Statins and fenofibrate affect skeletal muscle chloride conductance in rats by differently impairing ClC-1 channel regulation and expression.

BACKGROUND AND PURPOSE: Statins and fibrates can produce mild to life-threatening skeletal muscle damage. Resting chloride channel conductance (gCl), carried by the ClC-1 channel, is reduced in muscles of rats chronically treated with fluvastatin, atorvastatin or fenofibrate, along with increased resting cytosolic calcium in statin-treated rats. A high gCl, controlled by the Ca(2+)-dependent protein kinase C (PKC), maintains sarcolemma electrical stability and its reduction alters muscle function. Here, we investigated how statins and fenofibrate impaired gCl. EXPERIMENTAL APPROACH: In rats treated with fluvastatin, atorvastatin or fenofibrate, we examined the involvement of PKC in gCl reduction by the two intracellular microelectrodes technique and ClC-1 mRNA level by quantitative real time-polymerase chain reaction. Direct drug effects were tested by patch clamp analysis on human ClC-1 channels expressed in human embryonic kidney (HEK) 293 cells. KEY RESULTS: Chelerythrine, a PKC inhibitor, applied in vitro on muscle dissected from atorvastatin-treated rats fully restored gCl, suggesting the involvement of this enzyme in statin action. Chelerythrine partially restored gCl in muscles from fluvastatin-treated rats but not in those from fenofibrate-treated rats, implying additional mechanisms for gCl impairment. Accordingly, a decrease of ClC-1 channel mRNA was found in both fluvastatin- and fenofibrate-treated rat muscles. Fenofibric acid, the in vivo metabolite of fenofibrate, but not fluvastatin, rapidly reduced chloride currents in HEK 293 cells. CONCLUSIONS AND IMPLICATIONS: Our data suggest multiple mechanisms underlie the effect of statins and fenofibrate on ClC-1 channel conductance. While statins promote Ca(2+)-mediated PKC activation, fenofibrate directly inhibits ClC-1 channels and both fluvastatin and fenofibrate impair expression of mRNA for ClC-1.

Role of tumour necrosis factor alpha, but not of cyclo-oxygenase-2-derived eicosanoids, on functional and morphological indices of dystrophic progression in mdx mice: a pharmacological approach.

The role of tumour necrosis factor (TNF)-alpha or cyclo-oxygenase-2 (COX-2) eicosanoids in dystrophinopathies has been evaluated by chronically treating (4-8 weeks) adult dystrophic mdx mice with the anti-TNF-alpha etanercept (0.5 mg/kg) or the COX-2 inhibitor meloxicam (0.2 mg/kg). Throughout the treatment period the mdx mice underwent a protocol of exercise on treadmill in order to worsen the pathology progression; gastrocnemious muscles from exercised mdx mice showed an intense staining for TNF-alpha by immunohistochemistry. In vivo, etanercept, but not meloxicam, contrasted the exercise-induced forelimb force drop. Electrophysiological recordings ex vivo, showed that etanercept counteracted the decrease in chloride channel function (gCl), a functional index of myofibre damage, in both diaphragm and extensor digitorum longus (EDL) muscle, meloxicam being effective only in EDL muscle. None of the drugs ameliorated calcium homeostasis detected by electrophysiology and/or spectrofluorimetry. Etanercept, more than meloxicam, effectively reduced plasma creatine kinase (CK). Etanercept-treated muscles showed a reduction of connective tissue area and of pro-fibrotic cytokine TGF-beta1 vs. untreated ones; however, the histological profile was weakly ameliorated. In order to better evaluate the impact of etanercept treatment on histology, a 4-week treatment was performed on 2-week-old mdx mice, so to match the first spontaneous degeneration cycle. The histology profile of gastrocnemious was significantly improved with a reduction of degenerating area; however, CK levels were only slightly lower. The present results support a key role of TNF-alpha, but not of COX-2 products, in different phases of dystrophic progression. Anti-TNF-alpha drugs may be useful in combined therapies for Duchenne patients.

Effects of chronic treatment with statins and fenofibrate on rat skeletal muscle: a biochemical, histological and electrophysiological study.

BACKGROUND AND PURPOSE: Skeletal muscle injury by hypolipidemic drugs is not fully understood. An extensive analysis of the effect of chronic treatment with fluvastatin (5 mgkg(-1) and 20 mgkg(-1)), atorvastatin (10 mgkg(-1)) and fenofibrate (60 mgkg(-1)) on rat skeletal muscle was undertaken. EXPERIMENTAL APPROACH: Myoglobinemia as sign of muscle damage was measured by enzymatic assay. Histological and immunohistochemical techniques were used to estimate muscle integrity and the presence of aquaporin-4, a protein controlling water homeostasis. Electrophysiological evaluation of muscle Cl(-) conductance (gCl) and mechanical threshold (MT) for contraction, index of intracellular calcium homeostasis, was performed by the two-intracellular microelectrodes technique. KEY RESULTS: Fluvastatin (20 mgkg(-1)) increased myoglobinemia. The lower dose of fluvastatin did not modify myoglobinemia, but reduced urinary electrolytes, suggesting direct effects on renal function. Atorvastatin also increased myoglobinemia, with slight effects on urinary parameters. No treatment caused any histological damage to muscle or modification in the number of fibres expressing aquaporin-4. Either fluvastatin (at both doses) or atorvastatin reduced sarcolemma gCl and changed MT. Both statins produced slight effects on total cholesterol, suggesting that the observed modifications occur independently of HMGCoA-reductase inhibition. Fenofibrate increased myoglobinemia and decreased muscle gCl, whereas it did not change the MT, suggesting a different mechanism of action from the statins. CONCLUSIONS AND IMPLICATIONS: This study identifies muscle gCl and MT as early targets of drugs action that may contribute to milder symptoms of myotoxicity, such as muscle cramps, while the increase of myoglobinemia is a later phenomenon.

Skeletal muscle disuse induces fibre type-dependent enhancement of Na(+) channel expression.

Slow-twitch and fast-twitch muscle fibres have specific contractile properties to respond to specific needs. Since sodium current density is higher in fast-twitch than in slow-twitch fibres, sodium channels contribute to the phenotypic feature of myofibres. Phenotype determination is not irreversible: after periods of rat hindlimb unloading (HU), a model of hypogravity, a slow-to-fast transition occurs together with atrophy in the antigravity slow-twitch soleus muscle. Using cell-attached patch-clamp and northern blot analyses, we looked at sodium channel expression in soleus muscles after 1-3 weeks of HU in rats. We found that sodium channels in fast-twitch flexor digitorum brevis muscle fibres, soleus muscle fibres and 1- to 3-week HU soleus muscle fibres showed no difference in unitary conductance, open probability and voltage-dependencies of activation, fast inactivation and slow inactivation. However, muscle disuse increased sodium current density in soleus muscle fibres 2-fold, 2.5-fold and 3-fold after 1, 2 and 3 weeks of HU, respectively. The concentration of mRNA for the skeletal muscle sodium channel alpha subunit increased 2-fold after 1 week of HU but returned to the control level after 3 weeks of HU. In contrast, the concentration of mRNA for the ubiquitous sodium channel beta(1) subunit was unchanged after 1 week and had increased by 30% after 3 weeks of HU. The tetrodotoxin sensitivity of sodium currents in 3-week HU soleus muscles and the lack of mRNA signal for the juvenile skeletal muscle sodium channel alpha subunit excluded denervation in our experiments. The observed increase in sodium current density may reduce the resistance to fatigue of antigravity muscle fibres, an effect that may contribute to muscle impairment in humans after space flight or after long immobilization.

Alteration of excitation-contraction coupling mechanism in extensor digitorum longus muscle fibres of dystrophic mdx mouse and potential efficacy of taurine.

No clear data is available about functional alterations in the calcium-dependent excitation-contraction (e-c) coupling mechanism of dystrophin-deficient muscle of mdx mice. By means of the intracellular microelectrode "point" voltage clamp method, we measured the voltage threshold for contraction (mechanical threshold; MT) in intact extensor digitorum longus (EDL) muscle fibres of dystrophic mdx mouse of two different ages: 8 - 12 weeks, during the active regeneration of hind limb muscles, and 6 - 8 months, when regeneration is complete. The EDL muscle fibres of 8 - 12-week-old wildtype animals had a more negative rheobase voltage (potential of equilibrium for contraction- and relaxation-related calcium movements) with respect to control mice of 6 - 8 months. However, at both ages, the EDL muscle fibres of mdx mice contracted at more negative potentials with respect to age-matched controls and had markedly slower time constants to reach the rheobase. The in vitro application of 60 mM taurine, whose normally high intracellular muscle levels play a role in e-c coupling, was without effect on 6 - 8-month-old wildtype EDL muscle, while it significantly ameliorated the MT of mdx mouse. HPLC determination of taurine content at 6 - 8 months showed a significant 140% rise of plasma taurine levels and a clear trend toward a decrease in amino acid levels in hind limb muscles, brain and heart, suggesting a tissue difficulty in retaining appropriate levels of the amino acid. The data is consistent with a permanent alteration of e-c coupling in mdx EDL muscle fibres. The alteration could be related to the proposed increase in intracellular calcium, and can be ameliorated by taurine, suggesting a potential therapeutic role of the amino acid.

Pharmacological characterization of chloride channels belonging to the ClC family by the use of chiral clofibric acid derivatives.

The enantiomers of 2-(p-chlorophenoxy)propionic acid (CPP) and of its analogs with substitutions on the asymmetric carbon atom were tested on human ClC-1 channel, the skeletal muscle chloride channel, after heterologous expression in Xenopus laevis oocytes, to gain insight in the mechanism of action of these stereoselective modulators of macroscopic chloride conductance (gCl) of rat striated fibers. By means of two microelectrode voltage clamp recordings, we found that S(-)-CPP shifted the activation curve of the ClC-1 currents toward more positive potentials and decreased the residual conductance at negative membrane potential; both effects probably account for the decrease of gCl at resting potential of native muscle fibers. Experiments on expressed Torpedo marmorata ClC-0 channels and a mutant lacking the slow gate suggest that S(-)-CPP could act on the fast gate of the single protochannels constituting the double-barreled structure of ClC-0 and ClC-1. The effect of S(-)-CPP on ClC-1 was markedly increased at low external pH (pH = 6), possibly for enhanced diffusion through the membrane (i.e., because the compound was effective only when applied to the cytoplasmic side during patch clamp recordings). The R(+)-isomer had little effect at concentrations as high as 1 mM. The CPP analogs with an ethyl, a phenyl, or an n-propyl group in place of the methyl group on the asymmetric center showed a scale of potency and a stereoselective behavior on ClC-1 similar to that observed for blocking gCl in native muscle fibers. The tested compounds were selective toward the ClC-1 channel. In fact, they were almost ineffective on an N-terminal deletion mutant of ClC-2 that is volume- and pH-independent while they blocked wild-type ClC-2 currents only at high concentrations and independently of pH and drug configuration, suggesting a different mechanism of action compared with ClC-1. No effects were observed on ClC-5 that shows less than 30% homology with ClC-1. Thus, CPP-like compounds may be useful both to gain insight into biophysical properties of ClC-1 and for searching tissue-specific therapeutic agents.

Homologation of mexiletine alkyl chain and stereoselective blockade of skeletal muscle sodium channels.

The optical isomers (-)-(S)- and (+)-(R)-3-(2, 6-dimethylphenoxy)-2-methyl-1-propanamine (Me2), homologues of the antiarrhythmic and antimyotonic drug mexiletine (Mex), were synthesized and assayed as new potential antimyotonic agents. As observed with Mex, Me2 exhibits an enantioselective behaviour. Tests carried out on sodium currents of single muscle fibres of Rana esculenta demonstrated that (-)-(S)- and (+)-(R)-Me2 were less potent than Mex in producing tonic block, but showed a higher use-dependent block. (-)-(S)-Me2 and (-)-(R)-Mex were also used to study the excitability of muscle fibres of myotonic ADR mice, a phenotype of a recessive form of low G(Cl) myotonia. (-)-(S)-Me2 reduced spontaneous discharges and after discharges better than (-)-(R)-Mex in agreement with the use-dependent block of sodium currents.

Antimyotonic effects of tocainide enantiomers on skeletal muscle fibers of congenitally myotonic goats.

Tocainide is effective in the symptomatic treatment of myotonic syndromes for its ability to reduce the high frequency discharges of action potentials typical of the disease, by blocking voltage-gated sodium channels. However, its use is restricted by serious side effects. In spite of its chiral structure, tocainide is clinically used as a racemic mixture. Since the optical isomers may differ in their efficacy and toxicity, the present study was aimed at evaluating the antimyotonic activity of the pure R(-) and S(+) enantiomers of tocainide, on the abnormal membrane hyperexcitability of external intercostal muscle fibers of congenitally myotonic goats. The excitability parameters were recorded in vitro by means of the standard two-microelectrode current-clamp technique before and after the addition of the compounds. The R(-) enantiomer of tocainide at concentrations as low as 10 microM potently counteracted the abnormal excitability of myotonic fibers, by increasing the threshold current, and decreasing the latency of the action potential and firing capability. Also, this concentration of R-(-) tocainide almost completely abolished the abnormal spontaneous electrical activity occurring in about 70-80% of the myotonic fiber. The S(+) enantiomer was remarkably less potent since up to 100 microM did not restore the normal excitability pattern. The results show that most of the antimyotonic activity of tocainide resides in the R(-) enantiomer suggesting that its clinical use may allow a significant reduction of the doses and possibly of the side effects.

Aging-associated down-regulation of ClC-1 expression in skeletal muscle: phenotypic-independent relation to the decrease of chloride conductance.

In order to clarify the mechanism underlying the reduction of resting membrane chloride conductance (gCl) during aging, the levels of mRNA encoding the principal skeletal muscle chloride channel, ClC-1, were measured. Total RNA samples isolated from tibialis anterior muscles of aged (24-29 months old) and adult (3-4 months old) rats were examined for ClC-1 expression using Northern blot analysis, and macroscopic gCl was recorded from extensor digitorum longus muscle fibers from each adult and aged rat in vitro using a two intracellular microelectrode technique. Although interindividual variability was observed, aged rats exhibited a parallel reduction of both gCl and ClC-1 mRNA expression as compared to adult rats. A linear correlation exists between individual values of ClC-1 mRNA and gCl. These results provide evidence that ClC-1 is the main determinant of sarcolemmal gCl and demonstrate that the decrease of gCl observed during aging is associated with a down-regulation of ClC-1 expression in muscle.

Higher content of insulin-like growth factor-I in dystrophic mdx mouse: potential role in the spontaneous regeneration through an electrophysiological investigation of muscle function.

Insulin-like growth factor-I (IGF-I) is known to promote proliferation and differentiation of muscle cells during growth and regeneration. Both these conditions are characterized by acquisition of specialized muscle functions, such as a large macroscopic chloride conductance (GCl), a parameter that is a target of growth hormone (GH)/IGF-I axis action on skeletal muscle. The present study has been aimed at evaluating the role of IGF-I in the spontaneous regeneration occurring in hind limb muscle of dystrophic mdx mouse. IGF-I levels have been measured in hind limb muscles, plasma and liver of mdx and control mice of 8-10 weeks and 5 months of age by radioimmunoassay. In parallel the biophysical and pharmacological properties of muscle chloride channels of extensor digitorum longus (EDL) muscle fibers of mice belonging to the same age-group have been measured electrophysiologically in vitro. At 8-10 weeks of age, significantly greater amounts of IGF-I were found in plasma and hind limb muscles of mdx mice with respect to controls. Such a difference was only just detectable and no longer statistically significant at 5 months of age. No differences were found in hepatic IGF-I levels at either age. The EDL muscle fibers of mdx mice at 8-10 weeks of age were characterized by higher GCl values and by a different pharmacological sensitivity to the enantiomers of 2-(p-chlorophenoxy)-propionic acid (CPP), specific chloride channel ligands, with respect to age-matched controls. However, these differences were no longer detected at 5 months of age. Our results suggest a role of IGF-I in the high regenerative potential of muscles from mdx mice and support the hypothesis that the biophysical and pharmacological properties of chloride channels of EDL muscle fibers are sensitive indices of the action of regeneration-promoting factors on muscle function.

Effects of HMG-CoA reductase inhibitors on excitation-contraction coupling of rat skeletal muscle.

3-Hydroxy-3-methyl glutaryl coenzyme A (HMG-CoA) reductase inhibitors currently used as cholesterol-lowering drugs produce side effects in patients, one of which is myopathy. In the present study we compared the effect of a 3-month chronic treatment with two different compounds, simvastatin and pravastatin, on the excitation-contraction coupling of rat skeletal muscle fibers, the mechanism which links membrane depolarization to the movements of cytosolic Ca2+ from intracellular stores. The voltage threshold for mechanical activation of extensor digitorum longus muscle fibers in response to depolarizing pulses of various durations was studied in vitro by the two intracellular microelectrode method in 'point' voltage clamp mode. Simvastatin (5-50 mg/kg) modified the mechanical threshold of striated fibers in a dose-dependent manner. The muscle fibers of rats treated with 10 mg/kg and 50 mg/kg of simvastatin needed significantly less depolarization to contract than did untreated fibers at each pulse duration, suggesting that levels of cytosolic Ca2+ were higher. Consequently, the rheobase voltage for fiber contraction was significantly shifted toward more negative potentials with respect to controls by 2.4 mV and 7.1 mV in the 10 mg/kg and 50 mg/kg simvastatin-treated animals, respectively. Pravastatin treatment at 100 mg/kg did not produce any alteration of excitation-contraction coupling since the rheobase voltage was similar to that of controls. The different physicochemical properties of the two drugs may underlie the different effect observed because lipophilic agents, such as simvastatin, have been shown to affect sterol biosynthesis in many tissues, whereas the hydrophilic pravastatin is hepato-selective.

Phosphorylation and IGF-1-mediated dephosphorylation pathways control the activity and the pharmacological properties of skeletal muscle chloride channels.

1. In the present study we tested the hypothesis that insulin-like growth factor-1 (IGF-1) modulates resting chloride conductance (G(Cl)) of rat skeletal muscle by activating a phosphatase and that the chloride channel, based on the activity of phosphorylating-dephosphorylating pathways, has different sensitivity to specific ligands, such as the enantiomers of 2-(p-chlorophenoxy) propionic acid (CPP). 2. For this purpose G(Cl) in EDL muscle isolated from adult rat was first lowered by treatment with 5 nM 4-beta-phorbol 12,13 dibutyrate (4-beta-PDB), presumably activating protein kinase C (PKC). The effects of IGF-1 and of the enantiomers of CPP on G(Cl) were then tested. 3. IGF-1 (3.3 nM) had no effect of G(Cl) on EDL muscle fibres in normal physiological solution, whereas it completely counteracted the 30% decrease of G(Cl) induced by 4-beta-PDB. No effects of IGF-1 were observed on G(Cl) lowered by the phosphatase inhibitor okadaic acid (0.25 microM). 4. Ceramide, reported to activate on okadaic acid-sensitive phosphatase, mimicked the effects of IGF-1. In fact, N-acetyl-sphingosine (2.5-5 microM), not very effective in control conditions, increased the G(Cl) lowered by the phorbol ester, but not the G(Cl) lowered by okadaic acid. 5. In the presence of 4-beta-PDB, G(Cl) was differently affected by the enantiomers of CPP. The S(-)-CPP was remarkably less potent in producing the concentration-dependent reduction of G(Cl), whereas the R(+)-CPP caused an increase of G(Cl) at all the concentrations tested. 6. In conclusion, the PKC-induced lowering of G(Cl) is counteracted by IGF-1 through an okadaic acid sensitive phosphatase, and this effect can have therapeutic relevance in situations characterized by excessive channel phosphorylation. In turn the phosphorylation state of the channel can modulate the effects and the therapeutic potential of direct channel ligands.

Chronic administration of taurine to aged rats improves the electrical and contractile properties of skeletal muscle fibers.

A reduction of resting chloride conductance (GCl) and a decrease of the voltage threshold for contraction are observed during aging in rat skeletal muscle. The above alterations are also observed in muscle of adult rat after taurine depletion. As lower levels of taurine were found by others in aged rats compared to young rats, we tested the hypothesis that a depletion of taurine may contribute to the alteration of the electrical and contractile properties we found in skeletal muscle during aging. This was accomplished by evaluating the potential benefit of a pharmacological treatment with the amino acid. To this aim 25-mo-old Wistar rats were chronically treated (2-3 mo) with taurine (1 g/kg p.o. daily) and the effects of such a treatment were evaluated in vitro on the passive and active membrane electrical properties of extensor digitorum longus muscle fibers by means of current-clamp intracellular microelectrode technique. Excitation-contraction coupling was also evaluated by measuring the voltage threshold for contraction with the intracellular microelectrode "point" voltage clamp method. In parallel muscle and blood taurine contents were determined by high-performance liquid chromatography. Taurine supplementation significantly raised taurine content in muscle toward that found in adult rats. Supplementation also significantly increased GCl vs. the adult value, in parallel the excitability characteristics (threshold current and latency) related to this parameter were ameliorated. The increase of GCl induced by taurine was accompanied by a restoration of the pharmacological sensitivity to the R(+) enantiomer of 2-(p-chlorophenoxy) propionic acid, a specific chloride channel ligand. In parallel also the protein kinase C-mediated modulation of the channel was restored; in fact the potency of 4-beta-phorbol 12, 13-dibutyrate in reducing GCl was lower in taurine-treated muscles vs. untreated aged, being rather similar to that observed in adult. The treatment also improved the mechanical threshold for contraction of striated fibers which in aged rats is shifted toward more negative potentials, moving it toward the adult values. Our results suggest that the reduction of taurine content could play a role in the alteration of electrical and contractile properties observed during aging. These findings may indicate a potential application of taurine in ensuring normal muscle function in the elderly.

Partial recovery of skeletal muscle sodium channel properties in aged rats chronically treated with growth hormone or the GH-secretagogue hexarelin.

This study was aimed at investigating the effects of chronic treatment of aged rats with growth hormone (GH, 8 weeks) or the GH-secretagogue hexarelin (4 weeks) on the biophysical modifications that voltage-gated sodium channels of skeletal muscle undergo during aging, by means of the patch-clamp technique applied to fast-twitch muscle fibers. Two phenotypes of aged-rat fibers could be discriminated on the basis of channel conductance. In the young phenotype, sodium channels present a conductance of 18 pS as in young-adult rats. In the aged phenotype, channels present a conductance of 9 pS while ensemble average currents activate and inactivate more slowly. Nevertheless, in all situations, sodium channels shared a number of biophysical properties, such as open probability, mean open time, steady-state inactivation and use-dependent inhibition. Furthermore, channel density on extrajunctional sarcolemma was higher in aged rats, a result independent of the phenotype. Chronic treatment of aged rats with either GH or hexarelin restored current kinetics but not channel conductance and density. These results confirm the specific age-related changes in sodium channel behavior and show that treatment with either GH or hexarelin has partial restorative effects. Moreover, hexarelin restored the firing capacity of fast-twitch muscle fibers, as did GH in previous studies. These findings support the possible therapeutic value of the synthetic peptide in cases of GH deficiency, as in the elderly.

The biophysical and pharmacological characteristics of skeletal muscle ATP-sensitive K+ channels are modified in K+-depleted rat, an animal model of hypokalemic periodic paralysis.

We evaluated the involvement of the sarcolemmal ATP-sensitive K+ channel in the depolarization of skeletal muscle fibers occurring in an animal model of human hypokalemic periodic paralysis, the K+-depleted rat. After 23-36 days of treatment with a K+-free diet, an hypokalemia was observed in the rats. No difference in the fasting insulinemia and glycemia was found between normokalemic and hypokalemic rats. The fibers of the hypokalemic rats were depolarized. In these fibers, the current of sarcolemmal ATP-sensitive K+ channels measured by the patch-clamp technique was abnormally reduced. Cromakalim, a K+ channel opener, enhanced the current and repolarized the fibers. At channel level, two open conductance states blocked by ATP and stimulated by cromakalim were found in the hypokalemic rats. The two states could be distinguished on the basis of their slope conductance and open probability and were never detected on muscle fibers of normokalemic rats. It is known that insulin in humans affected by hypokalemic periodic paralysis leads to fiber depolarization and provokes paralysis. We therefore examined the effects of insulin at macroscopic and single-channel level on hypokalemic rats. In normokalemic animals, insulin applied in vitro to the muscles induced a glybenclamide-sensitive hyperpolarization of the fibers and also stimulated the sarcolemmal ATP-sensitive K+ channels. In contrast, in hypokalemic rats, insulin caused a pronounced fiber depolarization and reduced the residual currents. Our data indicated that in hypokalemic rats, an abnormally low activity of ATP-sensitive K+ channel is responsible for the fiber depolarization that is aggravated by insulin.

Potential targets for skeletal muscle impairment by hypogravity: basic characterization of resting ionic conductances and mechanical threshold of rat fast- and slow-twitch muscle fibers.

Prolonged hypogravity such as during space flights affects skeletal muscle function by inducing postural changes as well as reduced muscle strength and locomotion capacity. Also in rats, space flight as well as useful models of groundbased hypogravity induce marked atrophy in the slow-twitch soleus (SOL) muscle as opposed to slight or none in the fast-twitch ones such as extensor digitorum longus (EDL). Biochemical and histological studies on hindlimb suspended animals, showed a hypogravity-induced impairment of muscle function involving the transition of slow-twitch muscle type, responsible for postural control, toward the fast-twitch phenotype by modification of excitation-contraction pattern. In slow muscles of rats, hindlimb suspension induced upregulation of the fast isoform of myosin heavy-chain and increased expression of fast Ca2+ pump mRNA and protein, which is consistent with the increased Ca(2+)-dependent ATPase activity and the speeding of muscle relaxation, typical of fast muscles. Little is known about the modifications induced by hypogravity in the sarcolemmal ion channels function, which controls the pattern of muscle excitability and contractility. The normally high resting chloride conductance, which is required for the electrical stabilization of mammalian muscle fibers, may be a target of hypogravity modifications since a pharmacological block of this parameter determines, though an increase of excitability, the transition of the fast-twitch muscle phenotype toward the slow one either in adult or in developing rats. Hypogravity also induced increased expression of dihydropyridine receptors in soleus muscle, that are normally lower than that found in the fast ones. In this study, we characterized the electrical and contractile properties of rat extensor digitorum longus (EDL) and slow-twitch soleus SOL muscles fibers at the aim to better understand the molecular mechanisms leading to fiber transition.