Acetazolamide prevents vacuolar myopathy in skeletal muscle of K(+) -depleted rats.

BACKGROUND AND PURPOSE: Acetazolamide and dichlorphenamide are carbonic anhydrase (CA) inhibitors effective in the clinical condition of hypokalemic periodic paralysis (hypoPP). Whether these drugs prevent vacuolar myopathy, which is a pathogenic factor in hypoPP, is unknown. The effects of these drugs on the efflux of lactate from skeletal muscle were also investigated. EXPERIMENTAL APPROACH: For 10 days, K(+)-depleted rats, a model of hypoPP, were administered 5.6 mg kg(-1) day(-1) of acetazolamide, dichlorphenamide or bendroflumethiazide (the last is not an inhibitor of CA). Histological analysis of vacuolar myopathy and in vitro lactate efflux measurements were performed in skeletal muscles from treated and untreated K(+)-depleted rats, and also from normokalemic rats. KEY RESULTS: About three times as many vacuoles were found in the type II fibres of tibialis anterioris muscle sections from K(+)-depleted rats as were found in the same muscle from normokalemic rats. In ex vivo experiments, a higher efflux of lactate on in vitro incubation was found in muscles of K(+)-depleted rats compared with that found in muscles from normokalemic rats. After treatment of K(+)-depleted rats with acetazolamide, the numbers of vacuoles in tibialis anterioris muscle decreased to near normal values. Incubation with acetazolamide in vitro inhibited efflux of lactate from muscles of K(+)-depleted rats. In contrast, bendroflumethiazide and dichlorphenamide failed to prevent vacuolar myopathy after treatment in vivo and failed to inhibit lactate efflux in vitro. CONCLUSIONS AND IMPLICATIONS: Acetazolamide prevents vacuolar myopathy in K(+)-depleted rats. This effect was associated with inhibition of lactate transport, rather than inhibition of CA.

Impairment of skeletal muscle adenosine triphosphate-sensitive K+ channels in patients with hypokalemic periodic paralysis.

The adenosine triphosphate (ATP)-sensitive K+ (KATP) channel is the most abundant K+ channel active in the skeletal muscle fibers of humans and animals. In the present work, we demonstrate the involvement of the muscular KATP channel in a skeletal muscle disorder known as hypokalemic periodic paralysis (HOPP), which is caused by mutations of the dihydropyridine receptor of the Ca2+ channel. Muscle biopsies excised from three patients with HOPP carrying the R528H mutation of the dihydropyridine receptor showed a reduced sarcolemma KATP current that was not stimulated by magnesium adenosine diphosphate (MgADP; 50-100 microM) and was partially restored by cromakalim. In contrast, large KATP currents stimulated by MgADP were recorded in the healthy subjects. At channel level, an abnormal KATP channel showing several subconductance states was detected in the patients with HOPP. None of these were surveyed in the healthy subjects. Transitions of the KATP channel between subconductance states were also observed after in vitro incubation of the rat muscle with low-K+ solution. The lack of the sarcolemma KATP current observed in these patients explains the symptoms of the disease, i.e., hypokalemia, depolarization of the fibers, and possibly the paralysis following insulin administration.

A new benzoxazine compound blocks KATP channels in pancreatic beta cells: molecular basis for tissue selectivity in vitro and hypoglycaemic action in vivo.

BACKGROUND AND PURPOSE: The 2-propyl-1,4 benzoxazine (AM10) shows a peculiar behaviour in skeletal muscle, inhibiting or opening the ATP-sensitive K(+) (KATP) channel in the absence and presence of ATP, respectively. We focused on tissue selectivity and mechanism of action of AM10 by testing its effects on pancreatic KATP channels by means of both in vitro and in vivo investigations. EXPERIMENTAL APPROACH: In vitro, patch-clamp recordings were performed in native pancreatic beta cells and in tsA201 cells expressing the Kir6.2 Delta C36 channel. In vivo, an intraperitoneal glucose tolerance test was performed in normal mice. KEY RESULTS: In contrast with what observed in the skeletal muscle, AM10, in whole cell perforated mode, did not augment KATP current (I(KATP)) of native beta cells but it inhibited it in a concentration-dependent manner (IC(50): 11.5 nM; maximal block: 60%). Accordingly, in current clamp recordings, a concentration-dependent membrane depolarization was observed. On excised patches, AM10 reduced the open-time probability of KATP channels without altering their single channel conductance; the same effect was observed in the presence of trypsin in the bath solution. Moreover, AM10 inhibited, in an ATP-independent manner, the K(+) current resulting from expressed Kir6.2 Delta C36 (maximal block: 60% at 100 microM; IC(50): 12.7 nM) corroborating an interaction with Kir. In vivo, AM10 attenuated the glycemia increase following a glucose bolus in a dose-dependent manner, without, at the dose tested, inducing fasting hypoglycaemia. CONCLUSION AND IMPLICATIONS: Altogether, these results help to gain insight into a new class of tissue specific KATP channel modulators.

Inhaled tobramycin in children with acute bacterial rhinopharyngitis.

Antibiotic abuse for treating rhinopharyngitis induces the occurrence of resistant bacteria. As topical drugs might reduce this phenomenon, the aims of our study were to evaluate inhaled tobramycin in children with acute bacterial rhinopharyngitis and to compare it with oral amoxicillin/clavulanate. The trial was conducted as randomized, parallel group and double blind. Children, aged 3-6 years, with acute bacterial rhinopharyngitis were treated with 15 mg of aerosolized tobramycin (Group A) or 50 mg/Kg of amoxicillin/clavulanate (Group B) twice daily for 10 days. The following parameters were assessed: nasal obstruction, mucopurulent rhinorrhea, post-nasal drip, adenoidal hypertrophy, tympanic inflammation, tympanogram, rhinomanometry and cultures. Of 416 patients screened, 311 children (178 females and 133 males), median age 4.5 years, completed the study: 156 in Group A and 155 in Group B. Both treatments improved all parameters (p<0.01 for all). Intergroup analysis showed that inhaled tobramycin induced a better improvement versus amoxicillin/clavulanate concerning nasal obstruction (p<0.05), adenoidal hypertrophy (p<0.01), tympanic inflammation (p<0.01), rhinomanometry (p<0.01) and cultures (p<0.05). In conclusion, inhaled tobramycin may represent a valid treatment for acute bacterial rhinopharyngitis in children, as it is effective, safe, economic and simple to use.

Accelerated Infliximab Infusion: Safety, Factors Predicting Adverse Events, Patients' Satisfaction and Cost Analysis. A Cohort Study in IBD Patients.

BACKGROUND: Standard Infliximab infusion consists of a 2-hour intravenous administration. Recently, Infliximab shortened infusion has been included in the Infliximab label as possible maintenance regimen for patients tolerating Infliximab induction therapy. AIM: To verify if accelerated 1-hour Infliximab infusions are as safe as standard administrations, in patients with Inflammatory Bowel Disease. METHODS: Seventy-four patients treated between September 2008 and November 2014 were evaluated. Patients were eligible for 1-hour infusion if they had no history of infusion reactions during the previous 2-hour infusions. RESULTS: Twenty-three patients received 2-hour infusions, 16 patients received 1-hour infusions, 35 patients received 2-hour infusions followed by 1-hour infusions. A total of 1,123 Infliximab infusions were administered. The proportion of patients experiencing infusion reaction was: 4% over the 1-hour infusions and 9% over the 2-hour (P = 0.318). Adverse reaction/infusion rate was 0.55% over the 1-hour infusions and 0.66% over the 2-hour (P = 0.835). In the logistic model, accelerated infusion was the only statistically significant predictor of infusion reaction risk reduction (-90%; P = 0.024). Mean satisfaction was 8/10 (±0.84) with 1-hour regimen and 6/10 (±0.56) with 2-hour infusions (P = 0.000). The mean total cost was reduced by 47% with the 1-hour regimen (133.54€ and 250.86€ for 1-hour and 2-hour infusions, respectively). CONCLUSIONS: Accelerated Infliximab infusion does not increase the acute infusion reaction incidence. In patients with inflammatory bowel disease, the 1-hour regimen should be preferred to 2-hour protocol also due to positive effects on indirect costs and patient's satisfaction.

Taurine blocks ATP-sensitive potassium channels of rat skeletal muscle fibres interfering with the sulphonylurea receptor.

Taurine is a sulphonic aminoacid present in high amounts in various tissues including cardiac and skeletal muscles showing different properties such as antioxidative, antimyotonic and anti-schaemic effects. The cellular mechanism of action of taurine is under investigation and appears to involve the interaction of the sulphonic aminoacid with several ion channels. Using the patch-clamp technique we studied the effects of taurine in rat skeletal muscle fibres on ATP-sensitive K(+) channel (K(ATP)) immediately after excision and on channels that underwent rundown. The cytoplasmic application of 20 mM of taurine reduced the K(ATP) current; this effect was reverted by washout of the drug solution. In this experimental condition the IC(50) was 20.1 mM. After rundown, taurine inhibited the K(ATP) current with similar efficacy. Competition experiments showed that taurine shifted the dose-response inhibition curve of glybenclamide to the left on the log-dose axis without significantly affecting those of ATP or Ca(2+) ion. Single channel recording revealed that taurine affects the close state of the channel prolonging it and reducing the bursts duration. Our data indicate that taurine inhibits the muscular K(ATP) channel interfering with the glybenclamide site on the sulphonylurea receptor of the channel or on the site allosterically coupled to it. During ischaemia and hypoxia, the skeletal and heart muscles undergo several changes; for example, the activation of K(ATP) channels and loss of the intracellular taurine content. The depletion of taurine during ischaemia would contribute to the early activation of K(ATP) channels and salvage the intracellular ATP content.

Effects of mexiletine on ATP sensitive K+ channel of rat skeletal muscle fibres: a state dependent mechanism of action.

1. The effects of mexiletine were evaluated on the ATP-sensitive K+ channel (K(ATP)) of rat skeletal muscle fibres using patch clamp techniques. The effects of mexiletine were studied on macropatch currents 20 s (maximally activated), 8 min (early stage of rundown) and 15 min (late stage of rundown) after excision in the absence or in the presence of internal ADP (50-100 microM) or UDP (500 microM). In addition, the effects of mexiletine were tested on single channel. 2. In the absence of ADP and UDP, mexiletine inhibited the current through maximally activated channels with an IC50 of -5.58+/-0.3 M. Nucleoside diphosphates shifted the current versus mexiletine concentration relationship to the right on the log concentration axis. UDP (500 microM) was more efficacious than ADP (50-100 microM) in this effect. 3. At the early stage of rundown, the sensitivity of the channel to mexiletine was reduced and nucleoside diphosphates, particularly UDP, antagonized the effect of mexiletine. At the late stage of rundown, mexiletine did not affect the currents. 4. At the single channel level, 1 microM mexiletine reduced the mean burst duration by 63% and prolonged the arithmetic mean closed time intervals between the bursts of openings without altering the open time and closed time distributions. Mexiletine did not affect the single channel conductance. 5. These results show that in skeletal muscle, mexiletine is a state-dependent K(ATP) channel inhibitor which either acts through the nucleotide binding site or a site allosterically coupled to it.

Different sulfonylurea and ATP sensitivity characterizes the juvenile and the adult form of KATP channel complex of rat skeletal muscle.

We have described here the changes of the biophysical and pharmacological properties of the sarcolemmal ATP-sensitive K+ channels (KATP) of rat skeletal muscle fibres, occurring from an early postnatal period (5 days) to adulthood (210 days). The age-dependent changes of the mean current of the KATP channel (channel activity) and the effects of the blockers, ATP and glybenclamide, were examined by using the patch-clamp technique. Measurements of the single channel conductance, open probability and channel density were also performed. Excision of cell-attached patches into an ATP-free solution dramatically increased the KATP channel activity; however, the intensity of this activity was age dependent. The relative activity was low at 5-6 days of postnatal life, increased to a plateau at 12-13 days, then declined toward adult values after 37 days. Two distinct types of the KATP channel complex could be distinguished. The early developmental period (5-6 days) was dominated by a KATP channel having a conductance of 66 pS, a high open probability of 0.602, and an IC50 for ATP and glybenclamide of 123.1 microM and 3.97 microM, respectively. This type of channel disappeared with maturation of the muscle to be replaced by the adult form of the KATP channel. The later developmental period (from 56 days) was dominated by a KATP channel having a 71 pS conductance, but a low open probability of 0.222. This adult channel was also 3.2 and 73.5 times more sensitive to ATP and glybenclamide, respectively. We have also observed that the sensitivity of the KATP channel to ATP and glybenclamide develops differently. Indeed, the greater increase in the sensitivity of the channel to ATP was observed between 5 and 12 days of age. Conversely, the greater enhancement of the sensitivity of the channel to glybenclamide occurred between 12 and 37 days. A further increase of this parameter was also observed between 37 and 56 days of age. The differential age-dependent acquisition of the sensitivity of KATP channels to ATP and glybenclamide poses the hypothesis that in rat skeletal muscle the ATP regulatory site and sulfonylurea site are located on different subunits of the KATP channel complex. The intense KATP channel activity recorded between 12 and 37 days of postnatal life sustains the high resting macroscopic K+ conductance characteristic of the early postnatal development.

Effects of high energy phosphates and L-arginine on the electrical parameters of ischemic-reperfused rat skeletal muscle fibers.

In skeletal muscle, 4 h of ischemia followed by 30 min of reperfusion depolarizes the fibers, markedly increases the Cl- and glibenclamide-sensitive K+ conductances and reduces the excitability of the fibers. The ischemia-reperfusion also significantly decreases the ATP content of the muscles. In the present work, the electrical parameters of reperfused extensor digitorum longus muscle of rats were measured in vitro at 30 degrees C, by a computerized two-intracellular microelectrode technique, before and after in vivo pretreatment with equimolar doses of phosphocreatine disodium salt tetrahydrate, phosphocreatine di-L-arginine salt and L-arginine hydrochloride. In the same experimental situations the ATP content of the muscles was also measured. Both phosphocreatine salts prevented the increase of membrane ion conductance due to muscle reperfusion by preloading the muscle fibers with extra ATP. Phosphocreatine disodium salt also prevented the depolarization and restored the normal excitability of the reperfused fibers. In contrast, phosphocreatine di-L-arginine salt did not restore the resting potential nor the excitability of the fibers, but it decreased the amplitude of the action potential by reducing the overshoot. The pretreatment with L-arginine also failed to protect the electrical parameters of the fibers from the ischemic-reperfusion insult. Furthermore, the L-amino acid produced a more pronounced reduction of the excitability of the fibers by increasing the threshold current needed to elicit an action potential and reducing it overshoot. The in vitro application of L-arginine to the muscle also reduced the overshoot of the action potential, suggesting a direct interaction of the L-amino acid with Na+ channels.

Effects of taurine analogues on chloride channel conductance of rat skeletal muscle fibers: a structure-activity relationship investigation.

In rat skeletal muscle, taurine was proposed to interact with a low affinity binding site on sarcolemmal phospholipids near chloride channel, increasing chloride conductance (GCl). In an attempt to evaluate the structure-activity relationship between taurine and its binding site, a series of N-azacycloalkenyl analogues of taurine (A: N-(1'aza-cyclohepten-2'yl)-2-aminoethane sulfonic acid; B: N-(1'-aza-cyclopenten-2'-yl)-2-aminoethane sulfonic acid; C: N-(1'-aza-cyclohepten-2'-yl)-3-amino-propane sulfonic acid; D: N-(1'aza-cyclopenten-2'-yl)-3-aminopropane sulfonic acid) have been synthetized and tested in vitro on rat extensor digitorum longus (EDL) muscle. In spite of the presence of a bulky and lipophilic 5 or 7 membered heterocycle linked to the taurine amino group, analogues A and B determined an increase of GCl, although less potently than taurine. Also 3-amino-propane sulfonic acid (homotaurine), tested in comparison, showed less activity in increasing GCl with respect to taurine, probably for the increased distance between charged groups. Taurine analogues C and D, which differ from compounds A and B for an additional methylene group, showed much lower activity in increasing GCl. It has been reported that guanidinoethane sulfonate (GES) displaces taurine from the low affinity site on sarcolemma by only 7%. This compound, characterized by lower charge density on the guanidinium cationic head, applied in vitro on EDL muscle, show reduced taurine-like activity in increasing GCl.(ABSTRACT TRUNCATED AT 250 WORDS)

Management of chronic otitis media with effusion: the role of glutathione.

BACKGROUND: The inflammatory cells documented in chronic otitis media with effusion (OME) spontaneously release oxidants which can induce middle ear (ME) epithelial cell damage. Glutathione (GSH), a major extracellular antioxidant in humans, plays a central role in antioxidant defense. PURPOSE: To evaluate the effects of GSH treatment on chronic otitis media with effusion (OME). SUBJECTS AND INTERVENTION: Sixty children with chronic OME were enrolled, 30 of whom were randomly assigned to the treatment group and 30 to the placebo group. Patients in the treatment group received 600 mg glutathione in 4 mL saline per day subdivided into five 2-minute administrations given by nasal aerosol every 3 or 4 waking hours for 2 weeks. Patients in the control group received 4 mL saline per day following the same procedure as for GSH treatment. RESULTS: Three months after therapy improvement had occurred in 66.6% of patients in the GSH-treated group and in 8% of the control subjects (P <.01). CONCLUSION: On the basis of these results, GSH treatment could be considered for the nonsurgical management of chronic OME.

Specific binding of nicergoline on an alpha1-like adrenoreceptor in the rat retina.

Systemic treatment with nicergoline, an ergoline derivative showing alpha1-antagonist properties, causes vasodilatation in the eye without apparent untoward cardiovascular effects. In the present work we investigated the ability of nicergoline to inhibit the binding of radiolabelled prazosin in the rat retina and cortex. We found that nicergoline inhibited [3H]prazosin binding in both tissues, being more potent than unlabelled prazosin in the retinal tissue. The competition curves of the ergoline derivative were well fitted by a one-site model in the cortical tissue, with an IC50 (concentration of the drugs needed to inhibit the binding of labelled prazosin by 50%) of 2.54 x 10(-8) M, and by a two-site model in the retinal tissue, with IC50 values of 7.08 x 10(-12) M and 1.82 x 10(-5) M. 2-(2,6 dimetoxyphenoxyethyl) aminomethyl-1,4-benzodioxane hydrochloride (WB4101) and phentolamine, selective ligands for the high-affinity binding site for prazosin, in particular the alpha1A-site, fully inhibited prazosin binding in the cortex but only partially inhibited prazosin binding in the retina, being less potent in this tissue than either nicergoline or prazosin. Our results suggest that a binding component of alpha1-adrenoreceptors is expressed to a lesser extent in the retina than the cortex, leading to a reduced response of the retinal tissue to prazosin, and more particularly to WB4101 and phentolamine. The selective binding of the nicergoline on this retinal adrenoreceptor may explain the peculiar efficacy of the drug in ocular pathophysiology.

Developmental changes of membrane electrical properties of rat skeletal muscle fibers produced by prenatal exposure to carbon monoxide.

The effects of prenatal exposure to 75 and 150 ppm of carbon monoxide (CO) were evaluated in vitro on the electrical parameters of extensor digitorum longus muscle of newborn rats, by means of computerized two intracellular microelectrode technique. Within the first 2 months of postnatal life the muscles from offspring of mothers exposed to 150 ppm CO showed lower values of resting chloride conductance (G(CI)) with respect to age-matched controls, but no significant differences were recorded after 80 days of life. Between 40-200 days of life, the extensor digitorum longus muscles from offspring of 150 ppm CO exposed rats showed a pharmacological response to a specific chloride channel modulator, the R-(+)-enantiomer of 2-(p-chlorophenoxy)propionic acid (R-(+)-CPP), different from that of age-matched controls, but similar to that of normal developing rats aged 8-20 days. The prenatal exposure to 150 ppm CO also slightly delayed the developmental reduction of resting potassium conductance (G(K)); indeed higher values of G(K) were recorded with respect to controls until 60 days of postnatal life. In the offspring of 150 ppm CO treated rats a slight and reversible modification of the excitability parameters related to G(CI) were also observed along with a decreased sensitivity to tetrodotoxin. In the muscle from offspring of 75 ppm CO treated animals G(CI) and G(K) were not different with respect to controls, at any age; however the pharmacological modulation of G(CI) by R-(+)-CPP was similar to that observed in the rats prenatally exposed 150 ppm CO. These results suggest that the prenatal exposure to mild concentrations of CO produces developmental alterations of the ion channels responsible for the passive and active membrane electrical properties of rat skeletal muscle fibers.

Carboxylic acids and chloride conductance in skeletal muscle: influence on the pharmacological activity induced by the chain substituents and the distance between the phenolic group and the carboxylic function in 4-chloro-phenoxy alkanoic acids.

Some analogues of 2-(4-chloro-phenoxy)-propionic acid have been synthesized to evaluate the influence on the skeletal muscle chloride conductance of the distance between the phenolic and the carboxylic groups and/or the presence of substituents and chiral centers differently located on the carbon chain. Absolute configuration and/or chiroptical properties of the chiral compounds synthesized have been determined. These compounds, differently from fibrates, show a very low activity on chloride conductance and no remarkable correlation of this parameter with the absolute configuration.

Cyclic amidine analogues of taurine and homotaurine: synthesis and effects on rat skeletal muscle.

A series of amidinosulfonic acids IIIa-f, analogs of taurine and homotaurine, was synthesized by reacting 2-aminoethane or 3-aminopropane sulfonic acid with lactim ethers Ia-c and then cyclized to sultams IVa-c and Vd-f. The effects of some selected amidinosulfonic acids III on membrane ionic conductances of rat skeletal muscle are described.

[Mucociliary clearance after aerobic exertion in athletes]. / Il trasporto mucociliare dopo sforzo aerobico in atleti.

The Authors studied the modifications in nasal mucociliary clearance times before and after aerobic exertion in athletes. A total of 60 athletes with high-level training (age range 18-37 years) were selected for this study. Persons who smoked or had allergies, nasal sinus phlogosis or tumors, altered nasal cavity morphology (i.e. deviation of the septum and/or hypertrophy of the turbinates), fever or who were taking topic and/or systemic drugs or had previously undergone head and neck surgery were all ruled out of the study. The authors then studied the mucociliary clearance time (MCCT) in these subjects using the saccharin test. This test involves placing a small amount of saccharin on the medial face of the lower turbinate, approximately 1.5 cm from the anterior end and then evaluating the time that elapses before the patient perceives the sweet taste. This test was performed: in 30 subjects one hour prior to and 15 minutes after physical aerobic exertion; in the remaining subjects (controls) the test was performed twice with a 75 minute interval between them. The results showed that the mucociliary clearance time increased after exertion which was, on the average, 11.29 minutes. On the basis of these data, the Authors discuss the likely causes for the detected increase, and correlate it to changes in ventilation and nasal secretion viscosity during physical exercise. According to previous research, these variations are also found in untrained subjects who undergo physical exertion; for this reason, the Authors conclude that nasal clearance is not significantly affected by training.