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Biomedical subjects

R Latorre

Publications and source records attributed to R Latorre.

At least 73 records · Page 4Linked to original sources
PubMed1992

K(+)-channel blockers restore synaptic plasticity in the neuromuscular junction of dunce, a Drosophila learning and memory mutant.

The effects of K(+)-channel blockers on synaptic transmission in dunce (dnc), a Drosophila learning and memory mutant, were investigated. Larvae dnc mutants lack facilitation and post-tetanic potentiation (PTP) at their motor end-plates; dnc mutants are also deficient in a form of phosphodiesterase, and exhibit abnormally high levels of cyclic adenosine 3',5'-monophosphate (cAMP). A two-microelectrode voltage-clamp was used to record end-plate currents and spontaneous end-plate currents from longitudinal ventrolateral third-instar larval muscle. The K(+)-channel blockers 3,4-diaminopyridine (3,4-DAP) and tetraethylammonium (TEA), at micromolar concentrations, caused a reversible decrease in end-plate current amplitudes both in wild-type and mutant end-plates. In the presence of blockers, a period of high-frequency stimulation (tetanus) of the nerve gave way to a transient increase in the end-plate currents of dnc mutants resembling facilitation and PTP in normal end-plates; 3,4-DAP and TEA also restored facilitation and PTP in normal end-plates after incubation with a non-hydrolysable analogue of cAMP (8Br-cAMP). It is suggested that a specific K+ conductance might be relevant to the lack of synaptic plasticity at the dnc neuromuscular synapses.

4-Aminopyridine↗
PubMed1992

Functional reconstitution of a chloride channel protein from bovine trachea.

We characterized the electrophysiological properties of a chloride channel protein isolated from bovine trachea after incorporation into planar lipid bilayers, and studied the effects of thiol-modulating agents on channel regulation both in bilayers and vesicular iodide uptake studies. Our experiments showed that this protein formed perfectly anion-selective channels in the bilayer, with an anion permeability sequence of I- (2.1) > NO3- (1.7) > Br- (1.2) > Cl- (1.0). The conductance of this channel was 25-30 picosiemens in 150 mM Cl-, and saturated with increasing chloride concentration. This channel could be completely inhibited by 4,4'-bis(isothiocyano)-2,2'-stilbenedisulfonate. Immunoblot analysis, using polyclonal antibodies (anti-p38), revealed one major band at 140 kDa. Upon reduction with dithiothreitol, 64- and 38-kDa polypeptides were observed. Functional experiments showed that reduction was accompanied by loss of 125I- uptake and single-channel activity. In the presence of dithiothreitol, only the low molecular mass protein forms (64 and 38 kDa) were detected by anti-p38 antibodies on Western blots. Cross-linking of S-S bonds with Cu(2+)-o-phenanthroline led to activation of chloride channels in vesicles and bilayers. Over-aggregation of chloride channels by this S-S cross-linking reagent caused inhibition of 125I- uptake by 80-100% and the abolishment of single-channel activity. We propose that the native chloride channel from bovine trachea can exist in vivo in different structural and functional forms depending upon its thiol-disulfide oxidation reduction status. The oxidized form has a molecular mass of 140 kDa and represents a fully active chloride channel. Inactivation of this channel might occur by over-aggregation of protein subunits, or by dissociation of the 140-kDa subunit by disulfide bond reduction.

4,4'-Diisothiocyanostilbene-2,2'-Disulfonic Acid↗
PubMed1992

Internal blockade of a Ca(2+)-activated K+ channel by Shaker B inactivating "ball" peptide.

Shaker B inactivating peptide ("ball peptide", BP) interacts with Ca(2+)-activated K+ (KCa) channels from the cytoplasmic side only, producing inhibition of channel activity. This effect was reversible and dose and voltage dependent (stronger at depolarized potentials). The inhibition of KCa channels by BP cannot be mimicked by an inactive point mutation of the BP, L7E. BP binds to KCa channels in a bimolecular reaction (dissociation constant of 95 microM at +40 mV). The binding site is probably located in the internal "mouth" or conduction pathway, since both external K+ and internal tetraethylammonium relieve BP-induced inhibition. These results suggest that KCa channels possess a binding site for the BP with some properties similar to the ball receptor found in Shaker B K+ channels.

Amino Acid Sequence↗
PubMed1992

Mode of action of iberiotoxin, a potent blocker of the large conductance Ca(2+)-activated K+ channel.

Iberiotoxin, a toxin purified from the scorpion Buthus tamulus is a 37 amino acid peptide having 68% homology with charybdotoxin. Charybdotoxin blocks large conductance Ca(2+)-activated K+ channels at nanomolar concentrations from the external side only (Miller, C., E. Moczydlowski, R. Latorre, and M. Phillips. 1985. Nature (Lond.). 313:316-318). Like charybdotoxin, iberiotoxin is only able to block the skeletal muscle membrane Ca(2+)-activated K+ channel incorporated into neutral-planar bilayers when applied to the external side. In the presence of iberiotoxin, channel activity is interrupted by quiescent periods that can last for several minutes. From single-channel records it was possible to determine that iberiotoxin binds to Ca(2+)-activate K+ channel in a bimolecular reaction. When the solution bathing the membrane are 300 mM K+ internal and 300 mM Na+ external the toxin second order association rate constant is 3.3 x 10(6) s-1 M-1 and the first order dissociation rate constant is 3.8 x 10(-3) s-1, yielding an apparent equilibrium dissociation constant of 1.16 nM. This constant is 10-fold lower than that of charybdotoxin, and the values for the rate constants showed above indicate that this is mainly due to the very low dissociation rate constant; mean blocked time approximately 5 min. The fact that tetraethylammonium competitively inhibits the iberiotoxin binding to the channel is a strong suggestion that this toxin binds to the channel external vestibule. Increasing the external K+ concentration makes the association rate constant to decrease with no effect on the dissociation reaction indicating that the surface charges located in the external channel vestibule play an important role in modulating toxin binding.

Amino Acid Sequence↗
PubMed1992

Gating kinetics of batrachotoxin-modified Na+ channels in the squid giant axon. Voltage and temperature effects.

The gating kinetics of batrachotoxin-modified Na+ channels were studied in outside-out patches of axolemma from the squid giant axon by means of the cut-open axon technique. Single channel kinetics were characterized at different membrane voltages and temperatures. The probability of channel opening (Po) as a function of voltage was well described by a Boltzmann distribution with an equivalent number of gating particles of 3.58. The voltage at which the channel was open 50% of the time was a function of [Na+] and temperature. A decrease in the internal [Na+] induced a shift to the right of the Po vs. V curve, suggesting the presence of an integral negative fixed charge near the activation gate. An increase in temperature decreased Po, indicating a stabilization of the closed configuration of the channel and also a decrease in entropy upon channel opening. Probability density analysis of dwell times in the closed and open states of the channel at 0 degrees C revealed the presence of three closed and three open states. The slowest open kinetic component constituted only a small fraction of the total number of transitions and became negligible at voltages greater than -65 mV. Adjacent interval analysis showed that there is no correlation in the duration of successive open and closed events. Consistent with this analysis, maximum likelihood estimation of the rate constants for nine different single-channel models produced a preferred model (model 1) having a linear sequence of closed states and two open states emerging from the last closed state. The effect of temperature on the rate constants of model 1 was studied. An increase in temperature increased all rate constants; the shift in Po would be the result of an increase in the closing rates predominant over the change in the opening rates. The temperature study also provided the basis for building an energy diagram for the transitions between channel states.

Animals↗
PubMed1992

[Pre S2: a marker for chronicity and/or liver damage in hepatitis B infection?].

Pre S2 was investigated using a monoclonal ELISA in 100 samples from 52 patients with different forms of infection with Hepatitis B virus. Pre S2 was present in 18 of 19 patients with acute hepatitis. Its persistence for more than 150 days after beginning of symptoms was associated to a chronic hepatitis state. It was also present in 11 patients with chronic hepatitis or cirrhosis, 15 asymptomatic Hepatitis B carriers and 3 of 4 patients with hepatic carcinoma. Pre S2 became negative in only 1 of 3 patients treated with interferon who had a positive response according to HBe and HBs antigens. Thus, Pre S2 is present in patients with evidence of viral replication. It is more a marker of persistent Hepatitis B infection than of chronic liver damage.

Acute Disease↗
PubMed1992

[Microscopic colitis and collagenous colitis. An entity not yet reported in Chile].

Collagenous and microscopic colitis have been described as causes for abundant watery diarrhea with a normal radiologic and endoscopic aspect of the colonic mucosa. Microscopic colitis is characterized by diffuse intraepithelial lymphocytic infiltration and collagenous colitis by thickening of subepithelial collagen layer greater than 15 microns with or without inflammatory changes of the mucosa. Here are reported 5 patients with microscopic colitis and 4 with collagenous colitis. The mean age was 52 years (range 40 to 68) with an equal sex distribution. Diarrhea was of longstanding duration without weight loss, anemia or hypoalbuminemia. Occasionally the volume of diarrhea was over 1 L a day. One patient had steatorrhea that proved resistant to a gluten free diet. Endoscopic and radiologic findings were normal in every patient and the diagnosis was based on typical histology. Azulfidine therapy was successful in 7 patients and prednisone in another. Colonic biopsy should be performed in every patient with chronic diarrhea.

Adult↗
PubMed1992

[Significance of investigating viral DNA in serum from patients with hepatitis B].

The DNA-HBV was investigated in 109 serum samples from 75 patients with different forms of infection with the HB virus, using an RIA hybridization technique (Genostic-Abott). DNA-HBV was present in 12 of 18 cases of acute hepatitis, 1 of 3 patients with fulminant disease, 11 of 14 patients with chronic hepatitis and/or cirrhosis, 2 of 10 patients with hepatoma and 6 of 30 asymptomatic chronic carriers. Presence of DNA-HBV beyond 60 days in 7 patients with acute hepatitis established the chronic state. The highest levels were found in patients with chronic hepatitis or cirrhosis (mean 41 +/- 11 pg/ml) and the lowest in chronic carriers and patients with hepatoma. High levels of DNA-HBV denote persistent viral replication and would support antiviral treatment. Thus, investigation of DNA-HBV has diagnostic, prognostic and therapeutic implications in patients with Hepatitis B.

DNA, Viral↗
PubMed1992

[Peptic ulcer and endoscopic signs of portal hypertension in patients with primary biliary cirrhosis].

BACKGROUND: Controversies exist on the prevalence of peptic ulcer and on the relationship between the presence of portal hypertension associated lesions in the upper gastrointestinal tract and the histological and functional stages of primary biliary cirrhosis AIM: To evaluate the prevalence of peptic ulcer and endoscopic signs of portal hypertension in subjects with primary biliary cirrhosis. PATIENTS AND METHODS: A retrospective study based on upper gastrointestinal endoscopies in 36 patients with primary biliary cirrhosis and 45 subjects with chronic active hepatitis. SETTING: Gastroenterology Center, Hospital Clínico, University of Chile, Santiago. MAIN RESULTS: No differences were demonstrated between the two groups for the prevalence of peptic ulcer (11% in primary biliary cirrhosis vs 13% in chronic active hepatitis). Esophageal varices were found in 5 of 16 patients (31%) in prefibrotic stages of primary biliary cirrhosis compared with 10 of 20 (50%) in whom fibrosis or cirrhosis were noted; NS. Endoscopic signs suggesting hypertensive gastropathy were present in 2 patients with early primary biliary cirrhosis (13%) and in 5 (25%) in advanced stages. CONCLUSIONS: Peptic ulcer is not more prevalent in patients with primary biliary cirrhosis than in those suffering from chronic active hepatitis. Esophageal varices and endoscopic stigmata of portal hypertension induced changes in gastric mucosa can be observed in early stages of primary biliary cirrhosis.

Endoscopy, Gastrointestinal↗
PubMed1991

A cyclic AMP-activated K+ channel in Drosophila larval muscle is persistently activated in dunce.

Single-channel recording from longitudinal ventrolateral Drosophila larval muscle reveals the presence of a potassium-selective channel that is directly and reversibly activated by cAMP in a dose-dependent fashion. Activation is specific and it cannot be mimicked by a series of agents that include AMP, cGMP, ATP, inositol trisphosphate, and Ca2+. Channel current records obtained from larval muscle in different dunce mutants possessing abnormally high levels of cAMP show that, in the mutants, the channel displays an increased probability of opening.

8-Bromo Cyclic Adenosine Monophosphate↗
PubMed1991

Four cases of direct ion channel gating by cyclic nucleotides.

Four different nucleotide-gated ion channels are discussed in terms of their biophysical properties and their importance in cell physiology. Channels activated directly by cGMP are present in vertebrate and invertebrate photoreceptors. In both cases cGMP increases the fraction of time the channel remains in the open state. At least three cGMP molecules are involved in channel opening in vertebrate photoreceptors and the concentration of the cyclic nucleotide to obtain the half maximal effect is about 15 microM. The light-dependent channel of both vertebrates and invertebrates is poorly cation selective. The vertebrate channel allows divalent cations to pass through 10-15-fold more easily than monovalent ions. In agreement with their preference for divalent cations, this channel is blocked by l-cis Dialtazem, a molecule that blocks certain types of calcium channels. In olfactory neurons a channel activated by both cAMP and cGMP is found and, as in the light-dependent channel, several molecules of the nucleotide are needed to open the channel with a half maximal effect obtained in the range of 1-40 microM. The channel is poorly cationic selective. A K+ channel directly and specifically activated by cAMP is found in Drosophila larval muscle. At least three cAMP molecules are involved in the opening reaction. Half-maximal effect is obtained at about 50 microM. This channel is blocked by micromolar amount of tetraethylammonium applied internally. Interestingly, this channel has a probability of opening 10-20-fold larger in the mutant dunce, a mutant that possesses abnormally elevated intracellular cAMP level, than in the wild type.

Animals↗
PubMed1991

Ion permeation in normal and batrachotoxin-modified Na+ channels in the squid giant axon.

Na+ permeation through normal and batrachotoxin (BTX)-modified squid axon Na+ channels was characterized. Unmodified and toxin-modified Na+ channels were studied simultaneously in outside-out membrane patches using the cut-open axon technique. Current-voltage relations for both normal and BTX-modified channels were measured over a wide range of Na+ concentrations and voltages. Channel conductance as a function of Na+ concentration curves showed that within the range 0.015-1 M Na+ the normal channel conductance is 1.7-2.5-fold larger than the BTX-modified conductance. These relations cannot be fitted by a simple Langmuir isotherm. Channel conductance at low concentrations was larger than expected from a Michaelis-Menten behavior. The deviations from the simple case were accounted for by fixed negative charges located in the vicinity of the channel entrances. Fixed negative charges near the pore mouths would have the effect of increasing the local Na+ concentration. The results are discussed in terms of energy profiles with three barriers and two sites, taking into consideration the effect of the fixed negative charges. Either single- or multi-ion pore models can account for all the permeation data obtained in both symmetric and asymmetric conditions. In a temperature range of 5-15 degrees C, the estimated Q10 for the conductance of the BTX-modified Na+ channel was 1.53. BTX appears not to change the Na+ channel ion selectively (for the conditions used) or the surface charge located near the channel entrances.

Animals↗
PubMed1991

Proton modulation of a Ca(2+)-activated K+ channel from rat skeletal muscle incorporated into planar bilayers.

The effect of pH on the activation of a Ca-activated K+ [K(Ca)] channel from rat skeletal muscle incorporated into planar lipid bilayers was studied. Experiments were done at different intracellular Ca2+ and proton concentrations. Changes in pH modified channel kinetics only from the Ca-sensitive face of the channel. At constant Ca2+ concentration, intracellular acidification induced a decrease in the open probability (Po) and a shift of the channel activation curves toward the right along the voltage axis. The displacement was 23.5 mV per pH unit. This displacement was due to a change in the half saturation voltage (Vo) and not to a change in channel voltage dependence. The shifts in Vo induced by protons appeared to be independent of Ca2+ concentration. The slope of the Hill plot of the open-closed equilibrium vs. pH was close to one, suggesting that a minimum of one proton is involved in the proton-driven channel closing reaction. The change in Po with variations in pH was due to both a decrease in the mean open time (To) and an increase in the mean closed time (Tc). At constant voltage, the mean open time of the channel was a linear function of [Ca2+] and the mean closed time was a linear function of 1/[Ca2+]2. Changes in the internal pH modified the slope, but not the intercept of the linear relations To vs. [Ca2+] and Tc vs. 1/[Ca2+]2. On the basis of these results an economical kinetic model of the effect of pH on this channel is proposed. It is concluded that protons do not affect the open-closed reaction, but rather weaken Ca2+ binding to all the conformational states of the channel. Moreover, competitive models in which Ca2+ and H+ cannot bind to the same open or closed state are inconsistent with the data.

Animals↗
PubMed1991

[Spontaneous bacterial peritonitis: a frequent and recurrent complication in cirrhotic patients with ascites].

One hundred and twenty consecutive patients with liver cirrhosis and ascites were prospectively studied in order to analyze the frequency, clinical and bacteriological features, recurrence, and prognosis of spontaneous bacterial peritonitis (SBP). Two variants of SBP were defined: culture positive SBP and culture negative neutrocytic ascites (CNNA). During a follow-up of 6 +/- 2 months, thirty three episodes in 23 patients were identified. Nineteen episodes had ascites positive cultures (58%). The total mortality rate associated with SBP was 39%. (47% for culture positive form and 29% for CNNA). Seven of 15 cirrhotics who had recovered from a first episode of SBP (46%) had 10 recurrences. Mortality associated with SBP recurrence was 50%. Six-month survival probability was 65% in patients with sterile ascites and 33% in SPB (p < 0.05). Impairment of liver function was present in 23 episodes (70%) but abdominal complaints occurred only in one/third and 4 (12%) were asymptomatic. E coli was the most frequent agent involved in culture positive SBP. We confirm that SBP is a frequent, recurrent and severe complication of ascites in cirrhotics. Episodes of SBP without abdominal symptoms or with a silent course are not infrequent. Then, SBP recognition requires ample use of diagnostic paracentesis.

Adult↗