fibers. In this paper we studied the effect of GTP gamma S on charge movement and calcium currents (ICa) in rat and frog skeletal muscle, using the Vaseline gap technique. We observed an increase in both charge movement and ICa after the intracellular addition of 10-100 microM GTP gamma S. GDP beta S did not have any effect. Addition of protein kinase A catalytic subunit increased the ICa, probably through a phosphorylation process, but did not modify the charge movement. This suggests that protein kinase A and GTP gamma S are acting on different sites of the channel. It can be speculated that G-proteins may have a regulatory role in the excitation-contraction coupling mechanism by a direct effect on charge movement.

Animals↗
PubMed1990

Hormonal regulation of potassium currents in single myometrial cells.

Three potassium currents (IK) were recorded from myometrial cells isolated from the uterus of rats at estrus and diestrus and kept in culture for 1-6 days. IK were differentiated by their modulation with norepinephrine and/or by their onset kinetics. At +50 mV the activation time constants were about 0.7 ms, 6 ms, and 15 ms for the fast, the intermediate, and the slow IK, respectively. Norepinephrine (1 microM) potentiated the fast IK and reduced the intermediate IK. In addition, differences were found with respect to cells from animals at estrus and diestrus. The fast IK was preferentially expressed in cultures from animals at estrus, whereas the intermediate IK was more frequent in cells from rats at diestrus. These results indicate that K+ channels from myometrial cells are multiregulated. Regulation may occur by short-term signals (neurotransmitters) and/or by preferentially expressing distinct types of channels depending on the hormonal status of the animal.

Animals↗
PubMed1990

GTP-dependent regulation of myometrial KCa channels incorporated into lipid bilayers.

The regulation of calcium-activated K (KCa) channels by a G protein-mediated mechanism was studied. KCa channels were reconstituted in planar lipid bilayers by fusion of membrane vesicles from rat or pig myometrium. The regulatory process was studied by exploring the actions of GTP and GTP gamma S on single channel activity. KCa channels had a conductance of 260 +/- 6 pS (n = 25, +/- SE, 250/50 mM KCl gradient) and were voltage dependent. The open probability (Po) vs. voltage relationships were well fit by a Boltzmann distribution. The slope factor (11 mV) was insensitive to internal Ca2+. The half activation potential (V1/2) was shifted -70 mV by raising internal Ca2+ from pCa 6.2 to pCa 4. Addition of GTP or GTP gamma S activated channel activity only in the presence of Mg2+, a characteristic typical of G protein-mediated mechanisms. The Po increased from 0.18 +/- 0.08 to 0.49 +/- 0.07 (n = 7, 0 mV, pCa 6 to 6.8). The channel was also activated (Po increased from 0.03 to 0.37) in the presence of AMP-PNP, a nonphosphorylating ATP analogue, suggesting a direct G protein gating of KCa channels. Upon nucleotide activation, mean open time increased by a factor of 2.7 +/- 0.7 and mean closed time decreased by 0.2 +/- 0.07 of their initial values (n = 6). Norepinephrine (NE) or isoproterenol potentiated the GTP-mediated activation of KCa channels (Po increased from 0.17 +/- 0.06 to 0.35 +/- 0.07, n = 10). These results suggest that myometrium possesses beta-adrenergic receptors coupled to a GTP-dependent protein that can directly gate KCa channels. Furthermore, KCa channels, beta-adrenergic receptors, and G proteins can be reconstituted in lipid bilayers as a stable, functionally coupled, molecular complex.

Adenosine Triphosphate↗
PubMed1990

ANG II inhibits calcium-activated potassium channels from coronary smooth muscle in lipid bilayers.

Angiotensin II (ANG II) is a powerful vasoconstrictor of coronary vessels and other smooth muscles. One of the actions of ANG II is the inhibition of K+ currents, possibly contributing to depolarization and contraction. Therefore, we investigated the role of ANG II on the regulation of K+ channels at the single-channel level. We studied its effect on calcium-activated potassium (KCa) channels (congruent to 250 pS) from coronary smooth muscle incorporated into lipid bilayers. KCa channels were sensitive to externally applied ANG II at voltages from -20 to -70 mV and pCa between 6.5 and 4. The dose-response curve gave a concentration of half-inhibition (Ki1/2) of 58 nM and a Hill coefficient of 2.2, indicating a minimum of two sites in the process. ANG II modified the open and closed states of the channel, affecting their proportions and their values. In addition, a new much slower (congruent to 1 s) closed or "blocked" state appeared. We conclude that one of the mechanisms by which ANG II causes vasoconstriction of the coronary vessels is a direct inhibition of KCa channels contributing to depolarization and contraction.

Angiotensin II↗
PubMed1990

Decreased number of peripheral blood CD4 + CD29+ lymphocytes and increased in vitro spontaneous production of anti-DNA antibodies in patients with active systemic lupus erythematosus.

Flow cytometric 2-color analysis of peripheral blood lymphocytes from patients with systemic lupus erythematosus (SLE) showed a reduction of relative and absolute number of CD4+ CD29+ cells compared to matched healthy individuals. This abnormality was more marked in patients with active/very active disease. Absolute number of CD4+ CD29+ cells was negatively correlated with spontaneous anti-DNA Ig production that we demonstrated to be a laboratory index strongly correlated with a clinical disease activity score. A decrease of the percentage of CD8+ CD29+ lymphocytes in patients with active disease was also observed.

Adult↗
PubMed1990

Functional studies of Ca2+ channels from plasmalemma and sarcoplasmic reticulum membranes in muscle cells.

Physiological and biochemical studies (channel characteristics, intracellular Ca2+ determinations and, channel purification, cloning and expression) of the different components involved in the regulation of intercellular Ca2+ have provided new information about their specific role. Recent information favors a major role for plasmalemma Ca2+ channels in E-C coupling of cardiac muscle, while a major role for sarcoplasmic reticulum Ca2+ release channels (ryanodine receptors) is proposed for E-C coupling of skeletal muscle. In smooth muscle, both plasmalemma and sarcoplasmic reticulum (IP3 receptors) Ca2+ channels are involved in E-C coupling. These studies will be comparatively discussed for skeletal, cardiac and smooth muscle cells.

Animals↗
PubMed1990

[The effects of orthodontic treatment on periodontal findings in adults].

After periodontal therapy 12 patients with advanced alveolar bone loss were orthodontically treated and recalled for approximately 51.1 months. Orthodontic tooth movement resulted in a mean overjet reduction of 1.7 mm, a mean overbite reduction of 1.3 mm and closure of the anterior spaces. No significant differences were observed in the probing depths and attachment levels between the test and control teeth. 31% of the treated teeth showed signs of root resorption. The results indicate that orthodontic therapy in patients with successfully treated advanced periodontal disease does not lead to further attachment loss if the patients are periodontally well maintained.

Adult↗
PubMed1989

[3H]PN200-110 and [3H]ryanodine binding and reconstitution of ion channel activity with skeletal muscle membranes.

Skeletal muscle membranes derived either from the tubular (T) network or from the sarcoplasmic reticulum (SR) were characterized with respect to the binding of the dihydropyridine, [3H]PN200-110, and the alkaloid, [3H]ryanodine; polypeptide composition; and ion channel activity. Conditions for optimizing the binding of these radioligands are discussed. A bilayer pulsing technique is described and is used to examine the channels present in these membranes. Fusion of T-tubule membranes into bilayers revealed the presence of chloride channels and dihydropyridine-sensitive calcium channels with three distinct conductances. The dihydropyridine-sensitive channels were further characterized with respect to their voltage dependence. Pulsing experiments indicated that two different populations of dihydropyridine-sensitive channels existed. Fusion of heavy SR vesicles revealed three different ion channels; the putative calcium release channel, a potassium channel, and a chloride channel. Thus, this fractionation procedure provides T-tubules and SR membranes which, with radioligand binding and single channel recording techniques, provide a useful tool to study the characteristics of skeletal muscle ion channels and their possible role in excitation-contraction coupling.

Alkaloids↗
PubMed1989

Voltage-dependent inactivation of slow calcium channels in intact twitch muscle fibers of the frog.

Inactivation of slow Ca2+ channels was studied in intact twitch skeletal muscle fibers of the frog by using the three-microelectrode voltage-clamp technique. Hypertonic sucrose solutions were used to abolish contraction. The rate constant of decay of the slow Ca2+ current (ICa) remained practically unchanged when the recording solution containing 10 mM Ca2+ was replaced by a Ca2+-buffered solution (126 mM Ca-maleate). The rate constant of decay of ICa monotonically increased with depolarization although the corresponding time integral of ICa followed a bell-shaped function. The replacement of Ca2+ by Ba2+ did not result in a slowing of the rate of decay of the inward current nor did it reduce the degree of steady-state inactivation. The voltage dependence of the steady-state inactivation curve was steeper in the presence of Ba2+. In two-pulse experiments with large conditioning depolarizations ICa inactivation remained unchanged although Ca2+ influx during the prepulse greatly decreased. Dantrolene (12 microM) increased mechanical threshold at all pulse durations tested, the effect being more prominent for short pulses. Dantrolene did not significantly modify ICa decay and the voltage dependence of inactivation. These results indicate that in intact muscle fibers Ca2+ channels inactivate in a voltage-dependent manner through a mechanism that does not require Ca2+ entry into the cell.

Animals↗
PubMed1989

Decay of the slow calcium current in twitch muscle fibers of the frog is influenced by intracellular EGTA.

The mechanism(s) of the decay of slow calcium current (ICa) in cut twitch skeletal muscle fibers of the frog were studied in voltage-clamp experiments using the double vaseline-gap technique. ICa decay followed a single exponential in 10 mM external Ca2+ and 20 mM internal EGTA solutions in all pulse protocols tested: single depolarizing pulses (activation protocol), two pulses (inactivation protocol), and during a long pulse preceded by a short prepulse (400 ms) to 80 mV (tail protocol). In single pulses the rate constant of ICa decay was approximately 0.75 s-1 at 0 mV and became faster with larger depolarizations. ICa had different amplitudes during the second pulses of the inactivation protocol (0 mV) and of the tail protocol (-20 to 40 mV) and had similar time constants of decay. The time constant of decay did not change significantly at each potential after replacing 10 mM Ca2+ with a Ca2+-buffered solution with malate. With 70 mM intracellular EGTA and 10 mM external Ca2+ solutions, ICa also decayed with a single-exponential curve, but it was about four times faster (approximately 3.5 s-1 at 0 mV pulse). In these solutions the rate constant showed a direct relationship with ICa amplitude at different potentials. With 70 mM EGTA, replacing the external 10 mM Ca2+ solution with the Ca2+-buffered solution caused the decay of ICa to become slower and to have the same relationship with membrane potential and ICa amplitude as in fibers with 20 mM EGTA internal solution. The mechanism of ICa decay depends on the intracellular EGTA concentration: (a) internal EGTA (both 20 and 70 mM) significantly reduces the voltage dependence of the inactivation process and (b) 70 mM EGTA dramatically increases the rate of tubular calcium depletion during the flow of ICa.

Animals↗
PubMed1989

Relationship between myoplasmic calcium transients and calcium currents in frog skeletal muscle.

Ca2+ currents (ICa) and myoplasmic Ca2+ transients were simultaneously recorded in single muscle fibers from the semitendinosus muscle of Rana pipiens. The vaseline-gap voltage-clamp technique was used. Ca2+ transients were recorded with the metallochromic indicator dye antipyrylazo III. Ca2+ transients consisted of an early fast rising phase followed by a late slower one. The second phase was increased by experimental maneuvers that enlarged ICa, such as augmenting [Ca2+]o (from 2 to 10 mM) or adding (-)-Bay K 8644 (2 microM). When [Ca2+]o was increased, the second phase of the Ca2+ transients and ICa showed an average increase at 0 mV of 2 +/- 0.9 microM (4) and 1.4 +/- 0.3 mA/ml (4), respectively. (-)-Bay K 8644 increased the late phase of the Ca2+ transients and ICa at 0 mV by 0.8 +/- 0.3 microM (3) and 6.7 +/- 2.0 mA/ml (4), respectively. The initial fast rising phase of the Ca2+ transients was not modified. (-)-Bay K 8644 slowed the time constant of decay of the transients by 57 +/- 6 ms. In other experimental conditions, Ca2+ release from the sarcoplasmic reticulum (SR) was impaired with repetitive stimulation in 1 mM [EGTA]i-containing fibers. Under those circumstances, Ca2+ transients directly followed the time integral of ICa. Pulses to 0 mV caused a large Ca2+ transient that became suppressed when large pulses to 100 mV were applied. In fibers with functioning SR, pulses to 100 mV elicited somewhat smaller or similar amplitude Ca2+ transients when compared with those elicited by pulses to 0 mV. The increase in ICa after raising [Ca2+]o or adding (-)-Bay K 8644 cannot directly explain the change in Ca2+ transients in fibers with functioning SR. On the other hand, when Ca2+ release from the SR is impaired Ca2+ transients depend on ICa.

3-Pyridinecarboxylic acid, 1,4-dihydro-2,6-dimethy↗
PubMed1989

Electrical properties and innervation of fibers in the orbital layer of rat extraocular muscles.

1. The inferior rectus muscle of rat, one of the extraocular muscles, contains two populations of multiply innervated fibers (MIFs): orbital MIFs, located in the orbital layer of the muscle and global MIFs, found in the global layer. The electrical properties and the responses to nerve stimulation of orbital MIFs were studied with single intracellular electrodes and compared with those of twitch fibers of the orbital layer, MIFs of the global layer, and tonic fibers of the frog. 2. About 90% of the orbital MIFs did not produce overshooting action potentials. In these fibers the characteristics and time course of the responses to nerve stimulation varied along the length of the fibers. Within 2 mm of the end-plate band of the muscle, the responses consisted of several small end-plate potentials (EPPs) and a nonovershooting spike. Distal to 2 mm, the responses in most fibers consisted of large and small EPPs with no spiking response. Some fibers produced very small spikes surmounted on large EPPs. 3. Overshooting action potentials were observed in approximately 10% of the orbital MIFs recorded between the end-plate band and 2 mm distal. The presence or absence of action potentials was not related to the magnitude of the resting potential of the fibers. 4. The threshold of nerve stimulated responses in orbital MIFs was the same as that in orbital twitch fibers. A large number of orbital MIFs had latencies equal to those for the orbital twitch fibers recorded at the same distance from the end-plate band, but the average latency was greater in the MIFs. The latency of orbital MIFs was about one-half of that for the MIFs of the global layer. The values for the effective resistance and membrane time constant of orbital MIFs fell between those for orbital twitch fibers on the one hand, and global MIFs and frog tonic fibers on the other. 5. In order to compare electrical properties with innervation patterns, fibers identified electrophysiologically as orbital MIFs were injected with the fluorescent dye Lucifer yellow and then traced in Epon-embedded, serial transverse sections. In addition to numerous superficial endings distributed along the fibers, a single "en plaque" ending was also found in the end-plate band that resembled the end plates of the adjacent orbital twitch fibers. 6. From these results we conclude that the electrical activity of orbital MIFs varies along the length of the fibers.(ABSTRACT TRUNCATED AT 400 WORDS)

Animals↗
PubMed1988

Effect of Ca2+ channel blockers on K+ contractures in twitch fibres of the frog (Rana pipiens).

1. The effects of Ca2+ channel blockers (nifedipine, nitrendipine and diltiazem) were tested on K+ contractures in single muscle fibres of the frog, Rana pipiens. 2. Nifedipine (1 microM) reduced the area under K+ contractures to 24 +/- 9% (4) (100 mM-K+) and 34 +/- 24% (4) (190 mM-K+). Nitrendipine (0.1 microM) reduced the area to 30 +/- 10% (4) (120 mM-K+). The blockade of the contractures was reversible. 3. Diltiazem (1 microM) shortened the first 190 mM-K+ contracture without affecting the peak amplitude. The first contractures, performed at 15-20 min after the removal of diltiazem, were greatly reduced to 29 +/- 14% (4). This effect was reversed after three to five contractures in the absence of the drug. Similar results were obtained with 60 and 100 mM-K+. 4. The resting potential in control saline and after a brief exposure to 120 mM-K+ was not affected by the dihydropyridines and diltiazem. 5. Slow and fast Ca2+ currents were not modified by 1 microM-diltiazem at any stimulation rate or with pre-pulse depolarizations. Diltiazem (50 microM) did not affect the fast Ca2+ current and reduced the slow one to 48 +/- 10% (4). 7. The reduction of K+ contractures by Ca2+ channel blocking agents was not related to a blockade of Ca2+ currents. This can be tentatively explained by interactions of these compounds on membranes sites which regulate the coupling between membrane depolarization and contraction.

Action Potentials↗
PubMed1987

Ca2+ and K+ current in cultured vascular smooth muscle cells from rat aorta.

Ca2+ (ICa) and K+ (IK) currents were recorded in single cultured cells from rat aorta using the whole cell clamp technique with patch electrodes. ICa was detected at -30 mV, and at 20 mV it reached a peak in about 10 ms and decayed with a t1/2 = 50 ms. The mean maximum slope conductance (GCa) was 30 microseconds/cm2. lK was detected at -10 mV and at 20 mV reached its maximum with a t1/2 = 12 ms. For IK, GK = 200 microseconds/cm2. These channels can be activated during action potentials and play a role in the excitation and contraction of vascular smooth muscle cells.

Animals↗
PubMed1987

Appropriate conditions to record activation of fast Ca2+ channels in frog skeletal muscle (Rana pipiens).

Frog skeletal muscle fibres possess slow Ca++ channels which are currently studied. Only recently a fast Ca++ current in frog skeletal muscle was reported. This finding is related to the fact that this current is very labile. Here we report the appropriate conditions to record the fast calcium current (ICa-f) in skeletal muscle of the frog Rana pipiens, in isotonic solutions and resting fibre length.

Animals↗